Histological Assessment of Autogenous Bone Graft A Case
Report and Critical Evaluation
by
walled infrabony pocket. The lower right first and second bicuspids were missing, providing a large edentulous zone adjacent to the defect. The gingival tissues were chronically inflamed and exhibited hyperplastic and fibrotic changes. Some minor gingival recession was present in the area and suppuration was observed at the initial preoperative examination. There was negligible mobility of the teeth in the region. The oral hygiene
regime was exemplary. The preoperative radiograph
Bernard S. Moskow* Fraya Karsh* Stephen D. Stein* The development of the ability to restore periodontium lost as a result of periodontal disease is a concept which is attractive to the dental profession. In attempts to achieve this objective, various types of bone grafts have been employed in periodontal therapy.1"11 Such grafts generally have been recommended for use in intraosseous defects not amenable to other methods of treatment, but Dragoo and Sullivan have demonstrated apparent success with iliac crest grafts in supracrestal situations.12 Clinical evidence of the success of these procedures has been documented routinely following surgical reentry and the demonstration of newly formed bone in the treated region.6'8'10 Several reports, which included histologie evaluation of bone graft sites, have suggested that bone grafts have the potential to form a totally new attachment apparatus including the deposition of cementum on a previously denuded root surface and the forThe mation of a functional periodontal difficult from clinof material is procurement histologie ically successful graft sites. In addition, because histological observations on this material are often made on a few isolated sections, the interpretation is largely on a two dimensional plane. Serial sectioning permits more accurate interpretation in three dimensions, although there is always the everpresent possibility of errors in interpretation. The following single case report adds some further microscopic observations on the fate of the free osseous tissue autografi.
of the area (Fig. 2A) confirmed the clinical presence of a shallow interdental crater between the mandibular right first and second molars and a deep infraosseous lesion on the mesial of the first molar. Thickening of the periodontal ligament space and areas of cementai résorption could also be noted on the x-rays, as could thinning of the interradicular bone of the first molar. Therapy. After initial preparation, surgical exposure of the area was accomplished with buccal and lingual muco-periosteal flaps extending from the cuspid to the
ligament.12"16
Case
report
Clinical Examination. The patient, a 37-year-old male clinic patient, with a noncontributory medical background presented with a broad infrabony lesion on the mesial of the mandibular right first molar. The pocket measured 11 mm at the mesio-buccal line angle of this tooth and 5 mm on the buccal and lingual surfaces, and there was an early bifurcation involvement on the buccal (Fig. 1A). The orifice of the infraosseous defect was wide and revealed a buccal wall at its most cervical height and a lingual wall at a more apical level. Strictly speaking, this then, would be considered a one-, two- and three*
Department of Periodontics, Columbia University School of DenSurgery, 630 West 168th St., New York, NY 10032.
tal and Oral
Figure 1. Clinical photographs of management of infraosseous defect with autogenous bone graft. A. Preoperative clinical view of lesion to be treated. Note the extent ofpocketing on the mesial aspect of the mandibular first molar. B. Exposure of area with full thicknessflap revealing deep, wide infrabony lesion. The graft was obtained from the adjacent edentulous ridge. C. Surgical exposure of treated area 28 weeksfollowing autografi. The defect appears to be filled with newly formed bone. D. Probing confirms obliteration of original defect. E. Surgical block section including treated lesion obtained. F. Closure of surgical site following removal of block section (lingual view).
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Radiographs of various stages of treatment of lesion. Preoperative x-ray. Note the extent and depth of the infraosseous lesion on the mesial of the first molar. B. Apparent healing of lesion 9 weeksfollowing autogenous bone graft. C. Radiograph taken 22 weeks following autografi. Endodontic therapy performed on distal root in anticipation of hemisection of tooth and removal of block section. Figure 2. A.
Figure 1
region of the second molar. The surgical site was debrided of all soft tissue tabs and the contents of the infrabony lesion thoroughly curetted out. The tooth surface was lightly scaled, but no effort was made to remove
Volume 50 Number 6
X-ray of gross surgical specimen. B. Photograph of specimen showing mesial root offirst molar with adjacent
Figure 3 A.
gross
periodontal tissues.
all of the cementum from the denuded root surface. The
opening of the infrabony lesion was wide and contained
and two walls except for its very base which was three-walled (Fig. IB). Small particles of cancellous alveolar bone were obtained from the adjacent edentulous ridge and loosely packed into the intraosseous defect. The operative area was closed with 000 silk sutures and packed with a noneugenol surgical dressing. The sutures were removed after 7 days and after the second postoperative week, the final dressing was discarded. Healing of the grafted surgical site progressed uneventfully. Evaluation of Clinical Results. There was no evidence of graft rejection after 9 weeks and the clinical picture appeared somewhat improved. Radiographic examination revealed increased opacity within the defect, and a diminishing of the funneling effect on the x-ray was noted (Fig. 2B). At 22 weeks after the operation, it was decided that the continued questionable nature of the first molar precluded including this tooth in a fixed bridge, and it was decided to hemisect the tooth and one
Histology ofAutogenous Bone Graft
293
retain the distal root as one abutment. Accordingly, endodontic therapy was performed on the distal root (Fig. 2C). It was decided to then take a block surgical section including adjacent periodontal tissues, along with the mesial root, in order that results of the bone autografi could be histologically examined and evaluated. Some 28 weeks after the original treatment, the area was again exposed by means of buccal and lingual flaps extending from the second molar to the canine tooth. The grafted area was filled with new bone (Fig. 1C) which clinically appeared to be continuous with the adjacent alveolar process. The new bone felt hard to touch and was resistant to probing (Fig. 1D). The tooth was hemisected using high speed rotary instruments. The mesial root and a generous segment of periodontium surrounding it, including the graft site, were then removed (Fig. IE). The flaps were coapted using 000 surgical silk, but the socket of the extracted root was left open (Fig. IF). The surgical specimen (Fig. 3) was fixed in 10% buffered formalin, decalcified in a weak solution of nitric acid, embedded in paraffin, and prepared for histologie examination in the usual manner. Serial sections 5 µ thick were cut starting from the buccal line angle and proceeding towards the lingual surface. Specimens for study were stained with hematoxylin and eosin and the Bloom variation of the Mallory stain. Histologie Evaluation. Under low power magnification the bone comprising the proximal wall of the original osseous periodontal defect could be distinguished easily from the grafted bone by virtue of the organized and regular lamellar pattern (Figs. 4A and 5A). The marrow spaces were of varying sizes, but the general character of
294
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Moskow, Karsh, Stein
Figure 4. Low power photomicrograph of site of osseous autografi. Multiple particles of grafted bone can be seen within the former of the host site bone are defect and a small island of newly forming bone is present at the base of the lesion. Some of the marrow spaces FM new BG bone NB cells. with chronic bone, are some and fibrous marrow, IC inflammatory graft, highly infiltrated fibrosed inflammatory cells, Ce cementum, A separation artifact, (H & E, original magnification, x 25). B. Higher magnification of graft site seen in Figure 4A. Active Osteogenesis can be noted about the particles ofgraft bone. The area is infiltrated with chronic inflammatory cells. Note the intraepithelial tear in the junctional epithelium which is associated with the separation artifact (Original magnification, =
=
=
x
=
=
=
63).
