Tooth movement.

Critical Reviews in Oral Biology and Medicine,

2(4):411-450 (1991)

Tooth Movement Zeev Davidovitch Department of Orthodontics, The Ohio State University College of Dentistry, Columbus, Ohio ABSTRACT: This article reviews the evolution of concepts regarding the biological foundation of force-induced tooth movement. Nineteenth century hypotheses proposed two mechanisms: application of pressure and tension to the periodontal ligament (PDL), and bending of the alveolar bone. Histologic investigations in the early and middle years of the 20th century revealed that both phenomena actually occur concomitantly, and that cells, as well as extracellular components of the PDL and alveolar bone, participate in the response to applied mechanical forces, which ultimately results in remodeling activities. Experiments with isolated cells in culture demonstrated that shape distortion might lead to cellular activation, either by opening plasma membrane ion channels, or by crystallizing cytoskeletal filaments. Mechanical distortion of collagenous matrices, mineralized or non-mineralized, may, on the other hand, evoke the development of bioelectric phenomena (stress-generated potentials and streaming potentials) that are capable of stimulating cells by altering the electric charge on their membrane or their fluid envelope. In intact animals, mechanical perturbations on the order of about 1 min/d are apparently sufficient to cause profound osteogenic responses, perhaps due to matrix proteoglycan-related "strain memory". Enzymatically isolated human PDL cells respond biochemically to mechanical and chemical signals. The latter include endocrines, autocrines, and paracrines. Histochemical and immunohistochemical studies showed that during the early places of tooth movement, PDL fluids are shifted, and cells and matrix are distorted. Vasoactive neurotransmitters are released from periodontal nerve terminals, causing leukocytes to migrate out of adjacent capillaries. Cytokines and growth factors are secreted by these cells, stimulating PDL cells and alveolar bone lining cells to remodel their related matrices. This remodeling activity facilitates movement of teeth into areas in which bone had been resorbed. This emerging information suggests that in the living mammal, many cell types are involved in the biological response to applied mechanical stress to teeth, and thereby to bone. Essentially, cells of the nervous, immune, and endocrine systems become involved in the activation and response of PDL and alveolar bone cells to applied stresses. This fact implies that research in the area of the biological response to force application to teeth should be sufficiently broad to include explorations of possible associations between physical, cellular, and molecular phenomena. The goals of this investigative field should continue to expound on fundamental principles, particularly on extrapolating new findings to the clinical environment, where millions of patients are subjected annually to applications of mechanical forces to their teeth for long periods of time in an effort to improve their position in the oral cavity. Recently developed research tools such as cell culture techniques and immunologic probes, are the best hope for enhancing this development. KEY WORDS: orthodontic forces, distortion of cells and matrix,

I.

INTRODUCTORY REMARKS

Throughout their natural history, teeth move and migrate. Prior to their eruption into the oral cavity, changes in the position of tooth buds occur primarily due to the growth of dental structures, and the concomitant remodeling of neighboring tissues, i.e., alveolar bone, gingiva, and periodontal ligament (PDL), including the dental follicle. Following their emergence into the oral cavity, teeth reach a position in the dental arch,

neurotransmitters, cytokines, synergism.

dictated by the forces of the surrounding muscles of the tongue, cheeks, and lips, and by contact with teeth of the opposite jaw. During mastication, teeth can move slightly in the vertical and horizontal directions, within the constraints of the soft tissues of the PDL, and the bendability of the alveolar bone. Despite their large magnitude, masticatory forces do not alter the position of teeth, due to their short duration. However, in the presence of periodontal disease, when paradental tissues are gradually destroyed, teeth can

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migrate into new positions, where the masticatory and parafunctional forces reach equilibrium. Often, these new positions are aesthetically

undesirable. Tooth position may be deemed undesirable due to functional and aesthetic considerations, prompting patients to seek orthodontic care. In its most simplistic translation, "orthodontics" means straightening teeth. This "straightening", or movement of teeth into desirable positions, is accomplished by the application of forces to teeth, usually of small magnitude (on the order of a few grams per square centimeter of dental root surface) and long duration (usually about 2 years). Millions of people are subjected annually to orthodontic treatment worldwide, making this branch of dental care a widespread and lucrative specialty. The number of dentists in the U.S. who limit their practice to orthondontics is presently around 10,000. However, many general dentists worldwide provide orthodontic care to their patients, and all, specialists and generalists alike, base their therapeutic means upon the time-tested observation that teeth can be forced to move away from their position in the dental arch to new locations by means of applied mechanical forces. The following review focuses upon the phenomenon of tooth movement that can be brought about by the application of continuous mechanical forces to teeth. Excluded from this review are the phenomena of eruptive, pathological (periodontal disease related), and surgically induced tooth movements. Specifically, this review discusses biological aspects of force-induced tooth movement on the tissue, cellular, and molecular levels.

II. HISTORICAL PATHFINDERS The first recorded recommendation to use force for orthodontic reasons was made around the year 1 A.D. by Celsus, who suggested the application of finger pressure to teeth for alignment purposes.1 Seventeen centuries later, Fauchard was the first to publish a description and an illustration of an orthodontic appliance, which generated forces by using ligatures to tie teeth to a rigid arch.2 In the 18th century, Hunter3 provided the first biological explanation for ortho-

dontic tooth movement: "To extract an irregular tooth would answer but little purpose, if no alterations could be made in the situation of the rest; but we find that the very principle upon which teeth are made to grow irregularly is capable, if properly directed, of bringing them even again. This principle is the power which many parts (especially bones) have of moving out of the way of mechanical pressure." Two significant observations were made during the 19th century concerning the biological nature of orthodontic tooth movement. In 1815, Delabbare4 remarked that pain and swelling of paradental tissues occur following the application of orthodontic forces to teeth. In contemporary terms, Delabbare introduced the notion that inflammation is an integral part of orthodontic tooth movement. In 1888, Farrar5 hypothesized that tooth movement is due, at least in part, to bending of alveolar bone by applied forces. This notion was supported by Wolffss6 proposition in 1892 that the internal architecture of bone is dictated by the mechanical forces that act upon it. The first report on the histomorphology of tissues surrounding orthodontically treated teeth was published by Sandstedt in 1904 to 1905.7T8 That landmark experiment, which was performed in one dog, concluded that force-induced tissue changes are limited to the PDL and its alveolar bone margin. At the end of 3 weeks of treatment, Sandstedt observed new bone growth in the stretched PDL, and bone resorption in the area of PDL compression. Cell death occurred in the compressed PDL when the applied force was excessive, and the alveolar bone resorbed as a result of osteoclastic activity in adjacent marrow spaces

(undermining resorption). Six years later, Oppenheim9 reported on a histologic examination of the jaws of one juvenile baboon whose teeth had been treated by orthodontic forces for 40 d. In contrast to Sandstedt, Oppenheim saw no demarcation between the old and new alveolar bone near the moving teeth, but rather a trabecular structure that strongly suggested a complete transformation of the entire alveolar bone in that region. The bony trabeculae were all rearranged in the direction of the force. However, Oppenheim's conclusions that orthodontic forces were capable of transforming the entire alveolus were rejected by his contempor-

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aries as misinterpretations. He was also criticized for using an animal with deciduous teeth for his experiment, suggesting that the transformation he had seen was related to growth and development rather than being the outcome of applied mechanical forces. Oppenheim's "transformation" hypothesis might have supported Farrar's earlier contention that orthodontic forces bend the alveolar bone, and thus are able to stimulate all the cells in and around this bone. However, Farrar's clinical approach also was not popular, because he advocated the use of heavy forces that could indeed bend the bone. In fact, the pendulum swung furthest to the other side, when Schwarz10 recommended the use of light orthodontic forces. He defined those forces as being "not greater than the pressure in the blood capillaries" (15 to 20 mmHg, or about 20 to 26 g/cm2 of root surface). A 1957 publication by Fukada and Yasuda"l attracted wide attention. They observed that bending of bone by mechanical means evokes the generation of measurable electric potential spikes in areas of compression and tension. While this observation caused the rebirth of the field of applied exogenous electricity to bone nonunion fractures, it also precipitated the reintroduction of the concept of alveolar bone bending by orthodontic forces!2,13

III. HISTOMORPHOLOGY OF TOOTH MOVEMENT A. Observations

by Light Microscopy

pioneering work of Sandstedt and Oppenheim opened the door for comprehensive efforts to explore in detail the morphological changes in the stressed PDL. For over 4 decades, Reitan'4-'9 spearheaded this thrust with authority and confidence. The strength of his work was derived primarily from the extensive use of human material, whereby teeth that were to be extracted for orthodontic reasons were subjected to a variety of orthodontic force systems, i.e., light, heavy, continuous, intermittent, tipping, and translatory. At the end of the experimental period, the teeth were removed together with their surrounding tissues, and processed for histologic The

evaluation. Moreover, Reitan studied paradental tissues of animals subjected to orthodontic forces, particularly dogs and monkeys, exploring the effects of age, function, type of bone, force magnitude, duration, and direction, on the morphological characteristics of the tissues. He concluded that PDL cells in sites of tension proliferate, and that newly formed osteoid in these areas resorb slowly when subjected to pressure. In examining tissues from different species,20 Reitan observed that their responses varied, and attributed this variability to the differences in their structural composition, i.e., alveolar bone density, frequency and distribution of marrow spaces, and the cell and matrix constitution of the PDL. The realization that the rate of orthodontic tooth movement in humans varies and is unpredictable prompted Storey to suggest that it depends upon the magnitude of the applied force,21'22 or the presence of hormonal fluctuations, such as those associated with the menstrual cycle.23 These clinical observations motivated Storey24-27 to conduct a series of experiments in rodents in which he applied forces of different magnitudes to the maxillary incisors, causing lateral tooth movement and mid-premaxillary sutural widening. In the rabbit and rat the teeth moved faster, as the force was increased, but as in man, there seemed to be a range of force magnitudes that could be termed optimal.24 Near teeth treated with such a force, newly formed bone appeared more mature, while heavy forces were associated with the formation of a highly cellular, poorly calcified matrix. Heavy forces caused periodontal necrosis and other destructive changes in the PDL, while light forces appeared to be favorite when moving a tooth through a thin plate of bone, as apposition of bone on the labial surface seemed to lag behind the resorptive activity on the PDL side.25 In older animals, the rate of tooth movement decreased, perhaps due to reduced cellular

activity.26 Based on these observations, Storey,27 concluded that the process of tooth translation through bone consists of three different phenomena: bioelastic, bioplastic, and biodisruptive. The PDL and alveolar bone, due to their fluid-fiber composition, can be deformed elastically by external stresses, which also evoke cellular activities. When the tissue elastic limit is reached, it starts to deform plastically, with adaptive prolif413


erative and remodeling reactions. Prolonged forces that exceed the bioplastic limit result in biodisruptive deformation, with ischemia, cell death, inflammation, and repair. Clinically speaking, Storey asserted that light forces within the bioplastic range would cause slow tooth movement, while optimal forces that cause teeth to move faster are within the boundaries of the biodisruptive range.27 He concluded that within the optimum range, tooth movement is rapid, but the quality of remodeling bone is poor, increasing the potential for relapse, once external force application ceases. In contrast to Reitan, who was able to study human paradental tissues following the application of forces to teeth, Storey's histologic work was conducted solely on rodents. Storey observed that in humans teeth move at different rates when forces of various magnitudes are used, and he used rodent incisors to explore the reasons for this phenomenon. While generally Storey's observations correlated well with findings of other investigators, it is doubtful whether conclusions derived from experiments in rodents can be directly extrapolated to man, due to marked physiological differences, as pointed out by Reitan.20 Nevertheless, Storey's reports are significant, as he emphasized the development of an inflammatory process in the stressed PDL, even when light forces are being used.27 Reitan's and Storey's investigations demonstrated the complexity of the tissue reaction during tooth movement. It was no longer perceived as a simple phenomenon of applied force causing the tooth to move within the PDL, leading to tension and compression, and subsequent bone formation and resorption, but rather as a dynamic set of events that involved profound alterations in cellular functions and changes in matrix composition. Thus, histology facilitated the morphological description of changes in the dentoalveolar complex that followed the administration of orthodontic forces to teeth. While being unable to explain why alveolar bone and PDL cells are responsive to applied mechanical stresses, or how these physical entities evoke biochemical responses by the cells, the histological investigations unveiled the sites of cellular activity, and enabled other researchers to ask "why" and "how". These questions were explored by

the use of methods such as histochemistry, electron microscopy, autoradiography, and immunohistochemistry. In addition, PDL and alveolar bone, recognized as the prime targets for orthodontic forces, were obtained from animals and humans and were subjected to mechanical stresses in culture conditions, either in tissue form or as isolated cells.

B. Histochemical Changes Associated with Force-Induced Paradental Tissue

Remodeling Activities of oxidative enzymes and phosphatases were localized in the PDL of rats during force-induced tooth movement by Deguchi and Mori28 and Takimoto et al.20'30 They caused tooth movement by placing a piece of stretched rubber between the first and second molars, forcing the teeth to move in opposite directions. (This method, introduced by Waldo and Rothblatt,31 was later adopted by a number of investigators who used rats as experimental animals in studying tooth movement. It does not allow measurements of force magnitude, and the rubber pieces can traumatize gingival and periodontal tissues.) They reported on an increase in the number of osteoclasts displaying high succinic dehydrogenase activity in PDL pressure zones after 24 h. In contrast, they observed no changes in the distribution of acid phosphatase and lactate dehydrogenase in the stressed PDL. Rats were also used by Lilja et al.32 to study the distribution and activity of a number of enzymes associated with alveolar bone resorption. One maxillary molar in each rat was moved bucally by light (5 g) or heavy (36 g) forces generated by a spring attached to the incisors for either 10 h or 1, 3, 4, or 6 d. The activities of acid phosphatase increased in PDL cells in

compression zones, as well as in adjacent gingival cells and alveolar crest periosteum. Staining for acid phosphatase also increased in marrow cells and in osteocytes near the PDL pressure zone. Lactate dehydrogenase activity, a marker of vital cells, disappeared from areas of PDL pressure ("hyalinized zones"), where cells apparently died in both cases of light and heavy forces. Interestingly, prostaglandin synthetase


activity was found in alveolar bone marrow cells and in gingival cells, but not in PDL cells of the rat. Unfortunately, quantitative analysis of the data in this study was impossible, because each group consisted of only one animal. In a related experiment, Lilja et al.33 examined the PDL pressure zone of human premolars following force application for 1 to 30 d. Dilated blood vessels were seen near the hyalinized zone throughout the experiment. Intense activities of arylsulfatase and prostaglandin synthetase and moderate activity of aminopeptidase M were found in macrophages degrading the hyalinized zone. Adjacent osteoclasts displayed intense activity for succinic dehydrogenase and acid phosphatase. Again, each treatment time group consisted of one to two subjects, precluding statistical analysis of the data. The generation of osteoclasts in the compressed PDL did not escape attention, leading investigators to examine this zone in an effort to elucidate specific histochemical aspects of bone remodeling. Kurihara and Enlow34 stained rat maxillary second molar sections with ruthenium red, which demonstrates the presence of glycosaminoglycans (GAG). In areas of alveolar bone

resorption, reattachment of the PDL seemed to occur as a result of GAG secretion by fibroblasts and osteoblasts, serving to link new and old collagenous fibers. Here, too, quantitation was impossible due to inadequate sample size. In con-

trast, Martinez and Johnson35 were better able to assess the effect of orthodontic forces on alveolar bone GAG content in rats. They moved maxillary molars in groups of four rats, treated for 1, 3, or 5 d with a spring (25 g). They found that GAG staining in alveolar bone at PDL tension areas (achieved with alcian blue) increased at day 3. However, in an external control group that received an inactive spring, the GAG staining was lighter than in the untreated side of the maxilla of the rats treated with an active spring. Thus, this well-planned experiment demonstrated the need for a control group in which the orthodontic appliance remains inactive. Compression zone osteoclasts in rats were also the targets of an investigation by Noda,36 who sought to determine the effect of calcitonin on osteoclastic cytochrome c oxidase activity. First, maxillary molars were moved lingually with

a spring for periods of time ranging from 15 min to 72 h. Enzymatic activity was localized in mi-

tochondria of osteoclasts by electron microscopy. Calcitonin injections caused an early reduction in the number of mitochondria and enzymatic activity in detached osteoclasts, but this effect was abolished 72 h after calcitonin administration. These results demonstrate that locally induced bone resorption may be affected by a boneseeking hormone. The above-mentioned histochemical studies did not, for the most part, produce quantifiable data. Nonetheless, they painted a picture of enzymatically active cells, engaged in the remodeling of the stressed PDL and alveolar bone. In areas of PDL compression, oxidative enzymes and proteinases were localized in osteoclasts and in macrophages removing necrotic tissue from hyalinized zones, demonstrating heightened metabolic rates in cells involved in alveolar bone resorption and degradation of PDL matrix and dead cells. Further details on the activities of these cells, as well as those located in PDL tension sites, are derived from experiments in which electron microscopy was utilized as the investigative tool, as discussed in the following section.

C. Ultrastructural Changes in Paradental Tissues During Tooth Movement

Despite Reitan's reported observation'9 that paradental tissues of the rat are different than those of man in many respects, as, for example, morphologically and physiologically, rats remained the experimental animal of choice in transmission electron microscopic (TEM) studies of paradental tissues during tooth movement. Rygh and Selvig37 described finding degradation products of erythrocytes in enlarged blood vessels and in the extravascular space of the compressed PDL. The tension site in the PDL was studied by Ten Cate et al.,38 who later also ex-

amined stretched cranial sutures in rats.32 In both PDL and suture they observed that fibroblasts were apparently engaged in synthesizing as well as degrading collagen. Fibroblasts entering areas of matrix disruption were termed "pioneers", a term introduced earlier by Rygh40 for identifying cells entering hyalinized zones in the compressed


PDL. In the latter site, Rygh was undecided on whether these phagocytic cells were primarily macrophages or fibroblasts. In examining PDL tension zones in rats that had been subjected to force application (5 to 10 g) to a maxillary molar, Rygh41 observed distended blood vessels and mitotic PDL cells. Spaces appeared in the stretched PDL, which were filled with flocculent material. Collagen fibers were primarily oriented in the direction of tension, but many nonoriented fibrils, as well as elastic-like (oxytalan) fibers, were visible. The latter were detected earlier by Edwards42 in dogs. Teeth are attached to the alveolar bone with embedded PDL fibers (Sharpey's). Martinez and Johnson43 examined this attachment in rats using scanning electron microscopy (SEM). They reported that 5 d of tooth movement significantly reduced the diameter of the attached fibers, suggesting a reduction in the mechanical strength of the PDL. Kurihara and Enlow44 used TEM in an attempt to elucidate the nature of the attachment of PDL fibers to bone during resorptive activities. They concluded that the most prevalent type of attachment in resorptive sites is adhesive in nature. It consists of a layer of ground substance, deposited by fibroblasts on the naked surface of recently resorbed bone. Later, collagen fibrils are secreted into this layer, coalescing into fibers. In this fashion, partially released old bone matrix fibers intermingle with the newly formed PDL fibers. Since Kethcham's report in 1927 that orthodontic treatment is associated with radiographic evidence of significant dental root resorption in many moving teeth, this phenomenon was explored by numerous investigators in man and experimental animals using light microscopy, TEM, and SEM. Via SEM, Kvam45 examined human premolars that had been exposed to orthodontic forces. After 5 d of treatment, small areas of root resorption were found on the margins of the compressed PDL of all teeth, and after 25 d all treated teeth displayed resorption lacunae penetrating through the cementum into the dentin. Extensive external root resorption was observed in the teeth of patients whose palates had been expanded rapidly by Barber and Sims46 and by Langford and Sims.47 In this procedure, heavy forces were applied for about 14 d to teeth anchoring the ex-

pansion device, and the teeth were then retained in their new position for a few months to allow bone to fill the expanded mid-palatal suture. All anchor teeth exhibited root resorption lacunae, and the degree of resorption was directly related to the length of the retention period. Repair of root defects by cellular cementum was observed, but with little evidence of PDL fiber reattachment. Using light microscopy and TEM, Rygh48 investigated the PDL compression zones in rats whose molars were subjected to orthodontic forces (5, 10, or 25 g) for 2 h to 28 d. Root resorption lacunae were seen near the hyalinized zone, in close proximity to a rich PDL vascular network. Rygh suggested that root resorption might be a side effect of the cellular activity associated with the removal of the necrotic tissue of the hyalin-

ized zone, and described the removal of the cementoid layer as the elimination of the defense against resorption in an area that is strongly proresorptive. This proposed association between root resorption and PDL injury was supported recently by the results of an experiment in rats by Nakane and Kameyama.49 In that study the gingiva and PDL of a maxillary molar were injured repeatedly three times at 3-h intervals, by insertion of a 2-mm-long needle. Root resorption developed within 1 d near the traumatized, inflammatory PDL and continued through the 21 d experimental period, with concomitant repair by cementoblasts. Since mineralized tissues remodel under the influence of systemic and local factors, it was suggested that factors associated with maintenance of calcium homeostasis might regulate the activity of root resorbing cells. To test this hypothesis, tooth movement was performed by researchers in hypocalcemic rats. Goldie and King50 created calcium deficiency in adult lactating rats and applied 60 g force to a maxillary molar for 1 to 14 d. The teeth in these rats moved significantly faster than in the control animals, as their bones underwent extensive resorption. However, SEM measurements determined that the extent of root resorption was decreased in the calciumdeficient rats, suggesting that bone resorting cells are more responsive to bone seeking hormones than cells that resorb roots of teeth. In a more recent experiment, Engstrom et al.51 moved apart maxillary incisors in growing rats (30 d old) who


were being fed a calcium- and vitamin-D-deficient diet. Using light microscopy, they determined that root resorption in the moving teeth was enhanced in the hypocalcemic rats, in contrast to the finding of Goldie and King.50 This discrepancy may be a result of the age difference between the two groups of hypocalcemic rats, as well as because molars were tested in one study, while incisors were examined in the other.

D. Autoradiographic Examination of the PDL During Tooth Movement The PDL contains a mixed population of cells that can synthesize or degrade bone, cementum, and the nonmineralized PDL itself. Some of the metabolic processes associated with these activities have been investigated by the use of autoradiography. In this method, radiolabeled amino acids are injected into animals, and their timerelated location is determined by exposing tissue sections to radiation-sensitive photographic emulsions. The resulting silver-bromide salt crystals that form over radioactive sites can be localized microscopically. In tooth movementrelated studies, investigators utilized tritiated proline (3H-proline) to study the kinetics of collagen synthesis and tritiated thymidine (3H-Tdr) to study cell proliferation. Garant and his collaborators52-55 have studied extensively the process of collagen remodeling by PDL fibroblasts. They administered 3H-proline into mice and rats to determine the pattern of collagen synthesis by PDL fibroblasts by using both light microscopy and TEM. They described PDL fibroblasts as being elongated, polarized cells, with the nucleus positioned at one end and the cytoplasmic and secretory part at the other pole. Fibroblasts were found to be distributed evenly throughout the rodent PDL, and to migrate between the fibers, interacting during motion with adjacent matrix and cells. This motion appears to be facilitated by cellular microfilaments (actin and myosin), and by attachment to the matrix with glycoproteins (mostly fibronectin).56 Garant and Cho concluded that in tooth movement, PDL fibroblasts in tension sites express the phenotype

of actively migrating and matrix-secreting cells. Cells in the normal PDL proliferate and die regularly,57 but these events are markedly increased during tooth movement.58 Proliferative activities in the mechanically stressed rat PDL were studied extensively by Roberts et al.59-66 and by Yee et al.67'68 Uptake of 3H-Tdr by PDL cells in tension sites was increased significantly within 2 h of the insertion of an elastic material between the first and second maxillary molars. Most of the mitotic activity occurred near the bone and the middle of PDL, but not near the dental root. Some of the newly divided cells appeared to migrate in the direction of the alveolar bone, perhaps because they were preosteoblasts.61 Based on these observations, Roberts and his associates concluded that stretching the PDL causes G,arrested cells to enter mitosis, while G1 cells are stimulated to start synthesizing DNA. The latter cells are readily labeled by 3H-Tdr. In an effort to identify and classify PDL cells at different stages of differentiation, Roberts and Cox69 resorted to measurements of nuclear volume in these cells. This tedious method revealed that the nuclei of osteoblasts are larger than those of fibroblasts, a fact that can be used to identify committed osteoprogenitor cells. While fibroblasts were found predominantly near PDL blood vessels, osteoblastic progenitors were located further away from the vessels and closer to the bone and cementum surfaces. These reports create the impression that the PDL preosteoblastic population resides solely within the boundaries of the PDL. However, McCulloch et al.70 and McCulloch and Heersche71 have attracted attention to the finding that in mice many alveolar bone marrow spaces are directly connected through vascular channels with the PDL. Moreover, frequent injections of 3H-Tdr into large groups of mice, and subsequent autoradiographic examination of their mandibles revealed that the endosteal spaces contain many labeled progenitor cells. Thickened areas of cementum were found opposite openings of these channels in 64% of the examined specimens. Although it is presently unknown whether progenitor cells that originate in alveolar bone marrow spaces participate in the PDL response to mechanical stress, it is tempting to speculate that such an association indeed exists.


E. Tooth Movement: Visible Tissue

Changes Clinicians realized 2000 years ago (perhaps earlier) that teeth can be moved from one position in the mouth to another by being subjected to persistent mechanical forces. Merely 250 years ago, Hunter made the first attempt to explain the biological basis for this dental movement. Without having seen the involved tissues magnified by a microscope, he postulated that force-induced tooth movement was facilitated by what we may term today as bone remodeling. This notion was verified in the early years of the 20th century by Sandstedt, who described in detail the effects of mechanical force on the PDL-alveolar bone interface. His work illustrated clearly that orthodontic tooth movement is made possible by the resorption of alveolar bone near sites of compression in the PDL, while bone apposition occurs where the PDL is stretched. A controversy erupted a few years later when Oppenheim suggested that the entire alveolar bone near a moving tooth remodels, including endosteal surfaces. Most, if not all, of the investigators who were engaged in exploring this issue during the first half of the 20th century overwhelmingly supported Sandstedt's hypothesis because, clearly, the most dramatic initial histologic changes could be seen in the stressed PDL and its immediate bordering mineralized tissue surfaces, bone and dental root. Reitan, whose comprehensive histologic explorations spanned over 4 decades into the second half of this century, reported on bone remodeling in alveolar bone marrow spaces and gingival periosteum, both at a distance from the stressed PDL. These observations appeared to support Oppenheim's transformation hypothesis, and compelled Reitan to accept the century-old proposition of Farrar that orthodontic forces bend the alveolar bone. In the late 1960s Baumrind succeeded in demonstrating that such a bending effect indeed takes place, while others have measured spikes of altered electric potentials in teeth, PDL, and alveolar bone that had been subjected to orthodontic forces. The search for an optimal orthodontic force, a force that would be most efficient in moving teeth, led two investigators who examined paradental tissues microscopically to make contrast-

ing recommendations. Schwarz warned against using "heavy" forces, forces that occlude PDL capillaries and thus can damage the tissue. However, Storey recommended utilizing forces that do cause damage to the PDL, biodisruptive forces that introduce inflammation into this tissue. He also showed that within a certain range, tooth movement could be accelerated concomitantly with an elevation in force magnitude. Like Reitan, Storey associated slow tooth movement in adults with a slow rate of cellular activity. With the advent of electron microscopy, detailed information emerged in the last 2 decades on the ultrastructure of dento-alveolar tissues. Cells and matrices were studied in great detail by the use of TEM and SEM. Moreover, histochemical and autoradiographic investigations shed new light on biochemical events that occurred in the dento-alveolar tissue complex during normal

existence and while under altered states of mechanical stress. It became evident that cells that remodel the dento-alveolar complex are equipped with an elaborate system of cytoplasmic organelles that enable them to synthesize and secrete matrix components and the enzymes that participate in the degradation of this matrix. Garant and Ten Cate and their associates demonstrated that PDL fibroblasts can readily remodel the matrix, as well as migrate through it, while Jee and Roberts and their collaborators identified preosteoblasts both in the PDL, following their cell cycle kinetics, and through the stretched PDL. In the compressed PDL, Rygh, Kvam, and others have identified macrophages that seemed to remove necrotic tissue. Taken together, the above microscopic studies on both light and electronic levels have described in great detail morphological changes, and some fundamental physiological alterations that seem to occur in dento-alveolar tissues during tooth movement. However, with the exception of the proponents of the bone bending hypothesis, none of the above investigations have addressed the question of the mechanism of transduction of physical stimuli to biological reactions. Those who advocated the idea that bent bone generates electric potentials proposed that these potentials somehow stimulate cells in a mechanically stressed area by causing structural and enzymatic changes in the cellular plasma mem-


brane. However, interest in this question has increased rapidly in recent years, far beyond the boundaries of orthodontic tooth movement. Moreover, it appears increasingly evident that the regulation of cellular functions in most, if not all, tissues is under the influence of local and systemic factors that are derived from the endocrine, nervous, and immune systems. Consequently, Section IV reviews evidence in support of the hypothesis that dento-alveolar tissue remodeling during tooth movement is an outcome of cellular activities that are regulated by interactions between physical distortions and locally distributed humoral factors that act as endocrines, paracrines, or autocrines.

IV. MECHANISMS OF CELLULAR STIMULATION IN TOOTH MOVEMENT A. The Effect of Mechanical Stress on Cells Cells of all kinds are subjected at one time or another to compression, stretch, and shearing. There is growing evidence that most cells have ion channels that are potentially capable of regulating active and passive variations in cellular mechanics. According to Morris,72 these ion channels are mechanosensitive, i.e., their openstate probability depends on stress at the membrane. Such channels were postulated decades ago as a means of mechanoelectrical transduction in muscle and nerve cells. However, it now seems that most other cell types have such channel components in their membranes. These channels are ubiquitous, occurring at uniform density, on the order of 1/jim2 and in every cell.73-75 Calcium ions may enter cells in significant amounts through these channels.76-78 According to Morris and Sigurdson,79 tensions generated in patch electrodes to activate stretch sensitive channels are of the same order of magnitude as those measured in

migrating fibroblasts.80 Cell membrane tension may result from intracellular osmotic changes, contraction of cytoskeletal elements, or physical changes in the extracellular matrix. In 1985, Ingber and Folkman81 constructed three-dimensional cell models comprised of a discontinuous array of

compression-resistant struts, pulled open by connections with tension elements. The stability of such a structure depends on maintenance of tensional integrity. Based on these models, and observations of cellular behavior in vitro, these authors concluded that important functions are regulated by alterations in the integrity and composition of the extracellular matrix. Interconnections between mammalian cellular nuclei and the plasma membrane, as well as with the extracellular matrix, are through the continuous system of cytoskeletal filaments and cell surface transmembrane receptors. Physical forces, either those generated by the cytoskeleton or in the matrix, appear to be important regulators of cell and tissue growth. This interaction between force and cell function was observed to exist in skeletal myotubes,82 lymphocytes,83 arterial smooth mus-

cle cells,84 osteosarcoma cells,85 and endothelial cells.78 Ingbar and Folkman81 hypothesized that if physical stimuli can be translated into metabolic alterations through changes of intracellular structure, then mechanochemical transduction of these signals is most likely mediated by the structural linkages that join the cytoskeleton with the external milieu. In 1985, Ingber and Jamieson86 proposed that the cellular mechanism of mechanochemical stimuli is transduced into chemical information through local changes in thermodynamic parameters. In this fashion, activation energy of a reaction is produced by pressure and volume alterations, and various chemical reactions and macromolecular polymerization processes can be selectively promoted or inhibited as a result of mechanical perturbation of the cell surface. Indeed, Joshi et al.87 have been able to demonstrate that intracellular cytoskeletal polymerization can be modulated by mechanical forces applied to the cell surface in neurites. Similar changes may be caused by cell growth factor interactions. For instance, Herman and Pledger88 reported on alterations in the distribution of actin and vinculin in fibroblasts as a result of exposure to platelet-derived growth factor. Likewise, the arrangement and function of steroid hormone receptors may be very sensitive to mechanical perturbation because they appear to be associated physically with the nuclear protein matrix.89-91 Nicolini et al.92 reported that a specific in419


crease in nuclear size is necessary for S phase initiation, an observation that was reported earlier by Roberts and Cox69 to occur in PDL cells. Nuclear shape seems also to play an important role in regulating nuclear transport. For instance, Jiang and Schindler93 studied the effect of various growth factors on nuclear transport, and suggested that changes in nuclear shape may be permissive for delivery of growth factor-receptor complexes to their site of action in the nucleus. Experimenting with mammary epithelial cells, Emerman and Pitelka94 observed that cell rounding is usually associated with inhibition of cell growth and with promotion of cytodifferentiation. Thus, cells that produce specialized products usually appear round, a shape that might facilitate exposure of specific parts of their genome. In orthodontic tooth movement, such transformations in cellular shape are readily visible in mechanically stressed paradental cells (Figures 1 to 3). In unstressed PDL sites (Figure 1), alveolar bone osteoblasts appear flat, while those in areas of PDL tension (Figure 2) seem large and round. In areas of PDL compression (Figure 3), PDL fibroblasts assume a round shape. Histologic studies by Reitan'4-16 and Rygh41 have demonstrated that activated osteoblasts in PDL tension sites are engaged in producing a new bone matrix, while PDL cells in compression sites are primarily involved in enzymatic degradation of the compressed extracellular matrix.

B. The Effect of Mechanical Stress on Mineralized and Nonmineralized Connective Tissues 1. Regulation of Bone Remodeling In Vivo by Applied Stresses

Typically, orthodontic forces are applied continuously to teeth and their surrounding tissues. These forces evoke cellular activity, as has been demonstrated by investigators who examined affected tissues mircoscopically. However, it is unclear how long a force should be applied to stimulate target cells in particular areas of PDL and alveolar bone. Lanyon and his associates have addressed this issue in a series of experiments whereby controlled strains were applied to avian

FIGURE 1. "Flat" alveolar bone osteoblasts (arrows) 5-[pm horizontal section of cat maxilla, stained immunohistochemically for cGMP. Tissue near control, nonorthodontically treated canine. B, alveolar bone; P,

in a

periodontal ligament. (Magnification

x

1400.)

allowing the investigators to examine, radiographically, histologically, and histochemically, the effects of various strains on bone remodeling95-04 Their experimental model consisted of surgical removal of bone from both proximal and distal epiphyses of the ulna in turkeys and roosters, freeing the entire diaphysis from regular functional strains, leaving its nervous and vascular supplies intact.95 Mechanical loads are then introduced to this bone through stainless steel pins attached to an external loading apparatus. The operation caused removal of loadbearing by the ulna, followed by a loss of bone mass.96 This loss was prevented by 4 cycles per day of an externally applied loading regimen

bone in vivo,

(10,000 to 12,000 microstrain, 0.5 Hz), for 42

d. When the number of cycles was increased to 36 per day, bone formation increased significantly. Static (continuous) loads had no effect in


FIGURE 2. Alveolar bone osteoblasts (arrows) at PDL tension site after 14 d tipping force application to cat maxillary canine. Horizontal section, 5 ixm thick, stained immunohistochemically for cGMP. Notice the round appearance of the cells. B, alveolar bone; P, periodontal

ligament. (Magnification

x

1400.)

this preparation on bone remodeling, while similar loads applied intermittently for a total of only a few minutes daily increased bone mass substantially.97 The magnitude of the strain in the loaded bone appeared to be directly associated with the nature and degree of the remodeling response.4 Peak longitudinal strains below 0.001 were associated with bone loss, while peak strains above this level were associated with substantial enhancement of periosteal and endosteal bone formation.98 These effects were found also in birds suffering from calcium deficiency." These experiments demonstrated that bone cells in vivo are very sensitive to a small number of strain cycles daily. A maximal osteogenic response was obtained by only 72 s of load bearing. Moreover, it seemed like the creation of a static load environment is essentially ignored as an os-

FIGURE 3. Periodontal cells in PDL compression zone at the border of the necrotic "hyalinized zone" after 14 d of tipping force application to cat maxillary canine. Horizontal section, 5 ipm thick, stained immunohistochemically for cGMP. Notice the enlarged size of the cells in comparison to the thinner PDL cells seen in Figure 1. (Magnification x 1400.)

teoregulatory stimulus, suggesting that functional influence on bone architecture is derived solely from intermittent loading. 00 Lanyon101 then proposed a hypothesis to explain the mechanism by which bone adapts to functional load bearing. In his opinion, the osteocytes are the most likely

candidates to sense the distribution, rate of change, and magnitude of strain in the bone matrix. Following their recognition of a change in the strain situation, osteocytes communicate with the bone surface cells that remodel the bone. It seems like the important feature of strain in this respect is in the occurrence of an abnormally distributed strain rather than an unusually large strain. Evidence for osteocytic response to applied stress was found in the higher level of glucose-6-phosphate dehydrogenase in these cells, and a sixfold increase in the number of osteocytes 421


that have incorporated 3H-uridine into their RNA.102 Furthermore, Lanyon hypothesized that osteocytes respond not only to the transient effects of mechanical strain, but also to the persistent effect of mechanical strain on the matrix. As a candidate for such strain-sensitive matrix components, he chose proteoglycans.103 Examining bone sections in a polarizing microscope, Lanyon quantified their birefringence and observed matrix proteoglycan reorientation following short mechanical loading. These large, highly charged matrix molecules could be forced by strain to attach to cell surface receptors, or pass into the cell and attach to its cytoskeleton. Since the proteoglycan reorientation persists for 1 to 2 d, it could provide a physical basis for a "strain memory" in bone.'03 In a more recent study, Skerry et al.104 found that reversal reorientation of bone matrix proteoglycans also occurs in vitro, not only in avian bone, but in bones from rats and dogs as well. They attributed this molecular distortion to strain-generated fluid flow, usually preferentially oriented with the direction of collagen fibers. The experiments of Lanyon and his associates provide an attractive explanation of the longlasting effects of short-lived strains on bone cells. However, the exact mechanism by which proteoglycan reorientation could influence the activity of target cells in bone remains unknown. In orthodontics, functional appliances subject teeth to intermittent forces, leading to their gradual movement. In this situation, bone matrix distortion could perhaps be evoked and act in the fashion proposed by Lanyon and co-workers. In contrast, fixed-appliance orthodontics resorts to continuous force applications. In this mode of treatment, tissue effects often contain widespread damage, intimately associated with inflammatory and reparative responses. Moreover, orthodontic tooth movement is materialized by intense resorptive activity of alveolar bone, while Lanyon and associates' short-term strain application did not evoke any resorptive activity, but rather extensive formative function by bone cells. Nonetheless, the implication of bone matrix in the transduction of mechanical stimuli to physiological responses by bone cells serves to broaden our understanding of the mechanism of cell stimulation by externally applied forces. This seem-

ingly critical involvement of bone matrix in the response of bone cells to mechanical stress can explain, at least in part, Oppenheim's9 earlier observation of a transformation of the entire alveolar process during tooth movement.