the alveolar bone from the edentulous zone as seen on the microscopic sections could be regarded as dense. Some remnants of fat were contained within the marrow cavities, but for the most part the marrow spaces were fibrous in nature with a substantial infiltration of chronic inflammatory cells. An intense lymphocytic infiltrate was present in one large marrow space (Fig. 4A). The surface of the host bone at the graft interface was covered by a thin layer of osteoid and osteoblasts, indicating active Osteogenesis in this area. Numerous particles of grafted bone were readily visible within the fibrous connective tissue contained between the lamellar bone and the tooth (Figs. 4 and 5). The transplanted chips were devoid of osteocytes, but woven bone containing viable osteocytes was seen on their surfaces (Fig. 4B). Osteoblastic activity and ingrowths of new capillaries were associated with all of the particles of grafted bone. The connective tissue bed was infiltrated with inflammatory cells, particularly in the cervical one-half of the grafted zone (Fig. 4B). At the base of the original osseous defect, a small island of new bone which seemed unrelated to the implanted bone was
present. A slender trabecula of new bone also was evident
the tooth at a more coronal level (Figs. 4A An artifactual space was present between the root surface and the autografi (Figs. 4-6). Associated with this void was a thin layer of epithelial cells which
adjacent and
to
6A).
adherent to both the cementai surface and the wall containing the grafted bone chips (Figs. 4-6). Despite the thinness of the epithelium (two to three cells wide), the separation artifact was seen to be intraepithelial in nature. The epithelial cells were of the squamous type, and their nuclei were flattened and orientated parallel to the root surface as is generally the case in junctional epithelium (Fig. 6). Examination of serial sections revealed that thicker plaques of squamous epithelium were present at various intervals along the thin trail of downgrowing epithelium and that a continuity existed between the long epithelial attachment and the sulcular epithelium. At the marginal aspect of the defect, two small fragments of nonvital graft bone were encapsulated by were
epithelial ingrowths (Fig. 7). Changes in the root surface suggestive of previous aggressive root planing were not prominent. A distinct
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Histology ofAutogenous Bone Graft
Number 6
295
Figure 5A. Low power photomicrograph of serially cut specimen of bone graft site showing active apposition of new bone on to transplanted particles of bone. Separation artifact resulting from histologie processing and epithelial downgrowth along root surface can be noted. BG bone graft, FM fibrous marrow, A separation artifact, Ce cementum, Pdl periodontal ligament, (BloomMallory stain, original magnification, x 25). B. High power magnification of microscopic field seen in lower right corner of Figure 5A. Particle of grafted bone is displaced into intact periodontal ligament and is apparently encapsulated. Evidence of root instrumentation and epithelial downgrowth is clearly seen (Original magnification, x 63). =
=
=
layer of pre-existing cementum was evident, but newlydeposited cementum could not be detected generally. The root surface lying in the base of the defect proved an exception to this observation. Here, evidences of a completely new attachment could be observed. This included recognizable newly-deposited cementum, apposition of new bone lamellae and a functionally-oriented periodontal ligament (Fig. 8). Sharpey's fibers were easily visible, and cementoblasts were lined up at regular intervals adjacent to the fiber insertion. Within the newly formed periodontal ligament, small spicules of nonvital bone, presumably grafted chips, were undergoing résorption (Fig. 9). This newly formed region of periodontium was apical to and remote from the bulk of the bony autografi. Discussion
widespread conviction that various types of bone grafts may have the capacity to induce the formation of an entirely new periodontium requires further proof and clarification, particularly in light of the known biologic behavior of periodontal tissues.3,6· "'12 Bone marrow transplants and other types of intraoral The
=
=
bone autografts provide a ready source of available material for stimulating Osteogenesis in periodontal defects; however, the concept that such grafts also have an inherent capacity for laying down new cementum on a previously denuded root surface and creating a functional periodontal ligament has yet to be demonstrated unequivocally. Animal experiments in which surgically created or artificially-provoked periodontal defects are used for bone implant studies are not ideal to test this thesis, as it has been shown that such lesions have an inherent ability for spontaneous regeneration of lost periodontium.17"19 The histologie study of human block sections is also subject to difficulties in interpretation arising out of orientation and plane of section and the frequent lack of serial sections.11,12,14"1B Also, sometimes it is difficult to establish whether the healing seen in bone implant sites is associated with the inherent capacity of certain types of periodontal lesions to regenerate spontaneously, or whether, in fact, it is stimulated by the bone graft itself. Melcher has described the shortcomings of the concept of utilizing bone grafts in attempts to regenerate periodontium.20, 21 Bone is only one of the connective tissues
296
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Photomicrograph of region slightly apical to the graft site. A plaque of epithelium is clearly seen along the root surface and separation artifact intervenes between the intraepithelial split. A slender trabecula of new bone can be seen adjacent to the junctional epithelium, and a larger island of newlyforming bone is present at a more apical level. An area of new attachment exists at the lower portion of the microscopic field. BG bone graft, NB new bone, A separation artifact, Ce cementum, NCE new cementum, (H & E, original magnification, x 63). B. Higher magnification ofjunctional epithelial plaque seen on Figure 6A. Note new cementum apposition apical to the epithelial downgrowth (Original magnification, x 160). Figure 6 A. a
=
=
of the periodontium that provides a continually adapting mechanism for support and function of the teeth. A second is the periodontal ligament. This tissue attaches the tooth to the alveolar process of the jaws and contains cells that have the capacity to deposit and resorb bone, cementum, and the soft connective tissue of the ligament itself. A homeostatic mechanism that maintains the periodontal space throughout life apparently operates within the periodontium. It has been suggested that this mechanism is at least partially mediated by the ability of the Based periodontal ligament cells to inhibit on our known understanding of the physiology of the periodontium, it would be reasonable to assume that the synthesis and deposition of periodontal ligament, alveolar bone and cementum are largely functions of the cells of the periodontal ligament and the endosteal cells of the bone, and that maintenance of the alveolar bone is dependent upon the function of the tooth.23 The functional stimuli required for maintenance of the alveolar bone are unlikely to be transmitted unless an intact periodontal ligament provides continuity between the tooth and bone. Based on the above understanding, it does not seem unreasonable to set as a primary objective