2. Bioelectric Phenomena in Bone The dependency of bone tissue integrity and metabolism on mechanical stress has long been recognized. At the present time, growing evidence strongly suggests that osteoporosis can be curtailed or reversed by regular physical exercises, which subject skeletal elements to musclederived forces. 05106 Astronauts and animals that have participated in space flights or in experiments on simulated weightlessness demonstrated continuous loss of skeletal tissue due to the lack of gravitational forces.107"' These observations imply that mechanical stresses regulate the activity of skeletal cells, confirming Wolff s6 proposition that the structural architecture of bone depends on the nature of the mechanical stresses applied to it. Applying pressure and tension to chick embryo long bone rudiments in vitro, Glucksmannll2 observed in 1942 that optimal cartilaginous tissue structure was obtained only in the presence of mechanical stress. Experiments of this sort, which have persisted, demonstrated that skeletal tissues and cells can respond to applied mechanical stresses in vitro, even in the absence of other seemingly important systems, such as the nervous and vascular systems. Thus, bone loomed brightly as a self-contained tissue, whose response to mechanical stress is independent of any other tissue system. An example for this rather narrow approach, which attributed most of the control of the response of bone to mechanical stress on the bone cells themselves, can be found in the proceedings of a recent conference on functional adaptation in bone tissue. 13 Isolated bones or bone cells were presented at that conference as being fully capable of responding biochemically to applied mechanical stresses. A proposed major link in the cascade between the applied force and the biological response was stress-generated electric potentials.114,115 The concept of the inherent ability of bone to respond to applied mechanical stress was


boosted by Fukada and Yasuda's" report in 1957 that measurable electric potentials are evoked temporally in bent bone. They investigated dry specimens cut from femora of man and ox, and were able to measure direct and converse piezoelectric effects in those bones. They concluded that the piezoelectric effect appears only when a shearing force is applied to the collagen fibers of the bone lattice, causing them to slip past each other. More recently, Marino et al."6 investigated the piezoelectric characteristics of collagen films and concluded that these effects originate at the level of the tropocollagen molecule, or in molecules no larger than 50 A in diameter. Further support for the concept that collagen is the source of the piezoelectric effect came from studies on tendons17'118 and on decalcified bone. 19 Bassett and Becker120 reported that a net negative potential and a net positive potential appear, respectively, on the compression and tension sides of bone. These phenomena were recorded later by Cochran et al.'21 in a segment of a bovine mandible, and by Gillooly et al.12 and by Zengo et al. 122,123 in dog mandibles. Marino and Gross'24 recently compared piezoelectric surface charges of human bone with those of the cementum and dentin of whale teeth. They found that cementum and dentin were capable of producing only about 12% of the surface charge produced by cortical bone, and concluded that "piezoelectricity mediates alveolar (bone) remodeling". This is a typical statement, which narrows the effect of mechanical stress on bone to the generation of electric potentials by the stressed collagen. The possibility that alveolar bone is indeed bent by the application of tipping or translatory forces to teeth was first suggested by Farrar.5 This event was later confirmed by Baumrind13 in rats and by Grimm125 in humans. It led DeAngelis'26 to propose that the alteration of the electric environment within the stressed alveolar bone may regulate differentiation of bone progenitor cells. A mechanism by which these potentials may reach the surface of bone cells was suggested by Pollack et al. 15 According to this concept, bone is surrounded by an electric double layer in which electric charges flow in accordance with a stress-related fluid flow. This stress-generated potential may affect the charge of cell membranes, as well as that of macromolecules

in their vicinity. Borgens,127 examining intact and damaged mouse bones, detected endogenous ionic currents that he attributed to streaming potentials, rather than to piezoelectricity, due to the long (up to 30 min) current decay period. He suggested that the source of current in mechanically stressed bone is cells rather than matrix. Otter et al.,128 studying dry and wet specimens of bovine tibia, concluded that while in the dry state the current is primarily piezoelectric, in wet bone the dominant mechanism is streaming potentials.

Bioelectric measurements in alveolar bone'22,'23 have demonstrated that the com-

pressed (concave) side of the orthodontically treated bone is electronegative with respect to the tension (convex) side, suggesting that negative potentials during bone bending can generate bone deposition, while positive potentials are responsible for bone resorption. However, Borgens' experiments in fracture sites'27 failed to find such a correlation. Rather, his measurements showed that current enters the lesion, where its dispersion

(i.e., its pathway and density) remains unknown due to the complexity of the distribution of mineralized and nonmineralized matrices. Whichever is the source of electric potentials in bone, it seems that these endogenous currents are involved in bone repair, remodeling, and perhaps growth. This conclusion led numerous investigators129-"33 to apply weak currents to nonunion bone fractures in an effort to facilitate healing. Clinical successes in orthopedics prompted orthodontists to combine orthodontic force application with administration of weak electric currents to jaw tissues in an effort to determine whether a synergistic effect on the rate of tooth movement would be achieved. In the first experiment of this kind, Beeson et al. 134 implanted electrodes in cat mandibles and applied a 10-1pA direct current constantly for 5 weeks. No significant differences between electrically treated and control animals were found, regardless of whether the cathode or the anode were placed in the vicinity of the moving tooth. This absence of an effect seems to have stemmed from the placement of the electrodes near the apex of the moving teeth, rather than near the alveolar crest where most of the force-induced bone remodeling occurs when teeth are tipped. Different results were obtained by Davidovitch et al. ,135 136 who applied 423


a current of 20 ILA to gingiva near orthodontically tipped maxillary canines in young adult cats. Significant acceleration of the rate of tooth movement was observed after 7 and 14 d of combined application of force and electricity. This potentiating effect was explained by the placement of the anode very close to the site of PDL compression, where alveolar bone resorption occurs, while the cathode was placed in close proximity to the site of PDL tension, where new bone is deposited.

3. Biochemical Events in Cells Affected by Mechanical Forces In Vitro The ability to maintain cells and organs in culture permitted investigators to subject them to mechanical stress and to monitor their response to either tensile or compressive forces. These experiments targeted specific cellular functions, such as proliferation, synthesis, and secretion of extracellular matrix components, or the enzymes that degrade them. The rationale for these experiments is derived from the assumption that in vitro conditions facilitate the exposure of sensitive target cells to a specific stimulus, in the absence of any other factors that might exist in vivo, which would mask or "confound" the response that is observed in vitro. As discussed below, the situation in the intact mammalian organism differs from that of the culture system in that bone cells in the living animal may become subjected to mechanical stresses simultaneously with signal molecules derived from neighboring endothelial cells, fibroblasts, or migratory leukocytes.These molecules may amplify or diminish the effect of mechanical stress on the shape and cytoskeletal structure of the bone cells. Thus, for an in-depth understanding of the nature of the response of cells to mechanical stress it is essential to study combinations of these interactions, a design that is rarely performed for in vitro

investigations. Among the main investigative yardsticks that were utilized to explore the mechanism of activation of cells by mechanical forces were cyclic nucleotides, prostaglandins, DNA, phosphatases, and metalloproteinases. Adenosine 3',5'monophosphate (cyclic AMP, or cAMP) and guanosine 3',5'-monophosphate (cyclic GMP,

or cGMP) have been identified as mediators of the effects of external stimuli on bone cells in vivo'37-'39 and in vitro.'40'141 Fluctuations in the levels of these substances have been found to occur in bone treated by parathyroid hormone (PTH),120 calcitonin,138 and vitamin D3,139 as well as electric currents'40 and mechanical force.'41 Prostaglandins, particularly those of the E series, have been associated with bone remodeling activities resulting from malignancies,142 gingival inflammation,143 rheumatic joint disease,'44 and fracture.'45 Thus, fluctuations in the levels of cAMP and cGMP in mechanically stressed cells and PGE2 in their culture media were used frequently as indicators of cellular responsiveness. In 1975, Rodan et al.141 applied compressive forces to chick cartilaginous bone rudiments and observed a reduction in the levels of cAMP and cGMP in these cells within 15 min. Uchida et al.146 stretched rat costochondral chondrocytes from 1 min to 24 h, and reported an initial increase in cAMP content at 3 to 5 min, with a decline to control levels shortly thereafter. The incorporation of 35S into GAG by the stretched cells increased significantly, but their rate of DNA synthesis was not altered. Interestingly, when stretched chondrocytes were incubated in the presence of calcitonin, their cAMP levels at 24 h were much higher than those of control cells or cells treated with PTH. In 1980, Somjen et al.'47 stretched rat embryonic calvarial cells for 1 to 60 min. They observed sharp increases in cAMP and PGE2 levels in cells and media, respectively, with peaks at 15 to 20 min and subsequent declines. Both cAMP and PGE2 levels failed to increase when bone cells were stressed in the presence of indomethacin, a potent inhibitor of prostaglandin synthesis. More recently, Shen et al.'48 exposed osteoblasts cultured from rat fetal calvaria to different concentrations of PGE2, and within 10 min observed a transient change in shape from an epithelioid to stellate, with markedly increased numbers of gap junctions. Rat calvarial cells were also used by Hasegawa et al.149 in an effort to determine the effect of stretching on DNA and matrix component synthesis. Continuous or intermittent stretching for 2 h increased both the number of DNA synthes-


izing cells (by 64%) and the production of osteonectin-like molecules. These results suggest that bone cells respond to mechanical stress by increasing their numbers and by rearranging their contacts to neighboring structures. Similar effects on DNA synthesis were obtained with avian calvarial osteoblasts that were stretched for up to 5 d by Buckley et al. 50 In addition, the stretched cells were uniformly aligned perpendicular to the direction of the strain field. Compressive forces were applied to bone cells in a number of studies, usually by compressing the gas phase above the culture medium. In this fashion, Klein-Nulend et al.'51 applied intermittent pressure to mouse calvaria for 5 d. This treatment increased alkaline phosphatase activity and 45Ca uptake by the calvaria, while resorptive activities decreased. The net result was a 16% increase in the calvarial mineral content. Similar effects were observed by these investigators following the application of intermittent compressive forces (132 g/cm2, 0.3 Hz) to fetal mouse metatarsal bone rudiments.'52 They concluded that compression inhibits the migration and activity of osteoclasts and their precursors. Heavier, continuous compressive forces (3 atm) were applied by Ozawa et al.'53 to osteoblast-like cells, resulting in suppression of osteoblastic activities and marked enhancement of PGE2 production. Unquestionably, the main arena of tissue remodeling during tooth movement is the PDL. It is the prime target of tooth-moving mechanical forces, and has been the subject of numerous investigations aimed at elucidating details of the biological response of its cells to applied mechanical stress. Duncan et al.154 applied mechanical forces to mouse molars in vivo as well as in vitro. After 3 to 5 d in culture, large amounts of PGE2 and type II collagen were synthesized by the ligaments. A substitute model for the PDL was introduced by Meikle et al.,155 who used a spring to apply tensile stress to rabbit calvarial sutures in vitro. They reported an increase in the tissue levels of collagenase and a reduction in

the level of tissue inhibitors of metalloproteinases (TIMP) in these sutures. Fibroblasts isolated from chick embryos were grown and stretched on nylon meshes by Curtis and Seehar.'56 Intermittent stretching at 0.25 to 1.0 Hz caused significant increases in mitotic

frequency and in the proportion of cells in S phase. It was concluded that stress speeds up the mitotic cycle of fibroblasts, rather than switching cells from G, to S. However, Norton et al.157 reported recently that tensile forces failed to change the cytoskeletal configuration in PDL fibroblasts (as determined by immunofluorescence of tubulin, vimentin, and actin), suggesting that these cells are not responsive to tensile forces per se, but rather to injurious effects resulting from these forces. Human gingival fibroblasts were stretched by Ngan et al.,158 who reported that a 5% increase in cellular surface area for 5 min to 2 h caused significant elevations in the levels of cAMP and PGE2 in the cells and their media, respectively. In a related study, Ngan et al.'59 reported recently on similar effects in stretched human PDL fibroblasts. Despite the major role played by PDL fibroblasts in tooth movement, a surprisingly small number of investigations on the response of these cells to mechanical forces in vitro have been performed to date. However, the relative ease of obtaining these cells from extracted healthy human teeth,l60 and the availability of a number of mechanical systems that can apply various modes of compressive and tensile stresses to cells in culture promise to facilitate new experiments in the near future. However, since the PDL fibroblast-like cell population is heterogeneous, consisting of cells with differing phenotypes, and, as the PDL also includes numerous precursors of osteoblasts as well as epithelial and endothelial cells, strict identification and phenotyping of these cells will be required so that the data may be interpreted more meaningfully. Of great curiosity and perhaps importance is the possible role of the epithelial rests of Malassez in tooth movement. These clusters of epithelial cells, left behind during the growth of the dental root, are distributed throughout the PDL in close proximity to the root surface cementum layer. Their functional role continues to be an enigma. Reitan'7 noted that these epithelial clusters are eliminated from necrotic areas of compressed PDL and do not regenerate. He speculated that these cells may play a protective role in preventing force-induced root resorption. Brunette et al.'61 cultured monkey epithelial cells derived from rests of Malassez, together with 425


PDL fibroblasts. They observed no junctional structures between epithelial cells and fibroblasts, but in some cases islands of epithelial cells were sandwiched between two layers of fibroblasts. Later, Brunette et al.'62 detected large quantities of PGE and PGF in media in which epithelial cells derived from porcine rests of Malassez had been incubated. They proposed that these prostaglandins might participate in the regulation of alveolar bone remodeling, either by affecting bone cells directly or indirectly by interacting with PDL fibroblasts and endothelial cells. Support for the concept of interaction between PDL fibroblasts, epithelial, and endothelial cells was provided by Merrilees et al. 163 They found that the GAG synthesized in vitro by PDL fibroblasts is predominantly chondroitin sulfate, whereas epithelial cells produced primarily hyaluronic acid. However, endothelial cells or their conditioned media, when cocultured with fibroblasts, stimulated increased GAG synthesis, particularly hyaluronic acid. The question of whether epithelial cells from the rests of Malassez can produce factors that enhance bone resorption was addressed by Birek et al.'64 They cultured epithelial cells or their conditioned media with mouse calvaria for 4 d, causing significant increases in calcium release from the bones. Indomethacin inhibited this resorptive effect only partially, suggesting that factors other than prostaglandins are synthesized by the epithelial cells, which may account for the osteolytic effects of the epithelial cells. The fact that epithelial cells from the rests of Malassez respond in vitro to 65tensile forces was demonstrated by Brunette. In his experiment the number of 3H-Trd labeled cells doubled after 2 h of stretching. Moreover, stretched cells had a higher volume of filamentous structures and more desmosomes per unit length of cell membrane than unstretched cells. The above review indicates that bone cells, PDL fibroblasts, and epithelial cells from the rests of Malassez respond readily to applied mechanical stresses. These cellular responses include identifiable biochemical events that span the entire cellular domain. The plasma membrane, the cytoplasmic organelles, the filamentous skeleton, and the nucleus all seem to participate in the physical-to-chemical transduction process. How-

ever, one of the main foci of attention in this

investigative field has been the role of prostaglandins in this process. The following section briefly reviews this issue. 4. Prostaglandins and Force-Induced Bone Remodeling Since the report by Klein and Raisz'66 in 1970 that prostaglandins stimulate bone resorption in tissue culture, numerous publications have implicated prostaglandins, particularly of the E series, in the response of bone cells to chemical and mechanical stimuli. Out of the plethora of investigations emerged a hypothesis, introduced by Rodan and Martin,'67 which proposed that osteoblasts regulate the resorptive activities of osteoclasts. This hypothesis was based upon the findings that osteoblasts carry receptors to all the hormones involved in the maintenance of calcium homeostasis, such as PTH, calcitonin, and vitamin D3. Osteoblasts respond to these endocrine molecules, as well as to locally produced agents such as growth factors, with elevations in cAMP contents and prostaglandin synthesis. Thus, prostaglandins could serve as a stimulatory link or a coupling factor between osteoblasts and osteoclasts. Applying tensile forces to cells derived from mouse embryo calvaria, Binderman et al.168 stimulated the production of PGE2 and cAMP by these cells. This effect was abolished by agents that bind to membrane phospholipids (gentamicin and antiphospholipid antibodies), and thus reduce their availability for enzymatic changes. They concluded that mechanical forces exert their effect on bone cells by the following chain of events: activation of phospholipase A2, release of arachidonic acid, increased PGE synthesis, and elevated cAMP production. Taken together, these observations assign to PGE2 a central role in the regulation of force-induced bone cell activation. This is clearly an oversimplified concept that may apply to cell culture conditions, in which many factors found in the intact, living mammalian organism are absent. PGE was localized immunohistochemically in the cat PDL by Davidovitch et al.169'170 The application of tipping forces to cat canines for periods of time ranging from 1 h to 14 d caused a significant increase in


the staining for PGE in PDL and alveolar bone cells at sites of both tension and compression, suggesting that PGE may indeed be involved in the response of PDL and bone cells to mechanical stress. However, as discussed below, other local factors with bone cell stimulation capabilities have been localized recently in the mechanically stressed PDL, suggesting that prostaglandins may be only one agent in a battery of factors that regulate the response of cells to force. The great emphasis in the literature on the role of prostaglandins in the response of bone cells to applied force prompted Yamasaki and his associates to perform a series of experiments on rats, monkeys, and humans, where PGE was injected into the gingiva near orthodontically treated teeth.171-174 In rats,171 PGE, or PGE2 administration increased the number of osteoclasts in mechanically stressed alveolar bone within 12 h. In monkeys,'72 the rate of tooth movement doubled during 18 d of treatment. The drawback of this experiment was the sample size, which consisted of two subjects. Similar results were obtained in a study on human patients. 173174 Here, PGE, was injected monthly for 5 months into the gingiva near orthodontically treated canines, resulting in doubling of the rate of tooth movement. In explaining why they chose PGE administration as an adjunct to orthodontic tooth movement, Yamasaki et al. stated that the rationale for the decision had been the reported evidence, primarily from tissue-cell culture experiments, that implicated PGE2 in bone resorption. However, PGE2 has been shown to enhance metaphyseal bone growth in young rats,175 and PGE1 infusion for 3 weeks enhanced alveolar bone growth in beagle dogs.'76 Thus, in vivo, PGE may regulate bone resorption and formation. To investigate this dual role of PGE in bone as a stimulator of both resorption and formation, Nefussi and Baron'77 cultured 45Ca-labeled rat fetal long bones with PGE2 for 4 d. This treatment caused enhanced periosteal osteoblastic activity (in terms of percentage of osteoblastic surfaces), but increased osteoclastic resorption in medullary cavities. Thus, the effect of PGE on bone cells may differ, depending on their location. Pursuing this avenue further, Lee178 moved teeth in rats by inserting an elastic band between the first and second maxillary molars, according

to the method of Waldo and Rothblatt,31 for periods of 6 h to 5 d. In addition, the animals received either local gingival injections of PGE1 twice daily, or a constant systemic administration by a mini-osmotic pump. In both cases the number of alveolar bone osteoclastic lacunae in PDL pressure sites was increased significantly, compared to non-PGEl-treated animals, but the effect was more pronounced in the animals receiving PGE, systemically. This finding raises the possibility of administering PGE, or PGE2 systemically during orthodontic treatment in an effort to enhance the rate of tooth movement. However, side effects of such a treatment must not be ignored. These effects may include diarrhea, vomiting, corneal congestion, and phlebitis. Another feature of prostaglandins related to bone remodeling has come to the foreground in recent years. Prostaglandins have long been known as being potent mediators of the inflammatory process in many tissues, including cartilage and bone. This fact led to the widespread use of nonsteroidal anti-inflammatory drugs in combatting rheumatoid arthritis'79'180 and jawbone destruction due to endodontic lesions'8' and periodontal disease.182-'84 If tooth-moving forces indeed cause an inflammatory reaction in the PDL, then it should be expected that not only would prostaglandins be found there, but other inflammatory mediators as well. The following sections examine this issue.

V. REGULATION OF TOOTH MOVEMENT BY INFLAMMATORY MEDIATORS A. The Cells and Fluids of the Periodontal Ligament The PDL is a soft tissue envelope separating the tooth from the alveolar bone. Like all other connective tissues, it is comprised of cells and extracellular matrix, which consists of collagen and ground substance.'85 It contains an intricate network of blood vessels and nerve endings, and is very cellular. The majority of the PDL cells are fibroblasts, but some of these fibroblast-like cells are actually osteoprogenitor cells.65 Osteoblasts, either active or in the form of lining cells, occupy the alveolar bone surface bordering the 427


PDL, while cementoblasts cover the dental root surface that interfaces with the PDL. Clusters of epithelial cells, the rests of Malassez, are spread in the PDL in the vicinity of the root surface, while capillaries are usually more numerous in the center of the ligament and in the zone closer to the alveolar bone. Cells migrating out of these capillaries, such as lymphocytes, macrophages, and mast cells, can be observed throughout the PDL. Cells may also migrate into the PDL from neighboring marrow spaces in the alveolar bone.70,71 The PDL contains an elaborate network of neural filaments'86 that arise from the trigeminal nerve and send neural bundles through the apical alveolar bone in a coronal direction to the gingiva and PDL. Myelinated and unmyelinated fibers are found in the PDL, some terminating as "free" nerve endings, mostly in the inner part of the PDL, while others terminate as knob-like enlargements or as coiled nerve endings. Unmyelinated fibers usually follow PDL blood vessels and may have a vasomotor function. In this capacity, PDL nerve fibers may release, when stressed mechanically, vasoactive peptides that regulate movement of leukocytes out of

capillaries.

Nerve impulses resulting from tooth movement can be detected in afferent fibers. These impulses originate primarily in the PDL and not in the dental pulp, as shown in experiments where the pulp had been removed.'87 Mechanical stimulation activates PDL fibers that are associated with large-sized myelinated fibers. 188 The smaller C fibers also react to mechanical forces, but with a larger magnitude or longer duration.189 In forceinduced tooth movement, the PDL nerve fibers perform two main functions: transmission of nociceptive impulses centrally and release of neuropeptides peripherally. The latter (discussed below) may have an important role in regulating the local inflammatory response, primarily by interacting with cells of the vascular system. Examining the PDL in mouse molars, Freezer and Sims'90 observed that 88% of the blood vessels consisted of venules and 12% were capillaries. Gould et al. 91 and McCulloch and Melcher'92 found that 75 to 80% of the blood vessels were positioned in the bony portion of the PDL. To test the effect of orthodontic forces

(1 to 7 d) on the PDL vascular bed, Khouw and Goldhaber'93 perfused dogs and monkeys with a colloidal suspension of carbon particles. They reported on dilation of vessels in both areas of PDL tension and compression, in close proximity to sites of alveolar bone apposition and resorp-

tion, respectively. Examination by TEM of se-

vere PDL compression sites in rats94 revealed stasis and erythrocytic breakdown, with disintegration of vessel walls. However, this initial response was followed by a repair process, typified by an invasion of the hyalinized zone by a front of cellular and vascular elements. A markedly increased PDL vascularity was also detected in the inflamed PDL of beagles by Jeffcoat et al.195 using an angiographic method. In addition to providing the PDL with a variety of leukocytes, the vascular system also contributes to its fluid composition. Bien'96 thoroughly analyzed the dynamics of PDL fluid in relation to tooth movement, and identified three sources of fluid in the PDL: cellular, vascular, and interstitial. The latter is localized in the ground substance and acts as a thixotropic gel, which is jelly-like when not in motion, and flows quite easily under pressure. When subjected to a steady force, this fluid flows within the PDL out of areas of compression and into areas of tension. This fluid flow, which starts as soon as the force is applied to a tooth and is maintained over extended periods of time, is apparently a crucial step in the physicochemical behavior of the PDL. This fluid motion and rearrangement signifies the onset and progress of distortion of PDL cells and fibers. This distortion of PDL, which is seen microscopically as widening in areas of tension and narrowing in sites of compression, may result in the release of vasoactive neuropeptides, appearance of stress-generated potentials, and alterations of cellular shape. Storey27 observed vasodilation and migration of carbon particles out of capillaries in stressed PDL within 20 min of the application of an orthodontic force to guinea pig incisors. Thus, orthodontic forces seem to evoke an early response in the stressed PDL, which encompasses fluids, matrix fibers and ground substance, and cells. The chain of events above-reported led us to propose the following scheme to describe the initial effects of force on paradental tissues (Figure


4): (1) movement of fluids within the PDL; (2) gradual distortion of the PDL; (3) generation of streaming potentials; (4) alteration of cellular shape and ion channel permeability; (5) neuropeptide release from nerve endings; (6) capillary vasodilation and migration of leukocytes into extravascular areas; and (7) bending of the alveolar bone and generation of piezoelectric spikes. The outcome of such events is the introduction into the stressed PDL of agents derived from cells of the nervous and immune systems. In addition, products of the endocrine system are routinely delivered to the PDL through the circulation. Thus, on the biochemical level, mechanical forces can result in the simultaneous exposure of PDL cells to signals from the nervous, immune, and endocrine systems, leading to intricate and fascinating interactions and cellular responses.

B. Interactions between Cells and Products of the Nervous, Immune, and Endocrine Systems Recent attempts to understand the mode of regulation of cellular activities in various tissues have resulted in a rapidly growing volume of evidence in support of the contention that interactions between cells of the nervous, immune, and endocrine systems are pivotal parts of this mechanism. In our own research we have focused on the possibility of such interactions in the stressed PDL between neurotransmitters, cytokines, and hormones. The neurotransmitters we targeted are substance P (SP), vasoactive intestinal peptide (VIP), calcitonin gene-related peptide (CGRP), and methionine enkephalin (ME). The cytokines we chose are interleukin-lao and -

ORTHODONTIC FORCE

Movement of PDL fluids

Generation of streaming potentials that affect PDL

Gradual Distortion of PDL matrix and cells

1~~~~~~~\\^~

~and alveolar bone cells

|

Alteration of cellular

Neuropeptide release from

shape, cytoskeletal configuration, and ion channel permeability

PDL afferent nerve endings

Bending of the alveolar

Capillary vasodilation, migration of leukocytes into

bone

Piezoelectric effects

extravascular areas

Synthesis and release of cytokines, growth factors, PG's

FIGURE 4. Initial effects of orthodontic forces on

paradental tissues.


113 (IL-la and IL- 13), interleukin-2 (IL-2), tumor necrosis factor-a (TNF-a), and gamma interferon (IFN--y). The reason for choosing these particular molecules was the existing evidence implicating them in bone remodeling. Incubation of human blood monocytes with SP by Lotz et al.197 induced the release of IL-1, TNF-a, and IL-6 by these cells, demonstrating recognition of neurotransmitters by immune cells. Similar effects were seen in B lymphocytes that were stimulated to differentiate by SP,198 and in neutrophils that were stimulated by SP to enhance their oxidative metabolism. 199 ME had a biphasic effect on the proliferation of peripheral blood monocytes200 and stimulated 02 release by polymorphonuclear cells.201 Cells of the immune system were found to synthesize neurotransmitter-like molecules. Substance P was extracted from mouse liver granulomas by Weinstock et al.,202 203 and its messenger RNA (mRNA) was localized to the granuloma eosinophils by in situ hybridization. Roth et al.24 reported that the opioid precursor proenkephalin was secreted by activated T helper cells. Cells of the nervous system were reported to be reactive to products of immune cells.205 For instance, incubation of mouse anterior pituitary cells with IL-1 induced protein phosphorylation, but without cAMP elevations, which appears to be an early signal for the secretion of 13-endorphin. In another experiment, Fagarasan and Axelrod206 treated pituitary cells with IL-I in the presence of norepinephrine or isoproterenol, causing an additive effect on 3-endorphin secretion. Su et al.207 found identical steroid receptors in guinea pig brain and spleen, postulating that steroids can, in this fashion, alter the immune function and cause psychological changes.

C. Neuropeptides and Mineralized Tissues

SP, a neuropeptide present in parts of the CNS and PNS, such as C- type sensory fibers and autonomic afferents and efferents, is released from nerve endings in response to various stimuli. Its effects in peripheral tissues include vasodilation and an increase in capillary permeability. These effects may contribute to the plasma

extravasation and increased local blood flow that accompany inflammation. Moreover, SP can stimulate histamine release from mast cells at sites of injury and inflammation. These biological capabilities turned SP into a suspected prime contributor to the pathogenesis of rheumatoid arthritis. In 1984, Levine et al.208 caused adjuvant arthritis in rats, and observed that the severity of the disease was pronounced in joints that were densely innervated by SP-containing afferent neurons. Injection of SP into joints increased the severity of arthritis. Lotz et al.209 incubated synoviocytes from human arthritic joints with SP, causing increases in PGE2 and collagenase release. Yokoyama and Fujimoto210 demonstrated that lymphocytes from rheumatoid arthritis patients could be activated by SP to present high levels of HLA markers. Monocytes of these patients, when treated with SP, released high amounts of oxygen intermediates. A significant observation was made by O'Bryne et al.,211 who injected IL-la into rabbit knees and measured SP and PGE2 in the joint fluid at 4, 24, and 48 h. By 4 h, SP was increased; it further increased by 24 h and remained elevated at 48 h. PGE2 levels were highest at 4 h and remained elevated at 48 h. These results highlight the intimate association, at sites of inflammation, between cytokines, neurotransmitters, and prostaglandins. The neurogenic component of joint inflammation was demonstrated by Lam and Ferrell, who in one experiment212 have inhibited carrageenan-induced knee joint inflammation in rats by denervation, while in the other,213 abolished it by an intraarticular injection of capsaicin, a SP releaser and inhibitor. Immunolocalization of SP in dental tissues was first reported by Olgart et al.,214'215 who observed its presence in the feline dental pulp. In a more recent series of articles, Wakisaka et al.216-2'8 reported on the distribution and origin of SP in the rat molar pulp and PDL. They observed SP-containing nerves along blood vessels, primarily in the middle and apical regions of the PDL. During orthodontic tooth movement in cats, intense immunohistochemical staining for SP in PDL tension sites was seen by Davidovitch et al.219 within 1 h of treatment. Furthermore, administration of SP to human PDL fibroblasts in vitro significantly increased the


concentration of cAMP in the cells and PGE2 in the medium within 1 min.219 Another neurotransmitter that has been implicated recently as a stimulator of bone resorption is VIP, a 28-amino acid residual peptide, which was originally extracted from porcine duodenum.220 In vitro studies have demonstrated that VIP stimulates bone resorption dramatically, and that this activity is not mediated by PGE2.221 Moreover, this effect of VIP involves an 8- to 13-fold increase in bone cAMP levels.221 Binding studies222 revealed high-affinity receptors for VIP on human osteosarcoma cells. Hohmann et al.223 localized VIP in nerve fibers in the periosteum of porcine rib, tibia, and vertebra. Here, VIP was traced to sympathetic postganglionic neurons. Herness224 localized it in mouse PDL, mainly in the apical part around the blood vessels. During orthodontic tooth movement in cats,22 intense staining for VIP was localized in the compressed PDL near sites of bone resorption and in the pulp of moving teeth. The recent discovery of CGRP by Rosenfeld et al.226 raised the interest of investigators of neural control of bone remodeling. This neurotransmitter was localized in small to medium diameter sensory ganglion neurons and appeared to coexist with SP.226 In the cat, local intraarterial infusion of SP or CGRP caused a concentration-dependent increase in nasal blood flow.227 The widespread distribution of this 37-amino acid peptide has been demonstrated in cardiovascular tissues of several species, as well as in perivascular nerves in the rat mesenteric artery.22 In arthritic patients, Larsson et al.229 detected CGRP-like immunoreactivity in the knee synovial fluid; however, similar amounts of CGRP were found in synovial fluids of control patients. Recently, Silverman and Kruger230 localized CGRP in many rat orofacial sensory structures, while Wakisaka et al.231 studied its distribution in the feline dental pulp. In the latter tissue CGRP-like immunoreactivity was observed in nerves along blood vessels as well as in the subodontoblastic zone, while dentinal injury in rat molars evoked sprouting of CGRP-containing nerve fibers.232 CGRP was also localized in the rat PDL233 and mandibular periosteum.234 In the cat dental pulp, intraarterial infusion of SP and CGRP produced vasodilation, as measured by both laser Doppler

and 125I clearance.235 The effect of CGRP was ten times larger when given after SP than before it. Regarding an association between CGRP and acute inflammation, Takahashi et al.236 found CGRP-containing nerves in contact with dynorphin-containing dendrites in acutely inflamed rat hindpaw. In relation to tooth movement, Kvinnsland and Kvinnsland237 localized CGRP in the pulp and PDL of rats receiving orthodontic forces to maxillary molars for 5 d. In unstressed teeth, CGRP immunoreactivity was localized primarily in pulp and PDL nerves surrounding blood vessels. In moving teeth, the number of CGRP-containing nerves in both pulp and PDL increased, and their staining intensified, particularly in PDL tension sites. In these areas, dark "spots" were seen, which were probably fibroblasts that have bound CGRP released from stressed sensory nerve endings. Such a pattern of cellular staining for CGRP was observed in the pulp and PDL of moving teeth in cats by Okamoto et al.238 in the author's laboratory. In this group of 28 animals that had been treated by a translatory force ap-

flowmetry

plication to one maxillary canine for 1 h and/or 2, 7, or 28 d, CGRP immunoreactivity in the

PDL intensified within 1 h in tension sites, but was particularly intense in compression areas at day 28, in PDL cells located at the edge of the hyalinized zone and near osteoclasts (Figures 5 to 8). In the pulp of nonmechanically stressed teeth, staining for CGRP was widespread and distinct, localized in perivascular fibers and in numerous fibroblasts (Figure 9). Force application to a tooth did not seem to alter the CGRP immunoreactivity in dental pulp cells (Figure 10). While SP appears to be intimately involved in the transmission of painful sensations in the primary afferent system, enkephalins are related to morphine- and stimulation-produced analgesia. Both of these neuropeptides have been shown immunohistochemically to have a similar distribution in many regions of the rat CNS.239 This relationship between SP and ME could be of interest in investigating the regulation mechanism of bone remodeling, since Jessell and Iversen240 have reported that opiate analgesics inhibit SP release in the rat trigeminal nucleus. One group recently focused on this issue.241-243 In one experiment241 they measured ME levels in dental 431


FIGURE 5. Immunohistochemical localization of CGRP in unstressed cat PDL. Notice dark staining of nerve fiber, but light staining of PDL cells (arrows). (Magnification x 1400.)

FIGURE 6. Localization of CGRP in PDL tension site after 1 h of application of a translatory force to a cat maxillary canine. Notice dark staining over cells' periphery and cytoplasm

(arrows). (Magnification

x

1400.)


FIGURE 7. Dark staining for CGRP of cells in PDL tension zone after 28 d of translatory force application to cat maxillary canine (arrows). (Magnification x 1400.)

FIGURE 8. Cells in compressed PDL at edge of hyalinized zone, stained very darkly for CGRP, after 28 d of translatory force application to cat maxillary canine. (Magnification x 1400.)


FIGURE 9. Nerve fiber in unstressed cat canine pulp, intensely stained for CGRP, approaching blood vessel wall (B). (Magnification x 1400.)

FIGURE 10. Immunohistochemical localization of CGRP in pulp of cat canine subjected to translator force for 28 d. Staining deposits are seen in nerve fiber approaching blood vessel (B), as well as in adjacent cells. (Magnification x 1400.)


pulps of human teeth that had been moved by an orthodontic spring for a few hours prior to extraction. They found drastic declines in the concentrations of ME in the moved teeth, perhaps correlated with the early development of painful

sensations in orthodontic treatment. In another study242 ME was extracted from pulpless, decalcified human third molars, suggesting that ME is present in dentinal fibers that connect with the dental pulp. In a more recent investigation Robinson et al.243 measured the concentrations of P-endorphin, another opioid-like neuropeptide, in premolars of 30 young patients whose premolars were to be extracted for orthodontic reasons. As in the case of ME, an acute mechanical stress caused a gradual decrease in the level of 3-endorphin in the pulp. In an effort to determine whether ME fluctuates in the PDL of moving teeth, we attempted to localize it immunohistochemically in orthodontically treated teeth; however, only scarce and faint immunoreactivity for ME could be detected in the unstressed PDL, with a similar appearance in the PDL of the moving tooth (unpublished results).

D.

Cytokines and Mineralized Tissues The evidence implicating cytokines in the

regulation of the activities of mineralized and nonmineralized connective tissue cells is overwhelming. Following their initial discovery, cytokines were believed to be signal molecules produced by leukocytes, serving primarily as communication links between cells of the immune system. However, many other cell types were later found to synthesize cytokine-like molecules that could function in an autocrine or paracrine capacity. With respect to bone metabolism, cytokines with demonstrated or suspected effects are IL-1, IL-2, IL-3, IL-6, TNF-a, and IFN--y. Of these cytokines, the most potent stimulator of bone resorption in vitro is IL-1. It is produced by many cell types, including osteoblasts244.245 and chondrocytes.246 Secretion of IL-1 is triggered by a variety of stimuli, including other cytokines and whole microorganisms. It has two distinct forms, IL-la and IL1p, which are coded by separate genes, but both forms have similar biologic actions. In a recent

report Kaplan et al.247 listed the systemic effects of IL-1. This long list encompasses the CNS, as well as the vascular and immunologic constel-

lations. On the local level, IL-1 attracts leukocytes, stimulates fibroblast proliferation, and enhances bone resorption. It thus seems to be one of the major components of the inflammatory response. Osteoblast-like cells, derived from human trabecular bone, were incubated for 1 to 3 d with various doses of IL-1 by Gowen et al.48 They reported a significant increase in the uptake of [3H] Tdr by these cells in comparison to controls, and a marked increase in cell counts at day 3. Infusion of IL-1 into normal mice by Boyce et al.249 first resulted in a PGE2-related hypocalcemia after 3 h, followed by a hypercalcemia at 24 h. In another related experiment Sabatini et al.250 infused IL-1 into mice, causing hypercalcemia at 72 h, with increased numbers of osteoclasts and bone resorption surfaces. Continuous infusion of IL-1 for 14 d into rabbit knee joints by Feige et al.251 induced severe arthritic changes. In vitro, synovial fibroblasts were stimulated by IL-1 to produce PGE2 and collagenase,252 and Rafter253 proposed that polymorphonuclear leukocytes are the main source for IL-1 in arthritic joints. However, in a recent interesting experiment Johnson et al.254 discovered that rat synovial fibroblasts could be stimulated by lipopolysaccharides to produce and secrete IL-1, only following an initial exposure of the cells to IFN-y. In samples of gingival cervicular fluid and in gingival tissue, IL-la and IL-113 were detected in patients with periodontal disease.255 Marked reductions in IL-1 levels followed effective periodontal treatment. In bone, target cells for IL-1 appear to be osteoblasts, according to Thomson et al.,256 who incubated neonatal mouse tibial osteoclasts with human cortical bone and with IL-1 in the presence or absence of calvarial osteoblasts. Resorption of bone occurred only when osteoclasts and osteoblasts were present. When fetal rat long bones were incubated with IL-l a or IL-13p and with PTH by Dewhirst et al.,257 a synergistic effect on 45Ca release was recorded when both cytokine and hormone were introduced simultaneously. Moreover, the presence of small concentrations of IL-1 necessitated only a very small 435