Osteogenesis.22
=
=
=
regeneration of the periodontal ligament rather than regeneration of bone. As stated by Melcher, this concept rests on the premise that cells of regenerated periodontal ligament have the capacity to synthesize and deposit the extracellular substance of bone, cementum and periodontal ligament and thereby restore a functioning periodontium.21
The presence of new bone that was found on surgical reentry was substantiated by histologie examination. Trabeculae of new bone were consistently seen about particles of the implanted bone, some of which exhibited recognizable areas of résorption (Fig. 4). After 28 weeks, the implant material was readily discernible microscopically but was easily distinguishable from the adjacent wall of cancellous bone, although it appeared to be continuous with it (Figs. 4 and 5). Despite the careful closure of the wound, and the lack of clinical evidence of sequestration of graft particles, downgrowth of a thin layer of epithelial cells was seen. These cells were typical of junctional epithelium and formed a delicate network extending to the base of the defect, but coronal to the area of new attachment (Fig. 6). A detailed study of serial sections confirmed the presence of epithelium
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Histology of Autogenous Bone Graft
297
SBSggM 1
BG
s':
·'
-.'^
wir Figure 7. Marginal region of autografi site depicting epithelial invagination and encapsulation of bone graft particles 28 weeks following bone transplantation. Adjacent bone graft particles show apposition of new bone. BG bone graft, Ep epithelium, SEp sulcular epithelium, Ce cementum, (H & E, original magnification, x 63). =
=
=
=
along the side of the root, although in most instances it was only one or two cells thick. Thicker plaques of epithelium were noted occasionally, and at the orifice of the defect there was evidence of epithelium encompassing small particles of graft bone, which can be anticipated ultimately as being rejected (Fig. 7). As deeper sections through the surgical site into the periphery of the graft area were made, true evidence of a regenerated attachment could be noted (Fig. 8). While evidence of grafted
found close to the base of the defect, we observations to suggest that the regenerainterpret tion in the basal area of the periodontium represents spontaneous healing unrelated to the graft. We propose that microscopic evidence of graft bone continues to be present for months or perhaps years and becomes a permanent record of the biologic activity of the graft bone
was
our
Unless a grafted periodontal wound provides evidence of attachment of periodontal ligament fibers to bone deposited on recognizable bone graft particles and to new cementum at the same level on the root, it seems unlikely that any claim to the contribution of the graft to the regeneration of the periodontium can be made. In this regard, it is relevant to note that Frank have noted deposition of cemenet al.24 and tum to be most advanced in the apical aspects of ungrafted periodontal wounds in man and monkey respec-
region. some
Listgarten25
tively.
The separation of the newly deposited cementum from the root surface seen on the microscopic sections, showing an area of regeneration (Fig. 8), was undoubtedly artifactual. Similar artifactitious splits frequently have been noted in histologie material of healing periodontal
298
3. Periodontol. June, 1979
Moskow, Karsh, Stein
11
#
·
·&
>·.
vi
Figure 8. « of new attachment apical to and apparently unrelated to bone graft. Note laying down of new cementum on previously instrumented root surface and apposition of new alveolar bone proper. The periodontal ligament fibers appear to be functionally orientated. A separation artifact is present at the interface between the root surface and the new layer of cellular cementum. De dentin, NCe new cementum, Nb new bone, A separation artifact, Pdl periodontal ligament, (H & E, original magnification, X 63). =
=
wounds,1
=
=
and in all probability result from differential shrinkage in processing.20 It is doubtful whether this type of artifact has any clinical implication in reattachment therapy as suggested by Hawley and Miller.16 The epithelial downgrowth with concurrent osseous repair reported in this human material is strikingly similar to the case described by Caton and Zander in a provoked periodontal lesion in a Rhesus monkey.28 Complete healing and "fill" of an infrabony pocket had occurred; however, histologie sections showed a long junctional epithelial attachment extending to the most apical level of root instrumentation. Clinically, and in every other manner, this appeared to be a successfully treated lesion and indicates that bone remodeling can occur without connective tissue attachment to the root. The long term implication associated with a long
=
to bone does not appear be favorable, as the cellular potential to form new cementum (i.e. the periodontal ligament) is physically and functionally remote from the area in question. It seems more likely that this type of attachment would be weaker than a connective tissue attachment and would be vulnerable to future ingrowths of epithelium and eventual pocket formation. The observations made in this study suggest that the procedure of surgical reentry following bone grafting to confirm regeneration of the periodontium is of little value. The clinical evidence of newly formed bone does not necessarily imply apposition of new cementum on the root nor the presence of a functional periodontal ligament. Such judgements can be made only through careful study of histologie material.
epithelial attachment adjacent to
Volume 50 Number 6
Histology ofA utogenous Bone Graft
299
Figure 9. Small particles ofgraft bone undergoing résorption in area ofperiodontal ligament apical to bulk ofgrafted bone. Note the multinucleated giant cells associated with the resorptive process. This appears to be an area of new attachment with active cementum and bone formation taking place. De dentin, Nb new bone, NCe new cementum, Pdl periodontal ligament, bg bone graft, Gc giant cells (H & E, original magnification, x 119). =
=
=
=
=
=
Summary
and
Conclusions
A histological study of the site of a clinically successful bone graft revealed the presence of a downgrowing junctional epithelium between the root surface and the altered graft bone. No new cementum could be seen associated with the osseous autografi, nor was a functional periodontal ligament in evidence adjacent to the grafted bone. It is suggested that the presence of newly formed bone on surgical reentry following bone grafting in periodontal lesions does not necessarily imply that regeneration of the periodontium has occurred. Acknowledgments
The authors are sincerely grateful for the technical assistance provided by Mr. Arthur Bloom, histopathologic technician, and Ms. Ida Nathan, medical photographer. Professors A. H. Melcher and Bernard Wasserman were kind enough to review
the are
manuscript, and their greatly appreciated.