amount of PTH to cause a marked resorptive effect. This finding is potentially very significant, because it means that in cases of induced bone resorption, such as that occurring during tooth movement, suboptimal amounts of key local mediators in the PDL may be sufficient to evoke alveolar bone resorption, provided that minute amounts of systemic bone-seeking hormones also happen to be present in the same site at that time. The effects of interleukins on bone resorption were investigated by Gowen and Mundy258 using the mouse calvarial assay system. They observed marked stimulation of resorption by IL-1, but not by IL-2. Tatakis et al.259 administered IL-1, IL2, or IL-3 to osteoblasts isolated from rat fetal calvaria. Only IL-1 stimulated the release of large amounts of PGE2 by the cells, and this effect was augmented by PTH. However, when osteoclasts were incubated with mouse calvaria in the presence of IL-2, their acid production increased.260 During tooth movement in cats261 we did not observe immunohistochemical staining for IL-2 in the unstressed PDL, neither did we notice it in PDL tension sites. However, after 7 and 14 d of force application to teeth, intense staining for IL2 was observed in clusters of mononucleated cells in PDL compression sites in close proximity to osteoclasts and near the necrotic hyalinized zone. The timing of the appearance of IL-2 immunoreactivity in the compressed PDL may coincide with the localization of osteoclasts in Howship's lacunae in the alveolar bone. Similar timing was reported by Nieto-Sampedro and Chandy262 to occur in injured rat brain, where IL-2 activity in the tissue immediately adjacent to the injury reached a peak 10 d postlesion. This peak coincided with axonal sprouting and astrocyte division. In tooth movement, IL-2 may thus be associated with attraction and/or proliferation of osteoclast progenitors, as well as with a stimulation of acid production by active osteoclasts. TNF-a is another cytokine with well-documented potential as a stimulator of bone resorption. This 17-kDa molecule is synthesized mainly by monocytes and macrophages, and is an im-

portant component of the inflammatory process. It stimulates endothelial cells to secrete IL-1263 and increases their adherence to leukocytes.264 In fetal mouse calvaria TNF-a increased the pro-

duction of procollagenase and the activity of collagenase.266 In human osteoblast-like cells TNFa inhibited proliferation and alkaline phosphatase activity.267 This effect was not mediated by cAMP, and, moreover, TNF-a reduced the cAMP elevation caused by PTH.268 In terms of bone resorbing potency, IL-1 is much stronger than TNF-a; however, Stashenko et al.269 reported that suboptimal concentrations of IL-1 3 or IL-la, in combination with suboptimal concentrations of TNF-a, stimulate the formation of osteoclast-like cells in vitro from human marrow cells,270 and this effect is synergistic when both agents are added simultaneously to the cell cultures. In living animals, TNF-a administration has an inhibitory effect on bone fracture healing271 and it induces hypercalcemia.272 Using monoclonal antibodies, Hopkins and Meager273 detected low levels of TNF-a and INF-y in synovial fluids of patients with rheumatoid arthritis, while Maury and Teppo274 measured elevated levels of TNF-a in the circulation of 46% of patients with rheumatoid arthritis and 29% of patients with lupus erythematosus. Using immunohistochemical staining, Yocum et al.275 localized TNF-a in mononuclear cells of the joint lining layer, sublining, and perivascular areas. During tooth movement in cats, intense cellular staining for TNF-a was observed in the compressed PDL after 7 and 14 d of force application, particularly in fibroblasts near alveolar bone osteoclasts. However, PDL cells in tension sites also contained positive TNF-a immunoreactivity. While IL-1, IL-2, and TNF-a have been found to stimulate and enhance bone resorption in vivo and in vitro, IFN-y was discovered to interfere with resorptive processes. This cytokine is a lymphocytic product, which antagonizes the effects of a number of growth factors. Gowen et al.276 reported that IFN-y completely abolished the resorptive effects of IL-1 and TNF-a in mouse calvaria, but that it did not inhibit the resorptive effects of PTH and vitamin D3. Direct effects of IFN-y and TNF-a on human osteoblast-like cells were demonstrated by Gowen et al.277 Cellular proliferation and PGE2 production were stimulated by TNF-a, while alkaline phosphatase activity and osteocalcin release were inhibited; however, the effects IFN-y had on bone cell activities were the polar opposite. In osteosarcoma


cells with an osteoblastic phenotype, both TNFa and IFN-y inhibited DNA synthesis, and their inhibitory effect was even greater when their administration was simultaneous.278 A similar inhibitory pattern on collagen synthesis was also noted. In mice who received bone particle implants near their tibiae and seven daily injections of IFN-y, macrophages that fused to form osteoclast-like cells increased in number, although IFNy inhibited the fusion of alveolar macrophages in vitro, suggesting that IFN-y limits inflammation and favors tissue repair.279 In dental tissues, IFN-y was found by Melin et al.280 to stimulate proliferation of human dental pulp fibroblasts and to inhibit the synthesis of type I and III collagen and of fibronectin. We stained cat jaw sections for IFN-y,281 and found practically no immunoreactivity in the unstressed PDL or in PDL tension sites in moving teeth. However, after 7 and 14 d of tooth movement, numerous cells in PDL compression sites displayed positive staining for IFN--y. These cells were located primarily near alveolar bone osteoclasts, suggesting that IFN-y is involved in the regulation of bone resorption in vivo.

E. Neurotransmitters, Tooth Movement

Cytokines, and

The above review suggests strongly that a number of neurotransmitters and cytokines play regulatory roles in the remodeling of mineralized and nonmineralized connective tissues. Results from experiments in the author's laboratory have demonstrated that most of these signal molecules can be localized immunohistochemically in cat PDL and alveolar bone cells, and that their distribution and relative concentration are modified when the tissues are stressed mechanically. Taken together, these results imply that interactions between neurotransmitters, cytokines, and target cells occur in paradental tissues during tooth movement. However, localization of these molecules does not prove that some or all play active roles in stimulating or inhibiting cells in mechanically stressed paradental tissues. To determine whether these signal molecules can evoke a biochemical response by PDL fibroblasts, and whether such stimulation can affect the activity

of bone-resorbing and bone-forming cells, we resorted to cell and tissue culture experiments, which are summarized briefly below.

VI. THE EFFECTS OF NEUROTRANSMITTERS AND CYTOKINES ON BONE AND PDL FIBROBLASTS IN VITRO In these experiments, (1) neonatal mouse calvaria were subjected to increasing concentrations of neurotransmitters and cytokines to determine the effect of 45C release or [3H] proline incorporation; (2) human PDL fibroblasts were incubated with neurotransmitters or cytokines to determine whether this treatment altered the concentrations of cellular cAMP and PGE2; and (3) conditioned media from cytokine-treated or mechanically stressed PDL fibroblasts were tested to determine whether they would enhance bone resorption in vitro.

A. The Effects of Neurotransmitters and Cytokines on Bone Resorption and Formation In Vitro In an effort to determine whether bone cells can respond to a direct application of neurotransmitters or cytokines, we282 administered these agents to neonatal mouse calvaria in vitro for a

24 h incubation period. IL-la and IL-1iI stimulated bone resorption more potently than other cytokines. Bone formation was inhibited by PTH, IL-la, IL-1p, and TNF-a, but not by IFN-y. None of the neurotransmitters had any effect on the rate of bone formation. These data demonstrate that cytokines and neurotransmitters that have been found in the PDL during tooth movement can directly affect bone remodeling in vitro.

B. Effects of SP and Cytokines on PGE and cAMP in PDL Fibroblasts PDL fibroblasts, obtained from extracted

premolars of young orthodontic patients, were removed enzymatically and incubated with SP (1 x 10-6 M) for 1 to 120 min.219 Significant in437


creases in the levels of cellular cAMP and PGE in the medium occurred within 1 min, and persisted throughout the incubation period. Likewise, human PDL fibroblasts were incubated with graded doses of IL-la, IL- 13, TNF-a, and IFNy283 for 15, 30, and 60 min, or 2, 4, 24, 48, and 72 h. The cells responded to all the cytokines with dose- and time-related increases in the levels of cAMP and PGE. These increases were inhibited by indomethacin. In another experiment284 cytokines were added to cell-containing media, alone or in combination. The interactions between these cytokines varied in degree, depending on the particular combinations of cytokines. Moreover, the administration of cytokine combinations was found to be additive, synergistic, subtractive, or inhibitory on the production of PGE, depending on the length of incubation time. For instance, 24 h after the simultaneous administration of IL-1 p and TNF-a, the level of PGE in the medium increased synergistically, but at 72 h it was lower than that produced by control cells. In most of the time periods the addition of IFN-y in the presence of IL-1 or TNF-a caused a reduction in the level of PGE in the medium.

C. Bone Resorbing Activity Produced Human PDL Fibroblasts

by

To determine whether cytokine-stimulated PDL cells can enhance bone resorption, we285 administered IL- la, IL- 1 3, TNF-a, or IFN-y to human PDL fibroblasts for a 1-h incubation. The medium was then replaced with a fresh medium for an additional 24 h incubation period, and the latter medium (conditioned medium, CM) was added to mouse calvaria containing media. CM derived from unstimulated PDL fibroblasts increased the rate of bone resorption (45Ca release) by 2.5-fold, as compared to control (no CM) resorptive rate. CM derived from IL- lp-stimulated PDL fibroblasts caused a rate of resorption 3.5 times greater than control. Resorption in the presence of CM obtained from cells stimulated by IFN-y or IL- 13 + IFN--y was at the same level as that caused by CM of unstimulated cells. Indomethacin administration to the calvaria-containing medium abolished the synthesis of PGE, but reduced only partially the enhanced resorp-

tion rate caused by the IL-1 CM. These results suggest that PDL cells produce nonprostaglandin bone-resorbing factor(s), in addition to PGE2. The addition of anti-IL-lp monoclonal antibodies to the CM failed to inhibit the resorptive effect on the calvaria, suggesting that the resorptive factor produced by PDL fibroblasts was not IL1 3 (unpublished results). In another recent study286 we found that the resorptive effects of CM derived from PDL fibroblasts stimulated by either IL-la, IL-IL, or TNF-a could be augmented by the addition of PTH (1 U/ml) to the CM. This observation suggests that extensive resorption of bone may result from products of cytokine-stimulated PDL fibroblasts, when a systemic hormone such as PTH is also present in the vicinity of the bone target cells.

D. Interactive Effects of IL-1, and Mechanical Stress in PDL Fibroblasts Human PDL fibroblasts were stretched in viby the use of convex templates for 2 to 60 min.'59 The stretching caused significant elevations in PGE and cAMP within 15 min; however, the addition of IL-Il caused synergistic, additive, or even subtractive effects, depending on the duration of the incubation time. In another recent study287 CM derived from PDL cells stimulated by mechanical stress in the presence of IL-1 caused significantly more bone resorption than CM obtained from cells that had been treated by either factor alone. Again, indomethacin inhibited PGE synthesis, but reduced bone resorption only partially. Taken together, the results of the experiments summarized in this section demonstrate that human PDL fibroblasts are sensitive to mechanical stress, as well as to a variety of chemical signals derived from the nervous, immune, and endocrine systems. If levels of cellular cAMP or PGE in the medium are used as indicators, or if the resorptive activity of CM derived from stimulated or unstimulated PDL cells is used as such, it becomes clear that when two or more of these signals, mechanical or chemical, are presented to the PDL cell simultaneously, their response would usually differ from that seen following the application of one signal alone. Thus, it may be tro


possible to observe synergistic, additive, or subtractive effects. In the in vivo situation there are usually several factors present simultaneously in

the cell's environment. Therefore, it may be reasonable to assume that in tooth movement PDL fibroblasts do not respond merely to tension or compression, but also to other signals; products of neighboring cells; members of other systems, such as endothelial cells, monocytes, and nerve cells; and cells of the epithelial rests of Malassez. Moreover, it seems likely that prostaglandins are not the sole link between PDL fibroblasts and bone cells; however, the identity of the other connecting molecules is unknown at the present time, and deserves further probing.

VII. TOOTH MOVEMENT: CONCLUSIONS AND DIRECTIONS FOR FUTURE RESEARCH For at least 2000 years it has been known that a tooth can be moved gradually from one spot in the oral cavity to a more desirable one by the application of mechanical forces to the tooth's crown. In the middle of the 18th century, John Hunter explained that this movement is an outcome of the habit of the bone "to move out of the way of pressure". About 100 years later, John Farrar attributed force-induced tooth movement to "decalcification of the root socket and bending of the alveolar bone". In the years since, extensive investigations have confirmed that indeed physical and chemical alterations occur concomitantly in the paradental tissues, resulting in cellular activities that culminate in tissue remodeling and tooth movement. Histologic studies made it clear that tissues can be remodeled only by the action of cells. In the case of the PDL, the cells that form and degrade the periodontal extracellular matrix are primarily the fibroblasts. In the case of the alveolar bone, the cells that remodel it are the osteoblasts, osteoclasts, and osteocytes. When the PDL and alveolar bone are stressed by continuously applied forces, both cells and matrix are distorted and extracellular fluids are mobilized. Bone matrix distortion in vivo is associated with reorientation of its proteoglycans, a phenomenon that may serve as "strain memory", as the dis-

torted molecules return slowly to their original configuration. This strain-related distortion is also associated with the appearance of piezoelectric spikes, while fluid flow leads to slow-dissipating streaming potentials. These bioelectric phenomena can cause alterations in the polarity of the plasma membranes of cells, leading to activation of membrane enzymes and cell-matrix interactions. Stress-caused changes in the shape of cells can result in the crystallization of cytoskeletal filaments and opening or closing of stress-related membrane ion channels. Thus, fibroblasts and bone cells can respond, in vivo and in vitro, to the distortive effects of mechanical stress, which are expressed both in the cells and their surrounding matrix. Significantly, bone cells appear to be sensitive to short-duration exposures to mechanical loads whose distribution in the bone matrix deviates from their regular dispersion pattern. This finding may suggest that properly designed force systems may obviate the need for prolonged periods of force application, as is customary in most of the prevalent orthodontic techniques. However, while such brief loads are capable of evoking osteoblastic activity, it is not evident that resorptive functions can also be stimulated by these strains. The latter may perhaps necessitate the application of prolonged strains, particularly those that cause periodontal injury. Thus, from the orthodontic standpoint, a clinically effective force should be somewhat biodisruptive, i.e., be capable of causing an inflammatory/reparative reaction in the PDL and alveolar bone. Osteoclastic activity is quite intense in such a case, and it is the activity of these multinucleated cells that removes the alveolar bone that stands in the way of the moving teeth. The applied mechanical stress causes gradual fluid shifts within the PDL, resulting in nerve fiber distortion, which leads to the release of neuropeptides from nerve endings. Some of these neuropeptides have vasoactive properties, and their interaction with PDL capillaries causes vasodilation, plasma extravasation, and migration of leukocytes from the capillaries into the extravascular spaces of the PDL. These migratory leukocytes synthesize and secrete a variety of cytokines capable of stimulating fibroblasts, endothelial cells, and alveolar bone cells. The PDL fibroblasts are responsive to the neuropep439


tides, cytokines, and several growth factors produced by endothelial cells and bone cells. These fibroblasts, in areas of tension, proliferate and synthesize new matrix components, and in areas of compression they degrade the necrotic PDL. However, in both sites of tension and compression, the fibroblasts seem to produce factors that activate neighboring bone cells. Recent evidence suggests that osteoclasts are regulated by factors derived from adjacent osteoblasts, and PGE2 was proposed as being a major part of this bridge. However, in vitro experiments with PDL fibroblasts and mouse calvaria have demonstrated that factors produced by unstimulated or stimulated (mechanically or chemically) PDL fibroblasts can markedly enhance the rate of bone resorption. Thus, in tooth movement, PDL fibroblasts may not only be responsible for the remodeling of the periodontal matrix, but may also be actively involved in the regulation of the activity of the cells that remodel the alveolar bone. Osteocytes also seem to be sensitive to applied loads, and it was suggested that these cells, which are capable of recognizing and responding to molecular reorientation in their surrounding matrix, communicate these alterations to bone surface cells (primarily osteoblasts), providing them with an osteogenic stimulus. Orthodontic tooth movement is based predominantly on the application of mechanical forces to teeth in a judicious fashion supported by biomechanical principles. The rationale for investigating associated biological phenomena is derived from the desire to gain a thorough insight into these events in order to determine whether our clinical means to move teeth are effective and unharmful. Furthermore, this rationale is derived from our everlasting quest to improve our clinical approaches and from the realization that such progress can be derived from increased knowledge of biological principles on the tissue, cellular, and molecular levels. Many questions remain, however, with the biochemical and physical systems, only partly understood, but certainly deserving much further investigation. Among these issues are the effects of mechanical stresses on cells of the epithelial rests of Malassez, and on alveolar bone marrow spaces and the interaction of these cells with cells of the PDL. Another poorly understood phenom-

enon is the tooth movement-related dental root resorption. Above all remains our inability to predict the nature and pattern of the individual biological response to tooth moving forces. It seems likely that new information on these issues will be derived from experiments that correlate and integrate in vivo and in vitro systems and findings. The ability to maintain and challenge human PDL fibroblasts in vitro offers an opportunity to thoroughly examine one of the major cell types that is directly affected by orthodontic forces. Similar approaches can be applied to other PDL cell types, i.e., endothelial and epithelial cells. Interactions between these cell types and bone cells remain an intriguing issue within the framework of force-induced cellular activation and tissue remodeling. Ultimately, further explorations in this area should elucidate the cellular/molecular basis of tooth movement, and enable clinicians to include biological considerations in planning treatment for individual patients.

ACKNOWLEDGMENT

Supported by grant number DE-08428 from Institute of Dental Research. National the REFERENCES Weinberger, B. W., Orthodontics, An Historical Review of its Origin and Evolution, C. V. Mosby, St. Louis, 1926, 54. 2. Weinberger, B. W., Orthodontics, An Historical Review of its Origin and Evolution, C. V. Mosby, St. Louis, 1926, 138. 3. Hunter, J., The Natural History of the Human Teeth, J. Johnson, London, 1778, 99. 4. Delabarre, C. F., Odontologie ou Observations sans les Dents Humaines, Paris, 1815. 5. Farrar,J. N., Treatise on Irregularities of the Teeth and Their Correction, Vol. 2, DeVinne Press, New York, 1888, 758. 6. Wolff, J. D., Das Gesetz der Transformation der Knochen, Hirshwald, Berlin, 1982. 7. Sandstedt, C., Einige beitrage zur theori der zahnregulierung, Nord. Tandlaeg, Tidskr., 5, 236, 1904. 8. Sandstedt, C., Einige beitrage zur theori der zahnregulierung, Nord. Tandlaeg, Tidskr., 6, 1, 1905. 9. Oppenheim, A., Tissue changes, particularly of the bone, incident to tooth movement, Trans. Eur. Orthodont. Soc., 8, 11, 1911. 1.


Schwarz, A. M., Tissue changes incidental to orthodontic tooth movement, Int.J. Orthodont., 18, 331, 1932. 11. Fukada, E. and Yasuda, I., On piezoelectric effect of bone, J. Phys. Soc. Jpn., 12, 1158, 1957. 12. Gillooly, C. J., Hosley, R. T., Mathews, J. R., and Jewett, D. L., Electric potentials recorded from mandibular alveolar bone as a result of forces applied to the tooth, Am. J. Orthodont., 54, 649, 1968. 13. Baumrind, S., A reconsideration of the propriety of the "pressure-tension" hypothesis, Am. J. Orthodont., 55, 12, 1969. 14. Reitan, K., The initial tissue reaction incident to orthodontic tooth movement as related to the influence of function, Acta Odont. Scand., Suppl. 6, 1951. 15. Reitan, K., Tissue reaction as related to the age factor, Dent. Rec., 74, 271, 1954. 16. Reitan, K., Experiments of rotation of teeth and their subsequent retention, Trans. Eur. Orthodont. Soc., 34, 124, 1958. 17. Reitan, K., Behavior of Malassez' epithelial rests during orthodontic tooth movement, Acta Odont. Scand., 19, 425, 1961. 18. Reitan, K., Effects of force magnitude and direction of tooth movement on different alveolar bone types, Angle Orthodont., 34, 244, 1964. 19. Reitan, K., Initial tissue response behavior during apical tooth resorption, Angle Orthodont., 44, 68, 1974. 20. Reitan, K. and Kvam, E., Comparative behavior of human and animal tissue during experimental tooth movement, Angle Orthodont., 11, 1, 1971. 21. Storey, E. and Smith, R., Force in orthodontics and its relation to tooth movement, Aust. J. Dent., 56, 11, 1952. 22. Smith, R. and Storey, E., The importance of force in orthodontics, Aust. J. Dent., 56, 291, 1952. 23. Storey, E., Bone changes associated with tooth movement: the influence of the menstrual cycle on the rate of tooth movemement, Aust. J. Dent., 58, 80, 1954. 24. Storey, E., Bone changes associated with tooth movement: a histological study of the effect of force in the rabbit, guinea pig and rat, Aust. J. Dent., 59, 147, 1955. 25. Storey, E., Bone changes associated with tooth movement: a histological study of the effect of force for varying durations in the rabbit, guinea pig and rat, Aust. J. Dent., 59, 209, 1955. 26. Storey, E., Bone changes associated with tooth movement: a histological study of the effect of age and sex in the rabbit and guinea pig, Aust. J. Dent., 59, 220, 1955. 27. Storey, E., The nature of tooth movement, Am. J. Orthodont., 63, 292, 1973. 28. Deguchi, T. and Mori, M., Histochemical observations on oxidative enzymes in periodontal tissue during experimental tooth movement in rat, Arch. Oral Biol., 13, 49, 1968. 10.