critical comments and
suggestions
References 1. Beube, F. E., and Silvers, H. F.: Further studies on bone regeneration with the use of boiled heterogenous bone. J Periodontol 7: 17, 1936. 2. Scopp, I. W., Morgan, F. H., Dooner, J. J., Freddes, H. J., and Heyman, R. .: Bovine bone (Boplant) implant for infrabony oral lesions (Clinical trials in humans). Periodontics 4: 169, 1966. 3. Nabers, C. L., and O'Leary, T. J.: Autogenous bone transplant in the treatment of osseous defects. J Periodontol 36:
5, 1965. 4. Robinson, R. E.: Osseous coagulum for bone induction. / Periodontol 40: 503, 1969. 5. Schallhorn, R. G, Hiatt, W. H., and Boyce, W.: Iliac transplants in periodontal therapy. J Periodontol 41: 566, 1970. 6. Rosenberg, . M.: Free osseous predictable procedure. J Periodontol 42:
tissue autografts 195. 1971.
as a
300
Mosko w,
J. Periodontol. June, 1979
Karsh, Stein
7. Diem, C. R., Bowers, G. M., and Moffitt, W. C: Bone A technique for osseous implants. J Periodontol 43: 295, 1972. 8. Hiatt, W. H., and Schallhorn, R. G.: Intraoral transplants of cancellous bone and marrow in periodontal lesions. / Periodontol 44: 194, 1973. 9. Libin, B. M., Ward, H. L., and Fishman, L.: Decalcified, lyophilized bone allografts for use in human periodontal defects. / Periodontol 46: 51, 1975. 10. Mellonig, J. T., Bowers, G. M., Bright, R. W., and Lawrence, J. J.: Clinical evaluation of freezedried bone allografts in periodontal osseous defects. J Periodontol 47: 125, 1976. 11. Schallhorn, R. G.: Osseous grafts in the treatment of periodontal osseous defects. Periodontal Surgery: Biologic Basis and Technique, Springfield, Thomas, 1976. 12. Dragoo, M. R., and Sullivan, H. C: A clinical and histologie evaluation of autogenous iliac bone grafts in humans: Part I. Wound healing two to eight months. J Periodontol 44: 599, 1973. 13. Ross, S. E., and Cohen, D. W.: The fate of a free osseous tissue autografi, a clinical and histologie case report. Periodontics 6: 145, 1968. 14. Nabers, C. L., Reed, O. M., and Hamner, J. E.: Gross and histologie evaluation of an autogenous bone graft 57 months postoperatively. J Periodontol 43: 702, 1972. 15. Froum, S. J., Thaler, R., Scopp, I. W„ and Stahl, S. S.: Osseous autografts. II. Histologie responses to osseous coagulum-bone blend grafts. J Periodontol 46: 656, 1975. 16. Hawley, C. E., and Miller, J.: A histologie examination of a free osseous autograft. J Periodontol 46: 289, 1975. 17. Linghorne, W. J., and O'Connel, D. C: Studies in the regeneration and reattachment of supporting structures of the
blending:
teeth. II. Regeneration of alveolar process. J Dent Res 30: 604, 1951. 18. Linghorne, W. J., and O'Connel, D. C: Studies in the reattachment and regeneration of the supporting structures of the teeth. III. Regeneration in epithelialized pockets. J Dent Res 34: 164, 1955. 19. Ramfjord, S.: Experimental periodontal reattachment in Rhesus monkeys. J Periodontol 22: 67, 1951. 20. Melcher, A. H.: On the repair potential of periodontal tissues. J Periodontol 47: 256, 1976. 21. Melcher, A. H: Biological processes in résorption, deposition and regeneration of bone. Periodontal Surgery: Biologic Basis and Technique. Springfield, Thomas, 1976. 22. Melcher, A. H.: Repair of wounds in the periodontium of the rat. Influence of periodontal ligament on Osteogenesis. Arch Oral Biol 15: 1183, 1970. 23. Cohn, S. .: Disuse atrophy of the periodontium in mice. Arch Oral Biol 10: 909, 1965. 24. Frank, R., Fiore-Donno, G., Cimasoni, G., and Matter, J.: Ultrastructural study of epithelial and connective gingival reattachment in man. J Periodontol 45: 626, 1974. 25. Listgarten, . .: Electron microscopic study of the junction between surgically denuded root surfaces and regenerated periodontal tissues. J Periodont Res 7: 68, 1972. 26. Morris, M. L.: Healing of naturally occurring periodontal pockets about vital human teeth. J Periodontol 26: 285, 1955. 27. Jansen, M. T., Coppes, L., and Verdenius, H. H. W.: The healing of periodontal wounds in dogs. J Periodontol 26: 292, 1955. 28. Caton, J., and Zander, . .: Osseous repair of infrabony pocket without new attachment of connective tissue. J Clin Periodont 3: 54, 1976.
Abstracts Dental Treatment for Patients
with
Leukemia
Carl, W.
Quintessence Int 9: 9, August,
1978.
The oral changes of leukemia patients before and after chemotherapy and dental treatment for patients with leukemia were described. The first sign of the leukemia may appear in the mouth and usually as the disease progresses, the oral manifestations progress proportionately. By maintaining or establishing good oral hygiene in patients with leukemia, the oral manifestations of the disease can be greatly reduced. Many times, extractions have to be done, even when the patient is in an acute leukemic state and specific precautions are needed. Team work including other related departments is necessary to deal with all the complications that may develop during the course of this disease. Dr. Yau-Fwu Huang 66 Elm Street, Buffalo, NY 14263.
A Simple Filter Paper Strip Viewer Estimation
Wilson, C. ., Tay, W. M., and Morgan, Quintessence Int 9: 63, August, 1978.
for
Gingival Fluid
W. J.
The most widely used method of collecting gingival exúdate for the degree of inflammation in a gingival sulcus is to insert filter paper strips at varying depths into the gingival crevice. By application of ninhydrin reagent stains, the exúdate on the paper becomes visible and can be measured. A new design to measure the stained area of filter paper strip was described and illustrated. A 35 mm photographic slide viewer was used and Letratape strips were placed on a viewing lens with a Letratone grid. The viewer possesses the versatility of a microscope at a fraction of the cost. British Forces Dr. Yau-Fwu Huang Post Office 34, London, England.
assessing
Report and Critical Evaluation
by
walled infrabony pocket. The lower right first and second bicuspids were missing, providing a large edentulous zone adjacent to the defect. The gingival tissues were chronically inflamed and exhibited hyperplastic and fibrotic changes. Some minor gingival recession was present in the area and suppuration was observed at the initial preoperative examination. There was negligible mobility of the teeth in the region. The oral hygiene
regime was exemplary. The preoperative radiograph
Bernard S. Moskow* Fraya Karsh* Stephen D. Stein* The development of the ability to restore periodontium lost as a result of periodontal disease is a concept which is attractive to the dental profession. In attempts to achieve this objective, various types of bone grafts have been employed in periodontal therapy.1"11 Such grafts generally have been recommended for use in intraosseous defects not amenable to other methods of treatment, but Dragoo and Sullivan have demonstrated apparent success with iliac crest grafts in supracrestal situations.12 Clinical evidence of the success of these procedures has been documented routinely following surgical reentry and the demonstration of newly formed bone in the treated region.6'8'10 Several reports, which included histologie evaluation of bone graft sites, have suggested that bone grafts have the potential to form a totally new attachment apparatus including the deposition of cementum on a previously denuded root surface and the forThe mation of a functional periodontal difficult from clinof material is procurement histologie ically successful graft sites. In addition, because histological observations on this material are often made on a few isolated sections, the interpretation is largely on a two dimensional plane. Serial sectioning permits more accurate interpretation in three dimensions, although there is always the everpresent possibility of errors in interpretation. The following single case report adds some further microscopic observations on the fate of the free osseous tissue autografi.
of the area (Fig. 2A) confirmed the clinical presence of a shallow interdental crater between the mandibular right first and second molars and a deep infraosseous lesion on the mesial of the first molar. Thickening of the periodontal ligament space and areas of cementai résorption could also be noted on the x-rays, as could thinning of the interradicular bone of the first molar. Therapy. After initial preparation, surgical exposure of the area was accomplished with buccal and lingual muco-periosteal flaps extending from the cuspid to the
ligament.12"16
Case
report
Clinical Examination. The patient, a 37-year-old male clinic patient, with a noncontributory medical background presented with a broad infrabony lesion on the mesial of the mandibular right first molar. The pocket measured 11 mm at the mesio-buccal line angle of this tooth and 5 mm on the buccal and lingual surfaces, and there was an early bifurcation involvement on the buccal (Fig. 1A). The orifice of the infraosseous defect was wide and revealed a buccal wall at its most cervical height and a lingual wall at a more apical level. Strictly speaking, this then, would be considered a one-, two- and three*
Department of Periodontics, Columbia University School of DenSurgery, 630 West 168th St., New York, NY 10032.
tal and Oral
Figure 1. Clinical photographs of management of infraosseous defect with autogenous bone graft. A. Preoperative clinical view of lesion to be treated. Note the extent ofpocketing on the mesial aspect of the mandibular first molar. B. Exposure of area with full thicknessflap revealing deep, wide infrabony lesion. The graft was obtained from the adjacent edentulous ridge. C. Surgical exposure of treated area 28 weeksfollowing autografi. The defect appears to be filled with newly formed bone. D. Probing confirms obliteration of original defect. E. Surgical block section including treated lesion obtained. F. Closure of surgical site following removal of block section (lingual view).
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Radiographs of various stages of treatment of lesion. Preoperative x-ray. Note the extent and depth of the infraosseous lesion on the mesial of the first molar. B. Apparent healing of lesion 9 weeksfollowing autogenous bone graft. C. Radiograph taken 22 weeks following autografi. Endodontic therapy performed on distal root in anticipation of hemisection of tooth and removal of block section. Figure 2. A.
Figure 1
region of the second molar. The surgical site was debrided of all soft tissue tabs and the contents of the infrabony lesion thoroughly curetted out. The tooth surface was lightly scaled, but no effort was made to remove
Volume 50 Number 6
X-ray of gross surgical specimen. B. Photograph of specimen showing mesial root offirst molar with adjacent
Figure 3 A.
gross
periodontal tissues.
all of the cementum from the denuded root surface. The
opening of the infrabony lesion was wide and contained
and two walls except for its very base which was three-walled (Fig. IB). Small particles of cancellous alveolar bone were obtained from the adjacent edentulous ridge and loosely packed into the intraosseous defect. The operative area was closed with 000 silk sutures and packed with a noneugenol surgical dressing. The sutures were removed after 7 days and after the second postoperative week, the final dressing was discarded. Healing of the grafted surgical site progressed uneventfully. Evaluation of Clinical Results. There was no evidence of graft rejection after 9 weeks and the clinical picture appeared somewhat improved. Radiographic examination revealed increased opacity within the defect, and a diminishing of the funneling effect on the x-ray was noted (Fig. 2B). At 22 weeks after the operation, it was decided that the continued questionable nature of the first molar precluded including this tooth in a fixed bridge, and it was decided to hemisect the tooth and one
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retain the distal root as one abutment. Accordingly, endodontic therapy was performed on the distal root (Fig. 2C). It was decided to then take a block surgical section including adjacent periodontal tissues, along with the mesial root, in order that results of the bone autografi could be histologically examined and evaluated. Some 28 weeks after the original treatment, the area was again exposed by means of buccal and lingual flaps extending from the second molar to the canine tooth. The grafted area was filled with new bone (Fig. 1C) which clinically appeared to be continuous with the adjacent alveolar process. The new bone felt hard to touch and was resistant to probing (Fig. 1D). The tooth was hemisected using high speed rotary instruments. The mesial root and a generous segment of periodontium surrounding it, including the graft site, were then removed (Fig. IE). The flaps were coapted using 000 surgical silk, but the socket of the extracted root was left open (Fig. IF). The surgical specimen (Fig. 3) was fixed in 10% buffered formalin, decalcified in a weak solution of nitric acid, embedded in paraffin, and prepared for histologie examination in the usual manner. Serial sections 5 µ thick were cut starting from the buccal line angle and proceeding towards the lingual surface. Specimens for study were stained with hematoxylin and eosin and the Bloom variation of the Mallory stain. Histologie Evaluation. Under low power magnification the bone comprising the proximal wall of the original osseous periodontal defect could be distinguished easily from the grafted bone by virtue of the organized and regular lamellar pattern (Figs. 4A and 5A). The marrow spaces were of varying sizes, but the general character of
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Figure 4. Low power photomicrograph of site of osseous autografi. Multiple particles of grafted bone can be seen within the former of the host site bone are defect and a small island of newly forming bone is present at the base of the lesion. Some of the marrow spaces FM new BG bone NB cells. with chronic bone, are some and fibrous marrow, IC inflammatory graft, highly infiltrated fibrosed inflammatory cells, Ce cementum, A separation artifact, (H & E, original magnification, x 25). B. Higher magnification of graft site seen in Figure 4A. Active Osteogenesis can be noted about the particles ofgraft bone. The area is infiltrated with chronic inflammatory cells. Note the intraepithelial tear in the junctional epithelium which is associated with the separation artifact (Original magnification, =
=
=
x
=
=
=
63).