29.

Takimoto, K., Deguchi, T., and Mori, M., Histochemical detection of succinic dehydrogenase in

osteoclasts following experimental tooth movement, J. Dent. Res., 45, 1473, 1966.

Takimoto, K., Deguchi, T., and Mori, M., Histochemical detection of acid and alkaline phosphatase in periodontal tissue following experimental tooth movement, J. Dent. Res., 47, 340, 1968. 31. Waldo, C. M. and Rothblatt, J. M., Histologic response to tooth movement in the laboratory rat: procedure and preliminary observation, J. Dent. Res., 33, 481, 1954. 32. Lilja, E., Lindskog, S., and Hammarstrom, L., Histochemistry of enzymes associated with tissue degradation incident to orthodontic tooth movement, Am. J. Orthodont., 83, 62, 1983. 33. Lilja, E., Bjornestedt, T., and Lindskog, S., Cellular enzyme activity associated with tissue degradation following orthodontic tooth movement in man, Scand. J. Dent. Res., 91, 381, 1983. 34. Kurihara, S. and Enlow, D. H., A histochemical and electron microscopic study of an adhesive type of collagen attachment on resorptive surfaces of alveolar bone, Am. J. Orthodont., 77, 532, 1980. 35. Martinez, R. H. and Johnson, R. B., Effects of orthodontic tooth movement on the alcian blue staining patterns of rat alveolar bone: a histochemical study, Histol. Histopathol., 3, 39, 1988. 36. Noda, K., Localization of cytochrome c oxidase activity in osteoclasts treated with calcitonin during experimental tooth movement, Acta Histochem. Cytochem., 21, 301, 1988. 37. Rygh, P. and Selvig, K. A., Erythrocytic crystallization in rat molar periodontium incident to tooth movement, Scand. J. Dent. Res., 81, 62, 1973. 38. Ten Cate, A. R., Deporter, D. A., and Freeman, E., The role of the fibroblasts in the remodeling of periodontal ligament during physiological tooth movement, Am. J. Orthodont., 69, 155, 1976. 39. Ten Cate, A. R., Freeman, E., and Dickinson, J. B., Sutural development: structure and its response to rapid expansion, Am. J. Orthodont., 71,622, 1977. 40. Rygh, P., Elimination of hyalinized periodontal tissues associated with orthodontic tooth movement, Scand. J. Dent. Res., 82, 57, 1974. 41. Rygh, P., Ultrastructural changes in tension zone of rat molar periodontium incident to orthodontic tooth movement, Am. J. Orthodont., 70, 269, 1976. 42. Edwards, J. E., A study of the periodontium during orthodontic rotation of teeth, Am. J. Orthodont., 54, 441, 1968. 43. Martinez, R. H. and Johnson, R. B., Effects of orthodontic forces on the morphology and diameter of Sharpey fibers on the alveolar bone of the rat, Anat. Rec., 219, 10, 1987. 44. Kurihara, S. and Enlow, D. H., An electron microscopic study of attachments between periodontal fibers and bone during alveolar remodeling, Am. J. Orthodont., 77, 516, 1980. 30.


45.

Kvam, E., Scanning electron microscopy of tissue

changes on the pressure surface of human premolars following tooth movement, Scand. J. Dent. Res., 80,

ulated 61.

357, 1972. 46. Barber, A. F. and Sims, M. R., Rapid maxillary

expansion and external root resorption in man: a scanning electron microscope study, Am. J. Orthodont., 79, 630, 1981.

47.

Langford, S. R. and Sims, M. R., Root surface resorption, repair, and periodontal attachment following rapid maxillary expansion in man, Am. J. Orthodont., 81, 108, 1982.

Rygh, P., Orthodontic root resorption studied by electron microscopy, Angle Orthodont., 47, 1, 1977. 49. Nakane, S. and Kameyama, Y., Root resorption caused by mechanical injury of the periodontal soft

48.

tissue in rats, J. Periodont Res., 22, 390, 1987.

Goldie, R. S. and King, G. J., Root resorption and tooth movement in orthodontically treated, calciumdeficient, and lactating rats, Am. J. Orthodont., 85, 424, 1984. 51. Engstrom, C., Granstrom, G., and Thilander, B., Effect of orthodontic force on peridontal tissue metabolism, Am. J. Orthodont. Dentofacial Orthoped., 93, 486, 1988. 52. Garant, P. R. and Cho, M. I., Cytoplasmic polarization of periodontal ligament fibroblasts. Implications for cell migration and collagen secretion, J. 50.

53.

54.

55.

56.

57.

58. 59. 60.

Periodont. Res., 14, 95, 1979. Garant, P. R., Cho, M. I., and Cullen, M. R., Attachment of periodontal ligament fibroblasts to the extracellular matrix in the squirrel monkey, J. Periodont. Res., 17, 70, 1982. Cho, M. I. and Garant, P. R., Mirror symmetry of newly divided rat periodontal ligament fibroblasts in situ and its relationship to cell migration, J. Periodont. Res., 20, 185, 1985. Garant, P. R. and Cho, M. I., Fibroblast migration, cytoplasmic polarity, and matrix secretion in the periodontal ligament, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC Press, Boca Raton, FL, 1989, 29. Willingham, M. C., Yamada, K. M., Yamada, S. S., Pouyssegur, J., and Pastan, I., Microfilament bundles and cell shape are related to adhesiveness to substratum and are dissociable from growth control in cultured fibroblasts, Cell, 10, 375, 1977. McCulloch, C. A. G., Barghava, U., and Melcher, A. H., Cell death and the regulation of populations of cells in the periodontal ligament, Cell Tissue Res., 255, 129, 1989. Kvam, E., Cellular dynamics on the pressure side of the rat periodontium following experimental tooth movement, Scand. J. Dent. Res., 80, 369, 1972. Roberts, W. E. and Jee, W. S. S., Cell kinetics of orthodontically-stimulated periodontal ligament in the rat, Arch. Oral Biol., 19, 17, 1974. Roberts, W. E., Chase, D. C., and Jee, W. S. S., Counts of labeled mitoses in the orthodontically stim-

periodontal ligament of the rat, Arch. Oral

Biol., 19, 665, 1974.

Roberts, W. E., Cell kinetic nature and diurnal periodicity of the rat periodontal ligament, Arch. Oral

Biol., 20, 456, 1975.

Roberts, W. E., Aubert, M. M., Sparaga, J. A., and Smith, R. K., Circadian periodicity of cell kinetics of rat molar periodontal ligament, Am. J. Orthodont., 76, 316, 1979. 63. Smith, R. K. and Roberts, W. E., Cell kinetics of the initial response of orthodontically induced osteogenesis in rat molar periodontal ligament, Calcified Tissue Int., 30, 51, 1980. 64. Roberts, W. E. and Chase, D. C., Kinetics of cell proliferation and migration associated with orthodontically-induced osteogenesis, J. Dent. Res., 60, 174, 1981. 65. Roberts, W. E., Klinger, E., and Mozsary, P. G., Circadian rhythm of mechanically mediated differentiation of osteoblasts, Calcified Tissue Int., 36, S63, 1984. 66. Roberts, W. E. and Ferguson, D. J., Cell kinetics of the periodontal ligament, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC Press, Boca Raton, FL, 1989, 55. 67. Yee, J. A., Kimmel, D. B., and Jee, W. S. S., Periodontal ligament cell kinetics following orthodontic tooth movement, Cell Tissue Kinet., 9, 293, 1976. 68. Yee, J. A., Response of periodontal ligament cells to orthodontic force: ultrastructure identification of proliferating fibroblasts, Anat. Rec., 194, 603, 1979. 69. Roberts, W. E. and Cox, W. B., Morphometric quantification of nuclear volume changes during osteoblast histogenesis, in Bone Histomorphometry, Meunier, P. J., Ed., Armour Montagu, Paris, 1977, 267. 70. McCulloch, C. A. G., Nemeth, E., Lowenberg, B., and Melcher, A. H., Paravascular cells in endosteal spaces of alveolar bone contribute to periodontal ligament cell populations, Anat. Rec., 219, 233, 1987. 71. McCulloch, C. A. G. and Heersche, J. N. M., Lifetime of the osteoblast in mouse periodontium, Anat. Rec., 222, 129, 1988. 72. Morris, C. E., Mechanosensitive ion channels, J. Memb. Biol., 113, 93, 1990. 73. Brezden, B. L., Gardner, D. R., and Morris, C. E., A potassium-selective channel in isolated Lymnaea stagnalis heart muscle cells, J. Exp. Biol., 62.

123, 175, 1986.

74.

Moody, W. J. and Bosma, M. M., A nonselective cation channel activated by membrane deformation in oocytes of the ascidian Boltenia Villosa, J. Membr.

75.

Biol., 107, 179, 1989. Sachs, F., Mechanical transduction in biological systems, Crit. Rev. Biomed. Eng., 16, 141, 1988.

76.

Christensen, O., Mediation of cell volume regula-


tion

by Ca2+

influx

through stretch-activated chan-

nels, Nature, 330, 66, 1987. 77. Cooper, K. E., Tang, J. M., Rae, J. L., and Eisenberg, R. S., A cation channel in frog lens epithelia responsive to pressure and calcium, J. Membr. Biol., 93, 259, 1986. 78. Lansman, J. B., Hallam, T. J., and Rink, T. J.,

79. 80.

81.

82.

83.

84.

85.

86.

87.

88.

89. 90.

Single stretch-activated ion channels in vascular endothelial cells as mechanotransducers, Nature, 325, 811, 1987. Morris, C. E. and Sigurdson, W. J., Stretch-inactivated ion channels coexist with stretch-activated ion channels, Science, 243, 807, 1989. Harris, A. K., Wild, P., and Stopak, D., Silicone rubber substrata: a new wrinkle in the study of cell locomotion, Science, 208, 177, 1980. Ingber, D. E. and Folkman, J., Tension and compression as basic determinants of cell form and function: utilization of a cellular tensegrity mechanism, in Cell Shape: Determinants, Regulation, and Regulatory Role, Stein, W. D. and Bronner, F., Eds., Academic Press, San Diego, CA, 1989, 3. Vanderburgh, H. H. and Kaufman, S., Stretchinduced growth of skeletal myotubes correlates with activation of the sodium pump, J. Cell Physiol., 109, 205, 1981. Cogoli, A., Valluchi-Morf, M., Mueller, M., and Briegleb, W., Effect of hypogravity on human lymphocyte activation, Aviat. Space Environ. Med., 51, 29, 1980. Leung, D. Y. M., Glagov, S., and Mathews, M. B., Cyclic stretching simulates synthesis of matrix components by arterial smooth muscle cells in vitro, Science, 191, 475, 1976. Quinn, R. S. and Rodan, G. A., Enhancement of ornithine decarboxylase and Na+, K+ ATPase in osteosarcoma cells by intermittent compression, Biochem. Biophys. Res. Commun., 100, 1696, 1981. Ingber, D. E. and Jamieson, J. D., Cells as tensegrity structures: architectural regulation of histodifferentiation by physical forces transduced over basement membrane, in Gene Expression During Normal and Malignant Differentiation, Andersson, L. C., Gahmberg, C. G., and Ekblom, P., Eds., Academic Press, Orlando, FL, 1985, 13. Joshi, H. C., Chu, D., Buxbaum, R. E., and Heidermann, S. R., Tension and compression in the cytoskeleton of PC 12 neurites, J. Cell Biol., 101, 697, 1985. Herman, B. and Pledger, W. J., Platelet-derived growth factor-induced alterations in vinculin and actin distribution in BALB/c-3T3 cells, J. Cell Biol., 100, 1031, 1985. Berezney, R. and Coffey, D. S., Nuclear protein matrix: association with newly synthesized DNA, Science, 189, 291, 1975. Pardoll, D. M., Vogelstein, B., and Coffey, D. S., A fixed site of DNA replication in eucaryotic cells, Cell, 19, 527, 1980.

Barrak, E. R., and Coffey, D. S., Biological properties of the nuclear matrix: steroid hormone binding, Recent Prog. Hormone Res., 38, 133, 1982. 92. Nicolini, C., Belmont, A. S., and Martelli, A., Critical nuclear DNA size and distribution associated with S phase initiation, Cell Biophys., 8, 103, 1986. 93. Jiang, L.-W. and Schindler, M., Nuclear transport in 3T3 fibroblasts: effects of growth factors, transformation and cell shape, J. Cell Biol., 106, 13, 1988. 94. Emerman, J. T. and Pitelka, D. R., Maintenance and induction of morphological differentiation in associated mammary epithelium on floating collagen membranes, In Vitro, 13, 316, 1977. 95. Lanyon, L. E., Goodship, A. E., Pye, C. J., and MacFie, J. H., Mechanically adaptive bone remodeling, J. Biomech., 15, 767, 1982. 96. Rubin, C. T. and Lanyon, L. E., Regulation of bone formation by applied dynamic loads, J. Bone Jt. Surg., 66-A, 397, 1984. 97. Lanyon, L. E. and Rubin, C. T., Static vs. dynamic loads as an influence on bone remodeling, J. Biomech., 17, 897, 1984. 98. Rubin, C. T. and Lanyon, L. E., Regulation of bone mass by mechanical strain magnitude, Calcified Tissue Int., 37, 411, 1985. 99. Lanyon, L. E., Rubin, C. T., and Baust, G., Modulation of bone loss during calcium insufficiency by controlled dynamic load, Calcified Tissue Int., 38, 209, 1986. 100. Rubin, C. T. and Lanyon, L. E., Osteoregulatory nature of mechanical stimuli: function as a determinant for adaptive remodeling in bone, J. Orthoped. Res., 5, 300, 1987. 101. Lanyon, L. E., Functional strain in bone tissue as an objective, and controlling stimulus for adaptive bone remodeling, J. Biomech., 20, 1083, 1987. 102. Pead, M. J., Suswillo, R., Skerry, T. M., Vedi, S., and Lanyon, L. E., Increased 3H-uridine levels in osteocytes following a single short period of dynamic bone loading in vivo, Calcified Tissue Int., 43, 92, 1988. 103. Skerry, T. M., Bitensky, L., Chayen, J., and Lanyon, L. E., Loading-related reorientation of bone proteoglycan in vivo. Strain memory in bone tissue?, J. Orthoped. Res., 6, 547, 1988. 104. Skerry, T. M., Suswillo, R., El Haj, A. J., Ali, N. N., Dodds, R. A., and Lanyon, L. E., Loadinduced proteoglycan orientation in bone tissue in vivo and in vitro, Calcified Tissue Int., 46, 318, 1990. 105. Smith, E., Reddan, W., and Smith, P., Physical activity and calcium modalities for bone mineral increased in aged women, Med. Sci. Sports Exer., 13, 60, 1980. 106. Smith, E. L., Smith, P. E., Ensign, C. J., and Shea, M. M., Bone involution decrease in exercising middle-aged women, Calcified Tissue Int., 36, S 129, 91.

1984. 107. Mack, P. B., LaChance, P. A., Vose, G. P., and


Vogt, F., Bone demineralization of foot and hand of Gemini-Titan IV, V, and VII astronauts during orbital flight, Am. J. Roentgenol., 100, 503, 1967. 108. Cann, C. E. and Adachi, R. R., Bone resorption and mineral excretion in rats during space flight, Am. J. Physiol., 244, R327, 1983. 109. Rambaut, P. C. and Goode, A. W., Skeletal changes during space flight, Lancet, p. 1050, 9, November 1985. 110.

Bikle, D. D., Halloran, B. P., Cone, C. M., Globus, R. K., and Morey-Holton, E., The effect of sim-

ulated weightlessness on bone maturation, Endocrinology, 120, 678, 1987. 111. Yamaguchi, M., Ozaki, K., and Hoshi, T., Simulated weightlessness and bone metabolism: decrease of protein and DNA synthesis in the femoral diaphysis of rats, Res. Exp. Med., 189, 331, 1989. 112. Glucksmann, A., The role of mechanical stresses in bone formation in vitro, J. Anat., 76, 231, 1942. 113. Functional Adaptation in Bone Tissue, A Kroc Foundation Conference, July 1983, Cowin, S. C., Lanyon, L. E., and Rodan, G., Eds., Calcified Tissue Int., 36(Suppl. 1), 1, 1984. 114. Pienkowski, D. and Pollack, S. R., The origin of stress-generated potentials in fluid saturated bone, J. Orthoped. Res., 1, 30, 1983. 115. Pollack, S. R., Salzstein, R., and Pienkowski, D., The electric double layer in bone and its influence on stress-generated potentials, Calcified Tissue Int., 36, S77, 1984. 116. Marino, A. A., Spadaro, J. A., Fukada, E., Kah, L. D., and Becker, R. O., Piezoelectricity in collagen films, Calcified Tissue Int., 31, 257, 1970. 117. Fukada, E. and Yasuda, I., Piezoelectric effects in collagen, Jpn. J. Appl. Phys., 3, 117, 1964. 118. Marino, A. A. and Becker, R. O., Piezoelectricity in hydrated frozen bone and tendon, Nature, 253, 627, 1975. 119. Marino, A. A. and Becker, R. O., The origin of the piezoelectric effect in bone, Calcified Tissue Int., 8, 177, 1971. 120. Bassett, C. A. L. and Becker, R. O., Generation of electric potentials by bone in response to mechanical stress, Science, 137, 1063, 1962. 121. Cochran, G. V. B., Pawluk, R. J., and Bassett, C. A. L., Stress generated electric potentials in the mandible and teeth, Arch. Oral Biol., 12, 917, 1967. 122. Zengo, A. N., Pawluk, R. J., and Bassett, C. A. L., Stress-induced bioelectric potentials in the dentoalveolar complex, Am. J. Orthodont., 64, 17, 1973. 123. Zengo, A. N., Bassett, C. A. L., Pawluk, R. J., and Prountzos, G., In vivo bioelectric potentials in the dentoalveolar complex, Am. J. Orthodont., 66, 130, 1974. 124. Marino, A. A. and Gross, B. D., Piezoelectricity in cementum, dentine and bone, Arch. Oral Biol., 34, 507, 1989. 125. Grimm, F. M., Bone bending, a feature of ortho-

dontic tooth movement, Am. J. Orthodont., 62, 384, 1972. 126. DeAngelis, V., Observations on the response of alveolar bone to orthodontic force, Am. J. Orthodont., 58, 284, 1970. 127. Borgens, R. B., Endogenous ionic currents traverse intact and damaged bone, Science, 225, 478, 1984. 128. Otter, M., Shoenung, J., and Williams, W. S., Evidence for different sources of stress-generated potentials in wet and dry bone, J. Orthoped. Res., 3, 321, 1985. 129. Friedenberg, Z. B. and Kohanim, M., The effect of direct current on bone, Surg. Gynecol. Obstet., 127, 97, 1968. 130. Friedenberg, Z. B., Harlow, M. C., and Brighton, C. T., Healing of nonunion of the medial malleolus by means of direct current: a case report, J. Trauma, 11, 863, 1971. 131. Bassett, C. A. L., Pilla, A. A., and Pawluk, R. J., Non-operative salvage of surgically resistant pseudoarthroses and non-unions by pulsing electromagnetic fields, Clin. Orthoped., 124, 128, 1977. 132. Friedenberg, Z. B. and Brighton, C. T., Bioelectricity and fracture healing, Plastic Reconstr. Surg., 68, 435, 1981. 133. Marino, A. A., Electrical stimulation in orthopaedics: past, present and future, J. Bioelect., 3, 235, 1984. 134. Beeson, D. C., Johnston, L. E., and Wisotzky, J., Effect of constant currents on orthodontic tooth movement in the cat, J. Dent. Res., 54, 251, 1975. 135. Davidovitch, Z., Finkelson, M. D., Steigman, S., Shanfeld, J. L., Montgomery, P. C., and Korostoff, E., Electric currents, bone remodeling, and orthodontic tooth movement. I. The effect of electric currents on periodontal cyclic nucleotides, Am. J. Orthodont., 77, 14, 1980. 136. Davidovitch, Z., Finkelson, M. D., Steigman, S., Shanfeld, J. L., Montgomery, P. C., and Korostoff, E., Electric currents, bone remodeling, and orthodontic tooth movement. II. Increase in rate of tooth movement and periodontal cyclic nucleotide levels by combined force and electric current, Am. J. Orthodont., 77, 33, 1980. 137. Davidovitch, Z., Montgomery, P. C., and Shanfeld, J. L., Cellular localization and concentration of bone cyclic nucleotides in response to acute PTE administration, Calcified Tissue Int., 24, 81, 1977.

138. Murad, F., Brewer, B., Jr., and Vaughn, M., Effect of thyrocalcitonin on adenosine, 3',5'-cyclic phosphate formation by rat kidney and bone, Proc. Natl. Acad. Sci. U.S.A., 65, 446, 1970. 139. Rasmussen, H. and Bordier, P., Vitamin D and bone, Metab. Bone Dis. Relat. Res., 1, 7, 1978. 140. Kornstein, R., Somjen, D., Danon, A., Fisher, H., and Binderman, I., Pulsed capacitive electric induction of cyclic AMP changes, Ca45 uptake and


DNA synthesis in bone cells, Trans. Bioelectr. Repair Growth, 1, 34, 1981. 141. Rodan, G. A., Bourret, L. A., Harvey, A., and Mensi, T., Cyclic AMP and cyclic GMP: mediators of the mechanical effects on bone remodeling, Sci-

ence, 189, 467, 1975. 142.

Tashjian, A. H., Jr., Voelkel, E. F., Levine, L., and Goldhaber, P., Evidence that the bone resorption-stimulating factor produced by mouse fibrosarcoma cells is prostaglandin E2, J. Exp. Med., 136, 1329, 172.

Gomes, B. C., Hausmann, E., Weinfeld, N., and DeLuca, C., Prostaglandins: bone resorption stimulating factors released from monkey gingiva, Calcified Tissue Int., 19, 285, 1976. 144. Shiozawa, S., Williams, R. C., Jr., and Ziff, M., Immunoelectron microscopic demonstration of prostaglandin E in rheumatoid synovium, Arthr. Rheum., 25, 685, 1982. 145. Dekel, S., Lenthal, G., and Francis, M. J. 0., Release of prostaglandins from bone and muscle after tibial fracture, Bone Jt. Surg., 63B, 185, 1981. 146. Uchida, A., Yamashita, K., Hashimoto, K., and Shimomura, Y., The effects of mechanical stress on cultured growth cartilage cells, Connect. Tissue Res.,

143.

17, 305, 1988.

147.

Somjen, D., Binderman, I., Berger, E., and Harell, A., Bone remodeling induced by physical stress is prostaglandin E2 mediated, Biochim. Biophys. Acta, 627, 91, 1980.

148.

Shen, V., Rifas, L., Kohler, G., and Peck, W. A., Prostaglandins change cell shape and increase intercellular gap junctions in osteoblasts cultured frdm rat fetal calvaria, J. Bone Min. Res., 1, 243, 1986.

149.

Hasegawa, S., Sato, S., Saito, S., Suzuki, Y., and Brunette, D. M., Mechanical stretching increases the number of cultured bone cells synthesizing DNA and alters their pattern of protein synthesis, Calcified Tissue Int., 37, 431, 1985.

Buckley, M. J., Banes, A. J., Levin, L. G., Sumpio, B. E., Sato, M., Jordan, R., Gilbert, J., Link, G. W., and Tran Son Tay, R., Osteoblasts increase their rate of division and align in response to cyclic, mechanical tension in vitro, Bone Min., 4, 225, 1988. 151. Klein-Nutlend, J., Velhuijzen, J. P., de Jong, M., and Burger, E. H., Increased bone formation and decreased bone resorption in fetal mouse calvaria as a result of intermittent compressive force, in vitro, 150.

2, 441, 1987. E. Burger, H., Velhuijzen, J. P., Klein-Nulend, J., and Van Loon, J. J. W. A., Osteoclastic invasion and mineral resorption of fetal mouse long bone rudiments are inhibitied by culture under intermittent compressive force, Connect. Tissue Res., 20, 131, 1989.

Bone Min.,

152.

153.

Ozawa, H., Imamura, K., Etsuko, A., Takahashi, N., Hiraide, T., Shibasaki, Y., Fukuhara, T., and Suda, T., Effect of continuously applied compressive

pressure on mouse osteoblast-like cells (MC3T3-E1) in vitro, J. Cell Physiol., 142, 177, 1990. 154. Duncan, G. W., Yen, E. H. K., Pritchard, E. T., and Suga, D. M., Collagen and prostaglandin synthesis in force-stressed periodontal ligament, in vitro,

J. Dent. Res., 63, 665, 1984. Meikle, M. C., Heath, J. K., Atkinson, S. J., Hembry, R. M., and Reynolds, J. J., Molecular biology of stressed connective tissues at sutures and hard tissues in vitro, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC Press, Boca Raton, FL, 1989, 71. 156. Curtis, A. S. G. and Sechar, G. M., The control of cell division by tension or diffusion, Nature, 274, 52, 1978. 157. Norton, L. A., Anderson, K. L., Melsen, B., Arenholt Bindslev, D., and Celis, J. E., Buccal mucosa fibroblasts and periodontal ligament cells perturbed by tensile stimuli in vitro, Scand, J. Dent. Res., 98, 36, 1990. 158. Ngan, P. W., Crock, B., Varghese, J., Lanese, R., Shanfeld, J. L., and Davidovitch, Z., Immunohistochemical assessment of the effect of chemical and mechanical stimuli on cAMP and prostaglandin E levels in human gingival fibroblasts, in vitro, Arch. Oral Biol., 33, 163, 1988. 159. Ngan, P., Saito, S., Saito, M., Shanfeld, J., and Davidovitch, Z., The interactive effects of mechanical stress and interleukin 1 on prostaglandin E and cyclic AMP production in human periodontal ligament fiborblasts in vitro: comparison with cloned osteoblastic cells of mouse (MC3T3-E1), Arch. Oral Biol., 35, 717, 1990. 160. Ragnarsson, B., Carr, G., and Daniel, J. C., Isolation and growth of human periodontal ligament cells in vitro, J. Dent. Res., 64, 1026, 1985. 161. Brunette, D. M., Kanoza, R. J., Marmary, Y., Chan, J., and Melcher, A. H., Interactions between epithelial and fibroblast-like cells in cultures derived from monkey periodontal ligament, J. Cell Sci., 27, 127, 1977. 162. Brunette, D. M., Heersche, J. N. M., Purdon, A. D., Sodek, J., Moe, H. K., and Assuras, J. N., In vitro cultured parameters and protein and prostaglandin secretion of epithelial cells derived from porcine rests of Malassez, Arch. Oral Biol., 24, 199, 1979. 163. Merrilees, M. J., Sodek, J., and Aubin, J. E., Effect of cells of epithelial rests of Malassez and endothelial cells on synthesis of glycosaminoglycans by periodontal ligament fibroblasts in vitro, Dev. Biol., 97, 146, 1983. 164. Birek, C., Heersche, J. N. M., Jez, D., and Brunette, D. M., Secretion of a bone resorbing factor by epithelial cells cultured from porcine rests of Malassez, J. Periodont. Res., 18, 75, 1983. 165. Brunette, D. M., Mechanical stretching increases

155.


the number of epithelial cells

synthesizing DNA in

culture, J. Cell Sci., 69, 35, 1984. 166. Klein, D. C. and Raisz, L. G., Prostaglandins: stim-

ulation of bone resorption in tissue culture, Endocrinology, 86, 1436, 1970. 167. Rodan, G. A. and Martin, T. J., Role of osteoblasts in hormonal control of bone resorption a hypothesis, Calcified Tissue Int., 33, 349, 1981. 168. Bindeman, I., Zor, U., Kaye, A. M., Shimshoni, Z., Harell, A., and Somjen, D., The transduction

of mechanical force into biochemical events in bone cells may involve activation of phospholipase A2, Calcified Tissue Int., 42, 261, 1988.

169. Davidovitch, Z., Shanfeld, J. L., Montgomery, P. C., Lally, E., Laster, L., Furst, L., and Korostoff, E., Biochemical mediators of the effects of mechanical forces and electric currents on mineralized tissues, Calcified Tissue Int., 36, S86, 1984. 170. Davidovitch, Z., Nicolay, O., Alley, K., Zwilling, B., Lanese, R., and Shanfeld, J. L., First and second messenger interactions in stressed connective tissues in vivo, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC Press, Boca Raton, FL, 1989, 97. 171. Yamasaki, K., Miura, F., and Suda, T., Prostaglandin as a mediator of bone resorption induced by experimental tooth movement in rats, J. Dent. Res., 59, 1635, 1980. 172. Yamasaki, K., Shibata, Y., and Fukuhara, T., The effects of prostaglandins on experimental tooth movement in monkeys (Macaca fuscata), J. Dent. Res., 61, 1444, 1982. 173. Yamasaki, K., Shibata, Y., Imai, S., Tani, Y., Shibasaki, Y., and Fukuhara, T., Clinical application of prostaglandin E, (PGE,) upon orthodontic tooth movement, Am. J. Orthodont., 85, 508, 1984. 174. Yamasaki, K., Pharmacological control of tooth movement, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC Press, Boca Raton, FL, 1989, 287. 175. Jee, W. S. S., Ueno, K., Deng. Y. P., and Woodbury, D. M., The effect of prostaglandin E2 in growing rats: increased metaphyseal hard tissue and cortico-endosteal bone formation, Calcified Tissue Int., 37, 148, 1985. 176. Marks, S. C., Jr. and Miller, S. C., Local infusion of prostaglandin E, stimulates mandibular bone formation in vivo, J. Oral Pathol., 17, 500, 1988. 177. Nefussi, J.-R. and Baron, R., PGE2 stimulates both resorption and formation of bone in vitro: differential responses of the periosteum and the endosteum in fetal rat long bone cultures, Anat. Rec., 211,9, 1985. 178. Lee, W., Experimental study of the effect of prostaglandin administration on tooth movement with particular emphasis on the relationship to the method of PGE, administration, Am. J. Orthodont. Dentofacial Orthoped., 98, 231, 1990. 179.

Decker, J. L., Malone, D. G., Haroui, B., Wahl, S., Schrieber, L., Klippel, J., Steinberg, A. G.,

and Wilder, R., Rheumatoid arthritis: evolving concepts of pathogenesis and treatment, Ann. Intern. Med., 101, 810, 1984. 180.. Brandt, K. D. and Slowman-Kovacs, S., Nonsteroidal antiinflammatory drugs in treatment of osteoarthritis, Clin. Orthoped., 213, 84, 1986. 181. Torbinejad, M., Clagett, J., and Engel, D., A cat model for the evaluation of mechanisms of bone resorption: induction of bone loss by stimulated immune complexes and inhibition by indomethacin, Calcified Tissue Int., 29, 207, 1979. 182. Waite, I. M., Saxton, C. A., Young, A., Wagg, B. J., and Corbett, M., The periodontal status of subjects receiving non-steroidal anti-inflammatory drugs, J. Periodont. Res., 16, 100, 1981. 183. Saffar, J. L. and Lasfargues, J. J., A histometric study of the effect of indomethacin and calcitonin on bone remodeling in hamster periodontitis, Arch. Oral Biol., 29, 555, 1984. 184. Williams, R. C., Jeffcoat, M. K., Kaplan, M. L., Goldhaber, P., Johnson, H. G., and Wechter, W. J., Flurbiprofen: a potent inhibitor of alveolar bone resorption in beagles, Science, 277, 640, 1985. 185. Melcher, A. H., An overview of the anatomy and physiology of the periodontal ligament, in The Biology of Tooth Movement, Norton, L. A. and Burstone, C. J., Eds., CRC Press, Boca Raton, FL, 1989, 1. 186. van Steenberghe, D., The structure and function of periodontal innervations, J. Periodont. Res., 14, 185, 1979. 187. Pfaffman, C., Afferent impulses from the teeth due to pressure and noxious stimulation, J. Physiol., 97, 207, 1939. 188. Cash, R. M. and Linden, R. W. A., The distribution of mechanoreceptors in the periodontal ligament of the mandibular canine teeth of the cat, J. Physiol., 330, 439, 1982. 189. Jyvisjiirvi, E., Kniffki, K.-D., and Mengel, M. K. C., Functional characteristics of afferent C fibers from tooth pulp and periodontal ligament, Prog. Brain Res., 74, 237, 1988. 190. Freezer, S. R. and Sims, M. R., A transmission electron-microscopy stereological study of the blood vessels, oxytalan fibres and nerves of mouse-molar periodontal ligament, Arch. Oral Biol., 32, 407, 1987. 191. Gould, T. R. L., Melcher, A. H., and Brunette, D. M., Location of progenitor cells of periodontal ligament of mouse molar stimulated by wounding, Anat. Rec., 188, 133, 1977. 192. McCulloch, C. A. G. and Melcher, A. H., Cell density and cell generation in the periodontal ligament of mice, Am. J. Anat., 167, 43, 1983. 193. Khouw, F. E. and Goldhaber, P., Changes in vasculature of the periodontium associated with tooth movement in the rhesus monkey and dog, Arch. Oral Biol., 15, 1125, 1970. 194. Rygh, P., Ultrastructural vascular changes in pressure zones of rat molar periodontium incident to or-


195.

196.

197.

198.

thodontic movement, Scand. J. Dent. Res., 80, 307, 1972. Jeffcoat, M. K., Kaplan, M. L., Rumbaugh, C. L., and Goldhaber, P., Magnification angiography in beagles with periodontal disease, J. Periodont. Res., 17, 294, 1982. Bien, S. M., Fluid dynamic mechanisms which regulate tooth movement, Adv. Oral Biol., 2, 173, 1966. Lotz, M., Vaughan, J. H., and Carson, D. A., Effect of neuropeptides on production of inflammatory cytokines by human monocytes, Science, 241, 1218, 1988. Laurenzi, M. A., Persson, M. A. A., Dalsgaard, C.-J., and Ringden, O., Stimulation of human B lymphocyte differentiation by the neuropeptides substance P and neurokinin A, Scand. J. Immunol., 30, 695, 1989.

199. Hafstrom, I., Gyllenhammar, H., Palmblad, J., and Ringertz, B., Substance P activates and modulates neutrophil oxidative metabolism and aggregation, J. Rhemuatol., 16, 1033, 1989. 200. Roscetti, G., Ausiello, C. M., Palma, C., Gulla, P., and Roda, G., Enkephalin activity on antigeninduced proliferation of human peripheral blood mononucleate cells, Int. J. Immunopharm., 10, 819, 1988. 201. Marotti, T., Sverko, V., and Hrsak, I., Modulation of superoxide anion release from human polymorphonuclear cells by Met- and Leu-enkephalin, Brain, Behav. Immun., 4, 13, 1990. 202. Weinstock, J. V., Blum, A., Walder, J., and Walder, R., Eosinophils from granulomas in Schistosomiasis mansoni produce substance P, J. Immunol., 141, 961, 1988. 203. Weinstock, J. V. and Blum, A. M., Release of substance P by granuloma eosinophils in response to secretagogues in murine Schistosomiasis mansoni, Cell Immunol., 125, 380, 1990. 204. Roth, K. A., Lorenz, R. G., Unanue, R. A., and Weaver, C. T., Nonopiate active proenkephalin-derived peptides are secreted by T helper cells, FASEB J., 3, 2402, 1989. 205. Fagarasan, M. O., Bishop, J. F., Rinaudo, M. S., and Axelrod, J., Interleukin 1 induces early protein phosphorylation and requires only a short exposure for late induced secretion of P-endorphin in a mouse pituitary cell line, Proc. Natl. Acad. Sci. U.S.A., 87, 2555, 1990. 206. Fagarasan, M. 0. and Axelrod, J., Interleukin-1 amplifies the action of pituitary secretagogues via protein kinases, Int. J. Neurosci., 51, 311, 1990. 207. Su, T.-P., London, E. D., and Jaffe, J. H., Steroid binding at a receptors suggests a link between endocrine, nervous, and immune systems, Science, 240, 219, 1988. 208. Levine, J. D., Clark, R., Devor, M., Helms, C., Moskowitz, M. A., and Basbaum, A. I., Intraneuronal substance P contributes to the severity of experimental arthritis, Science, 226, 547, 1984. 209. Lotz, M., Carson, D. A., and Vaughan, J. H.,

210. 211.

212. 213. 214.

215.

Substance P activation of rheumatoid synoviocytes: neural pathway in pathogenisis of arthritis, Science, 235, 893, 1987. Yokoyama, M. M. and Fujimoto, K., Role of lymphocyte activation by substance P in rheumatoid arthritis, Int. J. Tissue React., 12, 1, 1990. O'Bryne, E. M., Blancuzzi, V. J., Wilson, D. E., Wong, M., Peppard, J., Simke, J. P., and Jeng, A. Y., Increased intra-articular substance P and prostaglandin E2 following injection of interleukin-1 in rabbits, Int. J. Tissue React., 12, 11, 1990. Lam, F. Y. and Ferrell, W. R., Inhibition of carrageenan-induced joint inflammation by substnace P antagonist, Ann. Rheum. Dis., 48, 928, 1989. Lam, F. Y. and Ferrell, W. R., Capsaicin suppresses substance P-induced joint inflammation in the rat, Neurosci. Lett., 105, 155, 1989. Olgart, L., Gazelius, B., Brodin, E., and Pernow, B., Localization of substance P-like immunoreactivity in nerves in the tooth pulp, Pain, 4, 153, 1977. Olgart, L., Gazelius, B., Brodin, E., and Nilsson, G., Release of substance P-like immunoreactivity from the dental pulp, Acta Physiol. Scand., 101, 510, 1977.

216. Wakisaka,

S., Nishikawa, S., Ichikawa,H.,

Matsuo, S., Takano, Y., and Akai, M., The distribution and origin of substance P-like immunoreactivity in the rat molar pulp and periodontal tissues, Arch. Oral Biol., 30, 813, 1985. 217.

Wakisaka, S., Ichikawa, H., Nishikawa, S., Uchiyama, T., Matsuo, S., Takano, Y., and Akai, M., Immunohistochemical study on regeneration of substance P-like immunoreactivity in rat molar pulp and periodontal ligament following resection of the inferior alveolar nerve, Arch. Oral Biol., 32, 225, 1987.

218.