the alveolar bone from the edentulous zone as seen on the microscopic sections could be regarded as dense. Some remnants of fat were contained within the marrow cavities, but for the most part the marrow spaces were fibrous in nature with a substantial infiltration of chronic inflammatory cells. An intense lymphocytic infiltrate was present in one large marrow space (Fig. 4A). The surface of the host bone at the graft interface was covered by a thin layer of osteoid and osteoblasts, indicating active Osteogenesis in this area. Numerous particles of grafted bone were readily visible within the fibrous connective tissue contained between the lamellar bone and the tooth (Figs. 4 and 5). The transplanted chips were devoid of osteocytes, but woven bone containing viable osteocytes was seen on their surfaces (Fig. 4B). Osteoblastic activity and ingrowths of new capillaries were associated with all of the particles of grafted bone. The connective tissue bed was infiltrated with inflammatory cells, particularly in the cervical one-half of the grafted zone (Fig. 4B). At the base of the original osseous defect, a small island of new bone which seemed unrelated to the implanted bone was
present. A slender trabecula of new bone also was evident
the tooth at a more coronal level (Figs. 4A An artifactual space was present between the root surface and the autografi (Figs. 4-6). Associated with this void was a thin layer of epithelial cells which
adjacent and
to
6A).
adherent to both the cementai surface and the wall containing the grafted bone chips (Figs. 4-6). Despite the thinness of the epithelium (two to three cells wide), the separation artifact was seen to be intraepithelial in nature. The epithelial cells were of the squamous type, and their nuclei were flattened and orientated parallel to the root surface as is generally the case in junctional epithelium (Fig. 6). Examination of serial sections revealed that thicker plaques of squamous epithelium were present at various intervals along the thin trail of downgrowing epithelium and that a continuity existed between the long epithelial attachment and the sulcular epithelium. At the marginal aspect of the defect, two small fragments of nonvital graft bone were encapsulated by were
epithelial ingrowths (Fig. 7). Changes in the root surface suggestive of previous aggressive root planing were not prominent. A distinct
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Figure 5A. Low power photomicrograph of serially cut specimen of bone graft site showing active apposition of new bone on to transplanted particles of bone. Separation artifact resulting from histologie processing and epithelial downgrowth along root surface can be noted. BG bone graft, FM fibrous marrow, A separation artifact, Ce cementum, Pdl periodontal ligament, (BloomMallory stain, original magnification, x 25). B. High power magnification of microscopic field seen in lower right corner of Figure 5A. Particle of grafted bone is displaced into intact periodontal ligament and is apparently encapsulated. Evidence of root instrumentation and epithelial downgrowth is clearly seen (Original magnification, x 63). =
=
=
layer of pre-existing cementum was evident, but newlydeposited cementum could not be detected generally. The root surface lying in the base of the defect proved an exception to this observation. Here, evidences of a completely new attachment could be observed. This included recognizable newly-deposited cementum, apposition of new bone lamellae and a functionally-oriented periodontal ligament (Fig. 8). Sharpey's fibers were easily visible, and cementoblasts were lined up at regular intervals adjacent to the fiber insertion. Within the newly formed periodontal ligament, small spicules of nonvital bone, presumably grafted chips, were undergoing résorption (Fig. 9). This newly formed region of periodontium was apical to and remote from the bulk of the bony autografi. Discussion
widespread conviction that various types of bone grafts may have the capacity to induce the formation of an entirely new periodontium requires further proof and clarification, particularly in light of the known biologic behavior of periodontal tissues.3,6· "'12 Bone marrow transplants and other types of intraoral The
=
=
bone autografts provide a ready source of available material for stimulating Osteogenesis in periodontal defects; however, the concept that such grafts also have an inherent capacity for laying down new cementum on a previously denuded root surface and creating a functional periodontal ligament has yet to be demonstrated unequivocally. Animal experiments in which surgically created or artificially-provoked periodontal defects are used for bone implant studies are not ideal to test this thesis, as it has been shown that such lesions have an inherent ability for spontaneous regeneration of lost periodontium.17"19 The histologie study of human block sections is also subject to difficulties in interpretation arising out of orientation and plane of section and the frequent lack of serial sections.11,12,14"1B Also, sometimes it is difficult to establish whether the healing seen in bone implant sites is associated with the inherent capacity of certain types of periodontal lesions to regenerate spontaneously, or whether, in fact, it is stimulated by the bone graft itself. Melcher has described the shortcomings of the concept of utilizing bone grafts in attempts to regenerate periodontium.20, 21 Bone is only one of the connective tissues
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Photomicrograph of region slightly apical to the graft site. A plaque of epithelium is clearly seen along the root surface and separation artifact intervenes between the intraepithelial split. A slender trabecula of new bone can be seen adjacent to the junctional epithelium, and a larger island of newlyforming bone is present at a more apical level. An area of new attachment exists at the lower portion of the microscopic field. BG bone graft, NB new bone, A separation artifact, Ce cementum, NCE new cementum, (H & E, original magnification, x 63). B. Higher magnification ofjunctional epithelial plaque seen on Figure 6A. Note new cementum apposition apical to the epithelial downgrowth (Original magnification, x 160). Figure 6 A. a
=
=
of the periodontium that provides a continually adapting mechanism for support and function of the teeth. A second is the periodontal ligament. This tissue attaches the tooth to the alveolar process of the jaws and contains cells that have the capacity to deposit and resorb bone, cementum, and the soft connective tissue of the ligament itself. A homeostatic mechanism that maintains the periodontal space throughout life apparently operates within the periodontium. It has been suggested that this mechanism is at least partially mediated by the ability of the Based periodontal ligament cells to inhibit on our known understanding of the physiology of the periodontium, it would be reasonable to assume that the synthesis and deposition of periodontal ligament, alveolar bone and cementum are largely functions of the cells of the periodontal ligament and the endosteal cells of the bone, and that maintenance of the alveolar bone is dependent upon the function of the tooth.23 The functional stimuli required for maintenance of the alveolar bone are unlikely to be transmitted unless an intact periodontal ligament provides continuity between the tooth and bone. Based on the above understanding, it does not seem unreasonable to set as a primary objective