Wakisaka, S., Ichikawa, H., Nishikawa, S., Matsuo, Y., Takano, Y., and Akai, M., Immu-

nohistochemical observation on the correlation between substance P- and vasoactive intestinal polypeptide-like immunoreactivities in the feline dental pulp, Arch. Oral Biol., 32, 449, 1987. 219. Davidovitch, Z., Nicolay, O. F., Ngan, P. W., and Shanfeld, J. L., Neurotransmitters, cytokines, and the control of alveolar bone remodeling in orthodontics, Dent. Clin. N. Am., 31, 411, 1988. 220. Said, S. I. and Mutt, V., Polypeptide with broad biological activity: isolation from small intestine, Science, 169, 1217, 1970. 221. Hohmann, E., Levine, L., and Tashjian, A. H., Jr., Vasoactive intestinal peptide stimulates bone resorption via a cyclic adenosine 3',5'-monophosphate-

dependent mechanism, Endocrinology, 112, 1233, 1983.

222.

Hohmann, E. L. and Tashjian, A. H., Jr., Functional receptors for vasoactive intestinal peptide of human osteosarcoma cells, Endocrinology, 114, 1321,

223.

Hohmann, E. L., Elde, R. P., Rysavy, J. A., Einzig, S., and Gebhard, R. L., Innervation of

1984.


periosteum and bone by sympathetic vasoactive intestinal peptide-containing nerve fibers, Science, 232,

868, 1986. Herness, M. S., Vasoactive intestinal peptide-like immunoreactivity in rodent taste cells, Neuroscience, 33, 411, 1989. 225. Motakef, M., Shanfeld, J., and Davidovitch, Z.,

224.

Localization of VIP at bone resorption sites in vivo, J. Dent. Res., 69, 253, 1990. 226. Rosenfeld, M. G., Mermod, J.-J., Amara, S. G., Swanson, L. W., Sawchenko, P. E., Rivier, J., Vale, W. W., and Evans, R. M., Production of a novel neuropeptide encoded by the calcitonin gene via tissue-specific RNA processing, Nature, 304, 129, 1983. 227. Stjarne, P., Lundblad, L., Angard, A., Hokfelt, T., and Lundberg, J. M., Tachykinins and calcitonin gene-related peptide: co-existence in sensory nerves of the nasal mucosa and effects on blood flow, Cell Tissue Res., 256, 439, 1989. 228. Fujimori, A., Saito, A., Kimura, S., Watanabe, T., Uchiyama, Y., Kawasaki, H., and Goto, K., Neurogenic vasodilation and release of calcitonin generelated peptide (CGRP) from perivascular nerves in the rat mesenteric artery, Biochem. Biophys. Res. Commun., 165, 1391, 1989. 229. Larsson, J., Ekblom, A., Henriksson, K., Lundeberg, T., and Theordorsson, E., Immunoreactive tachykinins, calcitonin gene-related peptide and neuropeptide Y in human synovial fluid from inflamed knee joints, Neurosci. Lett., 100, 326, 1989. 230. Silverman, J. D. and Kruger, L., Calcitonin-generelated-peptide immunoreactive innervation of the rat head with emphasis on specialized sensory structures, J. Comp. Neurol., 280, 303, 1989. 231. Wakisaka, S., Ichikawa, H., Nishikawa, S., Matsuo, S., Takano, Y., and Akai, M., The distribution and origin of calcitonin gene-related peptide-containing nerve fibers in feline dental pulp, Histochemistry, 86, 585, 1987. 232. Taylor, P. E., Byers, M. R., and Redd, P. E., Sprouting of CGRP nerve fibers in response to dentin injury in rat molars, Brain Res., 461, 371, 1988. 233. Kata, J., Ichikawa, H., Wakisaka, S., Matsuo, S., Sakuda, M., and Akai, M., The distribution of vasoactive intestinal polypeptides and calcitonin generelated peptide in the periodontal ligament of mouse molar teeth, Arch. Oral Biol., 35, 63, 1990. 234. Hill, E. L. and Elde, R., Calcitonin gene-related peptide-immunoreactive nerve fibers in mandibular periosteum of rat: evidence for primary afferent origin, Neurosci. Lett., 85, 172, 1988. 235. Gazelius, B., Edwall, B., Olgart, L., Lundberg, J. M., Hokfelt, T., and Fischer, J. A., Vasodilatory effects and coexistence of calcitonin gene-related peptide (CGRP) and substance P in sensory nerves of cat dental pulp, Acta Physiol. Scand., 130, 33, 1987. 236. Takahashi, O., Traub, R. J., and Ruda, M. A.,

Demonstration of calcitonin gene-related peptide immunoreactive axons containing dynorphin A (1-8) immunoreactive spinal neurons in a rat model of peripheral inflammation and hyperalgesia, Brain Res., 475, 168, 1988. 237. Kvinnsland, I. and Kvinnsland, S., Changes in CGRP-immunoreactive nerve fibers during experimental tooth movement in rats, Eur. J. Orthodont., 12, 320, 1990. 238. Okamoto, Y., Davidovitch, Z., and Shanfeld, J., CGRP in dental pulp and periodontal ligament during tooth movement (abstract), J. Dent. Res., submitted. 239. Tashiro, T., Takahashi, O., Satoda, T., Matsushima, R., and Mizuno, N., Immunohistochemical demonstration of coexistence of enkephalinand substance P-like immunoreactivities in axonal components in the lumbar segments of cat spinal cord, Brain Res., 424, 391, 1987. 240. Jessell, T. M. and Iversen, L. L., Opiate analgesics inhibit substance P release in rat trigeminal nucleus, Nature, 268, 549, 1977. 241. Walker, J. A., Tanzer, F. S., Harris, E. F., Wakelyn, C., and Desiderio, D. M., The enkephalin response in human tooth pulp to orthodontic force, Am. J. Orthodont. Dentofacial Orthoped., 92, 9, 1987. 242. Tanzer, F. S., Tolun, E., Fridland, G. H., Dass, C., Killmar, J., Tinsley, P. W., and Desiderio, D. M., Methionine-enkephalin peptides in human teeth, Int. J. Peptide Protein Res., 32, 117, 1988. 243. Robinson, Q. C., Killmar, J. T., Desiderio, D. M., Harris, E. F., and Fridland, G., Immunoreactive evidence of beta-endorphin and methionine-arg-glyleu in human tooth pulp, Life Sci., 45, 987, 1989. 244. Hanazawa, S., Ohmori, Y., Amano, S., Miyoshi, T., Kumegawa, M., and Kitano, S., Spontaneous production of interleukin-1-like cytokine from mouse osteoblastic cell line (MC3T3-E1), Biochem. Biophys. Res. Commun., 131, 774, 1985. 245. Hanazawa, S., Amano, S., Nakada, K., Ohmori, Y., Miyoshi, T., Hirose, K., and Kitano, S., Biological characterization of interleukin-1-like cytokine produced by cultured bone cells from newborn mouse calvaria, Calcified Tissue Int., 41, 31, 1987. 246. Rath, N. C., Oronsky, A. L., and Kerwar, S. S., Synthesis of interleukin-1-like activity by normal rat chondrocytes in culture, Clin. Immunol. Immunopathol., 47, 39, 1988. 247. Kaplan, E., Dinarello, C. A., and Gelfand, J. A., Interleukin-1 and the response to injury, Immunol. Res., 8, 118, 1989. 248. Gowen, M., Wood, D. D., and Russell, G. G., Stimulation of the proliferation of human bone cells in vitro by human monocyte products with interleukin-1 activity, J. Clin. Invest., 75, 1223, 1985. 249. Boyce, B. F., Yates, A. J. P., and Mundy, G. R., Bolus injections of recombinant human interleukin1 cause transient hypocalcemia in normal mice, Endocrinology, 125, 2780, 1989.


Boyce, B., Aufdemorte, T., Bonewald, L., and Mundy, G. R., Infusions of

250. Sabatini, M.,

recombinant human interleukins la and 1p cause hypercalcemia in normal mice, Proc. Natl. Acad. Sci. U.S.A., 85, 5235, 1988. 251. Feige, U., Karbowski, A., Rordorf-Adam, C., and Pataki, A., Arthritis induced by continuous infusion of hr-interleukin-la into the rabbit knee joint, Int. J. Tissue React., 11, 225, 1989. 252. Mizel, S. B., Dayer, J. M., Krane, S. M., and Mergenhagen, S. E., Stimulation of rheumatoid synovial cell collagenase and prostaglandin production by partially purified lymphocyte-activating factor (interleukin 1), Proc. Natl. Acad. Sci. U.S.A., 78, 2474, 1981. 253. Rafter, G. W., Interleukin 1 and rheumatoid arthritis, Med. Hypoth., 27, 221, 1988. 254. Johnson, W. J., Breton, J., Newman-Tarr, T., Connor, J. R., Meunier, P. C., and Dalton, B. J., Interleukin-1 release by rat synovial cells is dependent on sequential treatment with r-interferon and lipopolysaccharide, Arthr. Rheum., 33, 261, 1990. 255. Masada, M. P., Persson, R., Kenney, J. S., Lee, S. W., Page, R. C., and Allison, A. C., Measurement of interleukin-la and - 1 in gingival cervicular fluid: implications for the pathogenesis of periodontal disease, J. Periodont. Res., 25, 156, 1990. 256. Thomson, B. M., Saklatvala, J., and Chambers, T. J., Osteoblasts mediate interleukin 1 stimulation of bone resorption by rat osteoclasts, J. Exp. Med., 150, 104, 1986. 257. Dewhirst, F. E., Ago, J. M., Peros, W. J., and Stashenko, P., Synergism between parathyroid hormone and interleukin 1 in stimulating bone resorption in organ culture, J. Bone Min. Res., 2, 127, 1987. 258. Gowen, M. and Mundy, G. R., Actions of recombinant interleukin 1, interleukin 2 and interferongamma on bone resorption in vitro, J. Immunol., 136, 2478, 1986. 259. Tatakis, D. N., Schneebergy, G., and Dziak, R., Recombinant interleukin-l stimulates prostaglandin E2 production by osteoblastic cells: synergy with parathyroid hormone, Calcified Tissue Int., 42, 358, 1988. 260. Ries, W. L., Seeds, M. C., and Key, L. L., Interleukin-2 stimulates osteoclastic activity: increased acid production and radioactive calcium release, J. Periodont. Res., 24, 242, 1989. 261. Davidovitch, Z., Lynch, P., and Shanfeld, J. L., Interleukin 2 localization in the mechanically-stressed periodontal ligament, J. Dent. Res., 68, 332, 1989. 262. Nieto-Sampedro, M. and Chandy, K. G., Interleukin-2-like activity in injured rat brain, Neurochem. Res., 12, 723, 1987. 263. Bevilacqua, M. P., Pober, J. S., Majeau, G. R., Fiers, W., Cotran, R. S., and Gimbrone, Jr., Recombinant tumor necrosis factor induces procoagulant activity in cultured human vascular endothelium: characterization and comparison with the actions of

interleukin 1, Proc. Natl. Acad. Sci. U.S.A., 83, 4533, 1986. 264. Gamble, J. R., Harlan, J. M., Klebanoff, S. J., and Vadas, M. A., Stimulation of the adherence of

neutrophils to umbilical vein endothelium by human recombinant tumor necrosis factor, Proc. Natl. Acad. Sci. U.S.A., 82, 8667, 1985. 265. Bertolini, D. R., Nedwin, G. E., Bringman, T. S., Smith, D. D., and Mundy, G. R., Stimulation of bone resorption and inhibition of bone formation in vitro by human tumor necrosis factors, Nature, 319, 516, 1986. 266. Delaisse, J.-M., Eeckhout, Y., and Vaes, G., Boneresorbing agents affect the production and distribution of procollagenase as well as the activity of collagenase in bone tissue, Endocrinology, 123, 264, 1988. 267. Yoshihara, R., Shiozawa, S., Imai, Y., and Fujita, T., Tumor necrosis factor a and interferon T inhibit proliferation and alkaline phosphatase activity of human osteoblastic SaOS-2 cell line, Lymphokine Res., 9, 59, 1990. 268. Shapiro, S., Tatakis, D. N., and Dziak, R., Effect of tumor necrosis factor alpha on parathyroid hormone-induced increase in osteoblastic cell cyclic AMP, Calcified Tissue Int., 46, 60, 1990. 269. Stashenko, P., Dewhirst, F. E., Peros, W. J., Kent, R. L., and Ago, J. M., Synergistic interactions between interleukin 1, tumor necrosis factor, and lymphotoxin in bone resorption, J. Immunol., 138, 1464, 1987. 270. Pfeilschifter, J., Chenu, C., Bird, A., Mundy, G. R., and Roodman, G. D., Interleukin-1 and tumor necrosis factor stimulate the formation of human osteoclast-like cells in vitro, J. Bone Min. Res., 4, 113, 1989. 271. Hashimoto, J., Yoshikawa, H., Takaoka, K., Shimizu, N., Mashuhara, K., Tsuda, T.,

Miyamoto, S., and Ono, K., Inhibitory effects of tumor necrosis factor alpha on fracture healing in rats, Bone, 10, 453, 1989. 272.

Johnson, R. A., Boyce, B. F., Mundy, G. R., and Roodman, G. D., Tumors producing human tumor necrosis factor induce hypercalcemia and osteoclastic bone resorption in nude mice, Endocrinology, 124, 1424, 1989.

Hopkins, S. J. and Meager, A., Cytokines in synovial fluid. II. The presence of tumor necrosis factor and interferon, Clin. Exp. Immunol., 73, 88, 1988. 274. Maury, C. P. J. and Teppo, A.-M., Cachectin/ tumor necrosis factor-a in the circulation of patients 273.

with rheumatic disease, Int. J. Tissue React., 11, 189, 1989. 275. Yocum, D. E., Esparza, L., Dubry, S., Benjamin, J. B., Volz, R., and Scuder, P., Characteristics of tumor necrosis factor production in rheumatoid arthritis, Cell Immunol., 122, 131, 1989. 276. Gowen, M., Nedwin, G. E., and Mundy, G. R., Preferential inhibition of cytokine-stimulated bone re-


277.

278.

279.

280.

281. 282.

283.

sorption by recombinant interferon gamma, J. Bone Min. Res., 1, 469, 1986. Gowen, M., MacDonald, B. R., and Russell, G., Actions ofrecombinant human T-interferon and tumor necrosis factor a on the proliferation and osteoblastic characteristics of human trabecular bone cells in vitro, Arthr. Rheum., 31, 1500, 1988. Nanes, M. S., McKoy, W. M., and Marx, S. J., Inhibitory effects of tumor necrosis factor-a and interferon-T on deoxyribonucleic acid and collagen synthesis by rat osteosarcoma cells (ROS 17/2.8), Endocrinology, 124, 339, 1989. Vignery, A., Niven-Fairchild, T., and Shepard, M. H., Recombinant murine interferon-r inhibits the fusion of mouse alveolar macrophages in vitro, but stimulates the formation of osteoclast-like cells in implanted syngeneic bone particles in mice in vivo, J. Bone Min. Res., 5, 637, 1990. Melin, M., Hartmann, D. J., Magloire, H., Falcoffe, E., Auriault, C., and Grimaud, J. A., Human recombinant gamma-interferon stimulates proliferation and inhibits collagen and fibronectin production by human dental pulp fibroblasts, Cell Mol. Biol., 35, 97, 1989. Judis, G., Davidovitch, Z., and Shanfeld, J., Identification of IFN at in vivo bone resorption sites, J. Dent. Res., 69, 244, 1990. Rosol, T. J., Shanfeld, J. L., and Davidovitch, Z., Biochemical aspects of orthodontic tooth movement. II. Effects of neurotransmitters and cytokines on bone resorption and formation in vitro, J. Dent. Res., submitted. Saito, S., Ngan, P., Saito, M., Kim, K., Lanese, R., Shanfeld, J., and Davidovitch, Z., Effects of cytokines on prostaglandin E and cAMP levels in human periodontal ligament fiborblasts in vitro, Arch.

Oral Biol., 35, 387, 1990. Saito, S., Ngan, P., Saito, M., Lanese, R., Shanfeld, J., and Davidovitch, Z., Interactive effects between cytokines on PGE production by human periodontal ligament fibroblasts, in vitro, J. Dent. Res., 69, 1456, 1990. 285. Saito, S., Rosol, T. J., Saito, M., Ngan, P. W., Shanfeld, J., and Davidovitch, Z., Bone-resorbing activity and prostaglandin E produced by human per-

284.

iodontal

ligament cells in vitro, J. Bone Min. Res.,

5, 1013, 1990. 286. Saito, S., Saito, M., Ngan, P., Lanese, R., Shanfeld, J., and Davidovitch, Z., Effects of para-

thyroid hormone and cytokines on prostaglandin E synthesis and bone resorption by human periodontal ligament fibroblasts, Arch. Oral Biol., in press. 287. Saito, S., Saito, M., Ngan, P. W., Shanfeld, J., and Davidovitch, Z., Interleukin 1 beta and prostaglandin E are involved in the response of periodontal cells to mechanical stress in vivo and in vitro, Am. J. Orthodont. Dentofacial Orthoped., in press.


Prediction of orthodontic tooth movement.

Tooth eruption and orthodontic movement.

Osteoimmunology in orthodontic tooth movement.

Comment on differential tooth movement.

Differential tooth movement in "uphill cases".

Three-dimensional analysis of orthodontic tooth movement.

Adiponectin prevents orthodontic tooth movement in rats.

Mechanical and thermal hypersensitivities associated with orthodontic tooth movement: a behavioral rat model for orthodontic tooth movement-induced pain.

Numerical analysis of tooth movement in different plans of transparent tooth correction therapies.

Initial orthodontic tooth movement of a multirooted tooth: a 3D study of a rat molar.

Corticotomies and Orthodontic Tooth Movement: A Systematic Review.

The effect of caffeine on orthodontic tooth movement in rats.

Osteocyte death during orthodontic tooth movement in mice.

Computerized Analysis of Digital Photographs for Evaluation of Tooth Movement.

Are interventions for accelerating orthodontic tooth movement effective?

Nrf2 activation attenuates both orthodontic tooth movement and relapse.

Variables affecting orthodontic tooth movement with clear aligners.

A periodontal ligament driven remodeling algorithm for orthodontic tooth movement.

The role of hypoxia in orthodontic tooth movement.

External forces causing tooth movement: a case report.

The effect of corticosteroid-induced osteoporosis on orthodontic tooth movement.

Histomorphometric study of alveolar bone turnover in orthodontic tooth movement.

Age-related effects on osteoclastic activities after orthodontic tooth movement.

Effects of icariin on orthodontic tooth movement in rats.

Tooth movement.

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Suggest Documents

Prediction of orthodontic tooth movement.

Tooth eruption and orthodontic movement.

Osteoimmunology in orthodontic tooth movement.

Comment on differential tooth movement.

Differential tooth movement in "uphill cases".

Three-dimensional analysis of orthodontic tooth movement.

Adiponectin prevents orthodontic tooth movement in rats.

Mechanical and thermal hypersensitivities associated with orthodontic tooth movement: a behavioral rat model for orthodontic tooth movement-induced pain.

Numerical analysis of tooth movement in different plans of transparent tooth correction therapies.

Initial orthodontic tooth movement of a multirooted tooth: a 3D study of a rat molar.

Corticotomies and Orthodontic Tooth Movement: A Systematic Review.

The effect of caffeine on orthodontic tooth movement in rats.

Osteocyte death during orthodontic tooth movement in mice.

Computerized Analysis of Digital Photographs for Evaluation of Tooth Movement.

Are interventions for accelerating orthodontic tooth movement effective?

Nrf2 activation attenuates both orthodontic tooth movement and relapse.

Variables affecting orthodontic tooth movement with clear aligners.

A periodontal ligament driven remodeling algorithm for orthodontic tooth movement.

The role of hypoxia in orthodontic tooth movement.

External forces causing tooth movement: a case report.

The effect of corticosteroid-induced osteoporosis on orthodontic tooth movement.

Histomorphometric study of alveolar bone turnover in orthodontic tooth movement.

Age-related effects on osteoclastic activities after orthodontic tooth movement.

Effects of icariin on orthodontic tooth movement in rats.

Tooth movement.

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Follow Us