Osteogenesis.22
=
=
=
regeneration of the periodontal ligament rather than regeneration of bone. As stated by Melcher, this concept rests on the premise that cells of regenerated periodontal ligament have the capacity to synthesize and deposit the extracellular substance of bone, cementum and periodontal ligament and thereby restore a functioning periodontium.21
The presence of new bone that was found on surgical reentry was substantiated by histologie examination. Trabeculae of new bone were consistently seen about particles of the implanted bone, some of which exhibited recognizable areas of résorption (Fig. 4). After 28 weeks, the implant material was readily discernible microscopically but was easily distinguishable from the adjacent wall of cancellous bone, although it appeared to be continuous with it (Figs. 4 and 5). Despite the careful closure of the wound, and the lack of clinical evidence of sequestration of graft particles, downgrowth of a thin layer of epithelial cells was seen. These cells were typical of junctional epithelium and formed a delicate network extending to the base of the defect, but coronal to the area of new attachment (Fig. 6). A detailed study of serial sections confirmed the presence of epithelium
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297
SBSggM 1
BG
s':
·'
-.'^
wir Figure 7. Marginal region of autografi site depicting epithelial invagination and encapsulation of bone graft particles 28 weeks following bone transplantation. Adjacent bone graft particles show apposition of new bone. BG bone graft, Ep epithelium, SEp sulcular epithelium, Ce cementum, (H & E, original magnification, x 63). =
=
=
=
along the side of the root, although in most instances it was only one or two cells thick. Thicker plaques of epithelium were noted occasionally, and at the orifice of the defect there was evidence of epithelium encompassing small particles of graft bone, which can be anticipated ultimately as being rejected (Fig. 7). As deeper sections through the surgical site into the periphery of the graft area were made, true evidence of a regenerated attachment could be noted (Fig. 8). While evidence of grafted
found close to the base of the defect, we observations to suggest that the regenerainterpret tion in the basal area of the periodontium represents spontaneous healing unrelated to the graft. We propose that microscopic evidence of graft bone continues to be present for months or perhaps years and becomes a permanent record of the biologic activity of the graft bone
was
our
Unless a grafted periodontal wound provides evidence of attachment of periodontal ligament fibers to bone deposited on recognizable bone graft particles and to new cementum at the same level on the root, it seems unlikely that any claim to the contribution of the graft to the regeneration of the periodontium can be made. In this regard, it is relevant to note that Frank have noted deposition of cemenet al.24 and tum to be most advanced in the apical aspects of ungrafted periodontal wounds in man and monkey respec-
region. some
Listgarten25
tively.
The separation of the newly deposited cementum from the root surface seen on the microscopic sections, showing an area of regeneration (Fig. 8), was undoubtedly artifactual. Similar artifactitious splits frequently have been noted in histologie material of healing periodontal
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11
#
·
·&
>·.
vi
Figure 8. « of new attachment apical to and apparently unrelated to bone graft. Note laying down of new cementum on previously instrumented root surface and apposition of new alveolar bone proper. The periodontal ligament fibers appear to be functionally orientated. A separation artifact is present at the interface between the root surface and the new layer of cellular cementum. De dentin, NCe new cementum, Nb new bone, A separation artifact, Pdl periodontal ligament, (H & E, original magnification, X 63). =
=
wounds,1
=
=
and in all probability result from differential shrinkage in processing.20 It is doubtful whether this type of artifact has any clinical implication in reattachment therapy as suggested by Hawley and Miller.16 The epithelial downgrowth with concurrent osseous repair reported in this human material is strikingly similar to the case described by Caton and Zander in a provoked periodontal lesion in a Rhesus monkey.28 Complete healing and "fill" of an infrabony pocket had occurred; however, histologie sections showed a long junctional epithelial attachment extending to the most apical level of root instrumentation. Clinically, and in every other manner, this appeared to be a successfully treated lesion and indicates that bone remodeling can occur without connective tissue attachment to the root. The long term implication associated with a long
=
to bone does not appear be favorable, as the cellular potential to form new cementum (i.e. the periodontal ligament) is physically and functionally remote from the area in question. It seems more likely that this type of attachment would be weaker than a connective tissue attachment and would be vulnerable to future ingrowths of epithelium and eventual pocket formation. The observations made in this study suggest that the procedure of surgical reentry following bone grafting to confirm regeneration of the periodontium is of little value. The clinical evidence of newly formed bone does not necessarily imply apposition of new cementum on the root nor the presence of a functional periodontal ligament. Such judgements can be made only through careful study of histologie material.
epithelial attachment adjacent to
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299
Figure 9. Small particles ofgraft bone undergoing résorption in area ofperiodontal ligament apical to bulk ofgrafted bone. Note the multinucleated giant cells associated with the resorptive process. This appears to be an area of new attachment with active cementum and bone formation taking place. De dentin, Nb new bone, NCe new cementum, Pdl periodontal ligament, bg bone graft, Gc giant cells (H & E, original magnification, x 119). =
=
=
=
=
=
Summary
and
Conclusions
A histological study of the site of a clinically successful bone graft revealed the presence of a downgrowing junctional epithelium between the root surface and the altered graft bone. No new cementum could be seen associated with the osseous autografi, nor was a functional periodontal ligament in evidence adjacent to the grafted bone. It is suggested that the presence of newly formed bone on surgical reentry following bone grafting in periodontal lesions does not necessarily imply that regeneration of the periodontium has occurred. Acknowledgments
The authors are sincerely grateful for the technical assistance provided by Mr. Arthur Bloom, histopathologic technician, and Ms. Ida Nathan, medical photographer. Professors A. H. Melcher and Bernard Wasserman were kind enough to review
the are
manuscript, and their greatly appreciated.
critical comments and
suggestions
References 1. Beube, F. E., and Silvers, H. F.: Further studies on bone regeneration with the use of boiled heterogenous bone. J Periodontol 7: 17, 1936. 2. Scopp, I. W., Morgan, F. H., Dooner, J. J., Freddes, H. J., and Heyman, R. .: Bovine bone (Boplant) implant for infrabony oral lesions (Clinical trials in humans). Periodontics 4: 169, 1966. 3. Nabers, C. L., and O'Leary, T. J.: Autogenous bone transplant in the treatment of osseous defects. J Periodontol 36:
5, 1965. 4. Robinson, R. E.: Osseous coagulum for bone induction. / Periodontol 40: 503, 1969. 5. Schallhorn, R. G, Hiatt, W. H., and Boyce, W.: Iliac transplants in periodontal therapy. J Periodontol 41: 566, 1970. 6. Rosenberg, . M.: Free osseous predictable procedure. J Periodontol 42:
tissue autografts 195. 1971.
as a
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Mosko w,
J. Periodontol. June, 1979
Karsh, Stein
7. Diem, C. R., Bowers, G. M., and Moffitt, W. C: Bone A technique for osseous implants. J Periodontol 43: 295, 1972. 8. Hiatt, W. H., and Schallhorn, R. G.: Intraoral transplants of cancellous bone and marrow in periodontal lesions. / Periodontol 44: 194, 1973. 9. Libin, B. M., Ward, H. L., and Fishman, L.: Decalcified, lyophilized bone allografts for use in human periodontal defects. / Periodontol 46: 51, 1975. 10. Mellonig, J. T., Bowers, G. M., Bright, R. W., and Lawrence, J. J.: Clinical evaluation of freezedried bone allografts in periodontal osseous defects. J Periodontol 47: 125, 1976. 11. Schallhorn, R. G.: Osseous grafts in the treatment of periodontal osseous defects. Periodontal Surgery: Biologic Basis and Technique, Springfield, Thomas, 1976. 12. Dragoo, M. R., and Sullivan, H. C: A clinical and histologie evaluation of autogenous iliac bone grafts in humans: Part I. Wound healing two to eight months. J Periodontol 44: 599, 1973. 13. Ross, S. E., and Cohen, D. W.: The fate of a free osseous tissue autografi, a clinical and histologie case report. Periodontics 6: 145, 1968. 14. Nabers, C. L., Reed, O. M., and Hamner, J. E.: Gross and histologie evaluation of an autogenous bone graft 57 months postoperatively. J Periodontol 43: 702, 1972. 15. Froum, S. J., Thaler, R., Scopp, I. W„ and Stahl, S. S.: Osseous autografts. II. Histologie responses to osseous coagulum-bone blend grafts. J Periodontol 46: 656, 1975. 16. Hawley, C. E., and Miller, J.: A histologie examination of a free osseous autograft. J Periodontol 46: 289, 1975. 17. Linghorne, W. J., and O'Connel, D. C: Studies in the regeneration and reattachment of supporting structures of the
blending:
teeth. II. Regeneration of alveolar process. J Dent Res 30: 604, 1951. 18. Linghorne, W. J., and O'Connel, D. C: Studies in the reattachment and regeneration of the supporting structures of the teeth. III. Regeneration in epithelialized pockets. J Dent Res 34: 164, 1955. 19. Ramfjord, S.: Experimental periodontal reattachment in Rhesus monkeys. J Periodontol 22: 67, 1951. 20. Melcher, A. H.: On the repair potential of periodontal tissues. J Periodontol 47: 256, 1976. 21. Melcher, A. H: Biological processes in résorption, deposition and regeneration of bone. Periodontal Surgery: Biologic Basis and Technique. Springfield, Thomas, 1976. 22. Melcher, A. H.: Repair of wounds in the periodontium of the rat. Influence of periodontal ligament on Osteogenesis. Arch Oral Biol 15: 1183, 1970. 23. Cohn, S. .: Disuse atrophy of the periodontium in mice. Arch Oral Biol 10: 909, 1965. 24. Frank, R., Fiore-Donno, G., Cimasoni, G., and Matter, J.: Ultrastructural study of epithelial and connective gingival reattachment in man. J Periodontol 45: 626, 1974. 25. Listgarten, . .: Electron microscopic study of the junction between surgically denuded root surfaces and regenerated periodontal tissues. J Periodont Res 7: 68, 1972. 26. Morris, M. L.: Healing of naturally occurring periodontal pockets about vital human teeth. J Periodontol 26: 285, 1955. 27. Jansen, M. T., Coppes, L., and Verdenius, H. H. W.: The healing of periodontal wounds in dogs. J Periodontol 26: 292, 1955. 28. Caton, J., and Zander, . .: Osseous repair of infrabony pocket without new attachment of connective tissue. J Clin Periodont 3: 54, 1976.
Abstracts Dental Treatment for Patients
with
Leukemia
Carl, W.
Quintessence Int 9: 9, August,
1978.
The oral changes of leukemia patients before and after chemotherapy and dental treatment for patients with leukemia were described. The first sign of the leukemia may appear in the mouth and usually as the disease progresses, the oral manifestations progress proportionately. By maintaining or establishing good oral hygiene in patients with leukemia, the oral manifestations of the disease can be greatly reduced. Many times, extractions have to be done, even when the patient is in an acute leukemic state and specific precautions are needed. Team work including other related departments is necessary to deal with all the complications that may develop during the course of this disease. Dr. Yau-Fwu Huang 66 Elm Street, Buffalo, NY 14263.
A Simple Filter Paper Strip Viewer Estimation
Wilson, C. ., Tay, W. M., and Morgan, Quintessence Int 9: 63, August, 1978.
for
Gingival Fluid
W. J.
The most widely used method of collecting gingival exúdate for the degree of inflammation in a gingival sulcus is to insert filter paper strips at varying depths into the gingival crevice. By application of ninhydrin reagent stains, the exúdate on the paper becomes visible and can be measured. A new design to measure the stained area of filter paper strip was described and illustrated. A 35 mm photographic slide viewer was used and Letratape strips were placed on a viewing lens with a Letratone grid. The viewer possesses the versatility of a microscope at a fraction of the cost. British Forces Dr. Yau-Fwu Huang Post Office 34, London, England.
assessing
