/, Periodonlat Res. 13: 498-503, 1978
A scanning electron microscopic study of the destruction of human alveolar crest in periodontal disease K. OoYA AND H. YAMAMOTO
Department of Oral Pathology, School of Dentistry, Tohoku University. Japan The crestal surfaces of human alveolar bone obtained at autopsy were studied by scanning electron microscopy. Healthy alveolar crests v/ere covered by supracrestal fibers, making it difficult to prepare and visualize collagen-free crestal bone. The incipient stage of the iXcstruction o( Ihe aJveolar crestal bone was associated with a loosening of the supracresta! fibers. Bone surfaces free of supracrestal fibers were easily prepared from the active inflammatory lesions. These denuded bone surfaces were characterized by numerous Howship's lacunae. In the resting stage, the alveolar crest had a honeycomb appearance with a covering plexus of collagen fibers and mineralized-like collagen matrix. It was concluded that the destruction of supracrestal fibers was important in the process of the destruction of the alveolar cresta! bone, which proceeded by osteoclastic resorption. (Accepted for publication February 23, 1978)
Introduction
Bone destruction takes place in a wide variety of pathological conditions. Alveolar bone destruction is an essential phenomenon in periodontal disease. Many theories have been proposed regarding the etiology of bone destruction in periodonJal disease (Weinmann 1941, Glickman & Wood 1942, Reichborn-Kjennerud 1963, Hancox & Boothroyd 1963). Bone destruction in periodontal disease is not necessarily a continuous process. The destruction of the alveolar bone in periodontal disease is determined by the balance of resorption and regeneration of bone (Thoma & Goldman 1937). Alveolar bone is often destroyed by osteoclastic resorption adjacent to sites of inflamed connective tissue. The purpose of this investigation was to observe the changes
on the surface of the alveolar crest by means of light microscopy, microradiography and scanning electron microscopy. Special attention was paid to the fine structural alteration of the connections between supracrestal fibers and the surface of the alveolar crestal bone.
Materials and Methods
The materials used were obtained within five hours after death from one hundred and twenty-two cadavers aged from 15 ;o 75 years which were secured at autopsy. Two hundred upper and lower jaws were fixed in 10 % neutral, buffered formalin. One hundred specimen blocks were pri^ pared by cutting in a buccolingual pla e parallel to the long axis of the teeth. H: If of the block was decalcified for 4S hrs a
DESTRUCTION OF H U M A N ALVEOLAR CREST 5.2 % nitric acid, dehydrated in graded alcohols, embedded in paraffin, serially sectioned at 8 microns and stained with haematoxylin-eosin stain and silver impregnation. The other half was intended for scanning ekctTon microscopy and the connective tissue of the gingiva was removed mechanically under a binocular microscope from the alveolar crest. Twenty specimens for scanning electron microscopy were dehydrated in acetone, cleaned by ultrasonic means to remove bone dust and other debris prior to coating with carbon and gold under continuous tilting and rotation, and examined in the JSM U-3 scanning electron microscope. The same specimens were examined by microradiography. They were cut on a Giliing.s Bronwill thin sectioning machine and the thickness reduced by grinding to 80 ^m. Microradiographs were produced on Kodak spectroscopic plates 649-0 with a Softex CMR type soft x-ray machine operated at 7 kV and 3 mA.
ResuHs
In healthy aiveoiar bone, the alveolar crest was difficult to prepare free from supracrestal fibers. The alveolar crest was covered by supracrestal fibers which ran from cementum to the crestal bone (Fig. la). The principal fibers on the alveolar crestal bone appeared to be the predominant constituent and were made up of fibrils 700 A to 1300 A in diameter, as seen by scanning electron microscopy (Fig. lb). Microradiography showed that the healthy alveolar crestal bone consisted of two lamina: a thin marginal supporting layer and an alveolar bone proper enclosing small marrow spaces (Fig. lc). In the incipient stage of the destruction of the alveolar crest which indicated the in'iimal nature of the inflammatory state, th suptacrestal fibers revealed slight irregul. r arrangement and loosening of the con-
499
nections with the alveolar crestal bone (Fig. 2a). Individual supracrestal fibers were less distinct and shown as bundles of coherent coUagenous fibers, in contrast to the normal supracrestal fibers (Fig. 2b). The marginal supporting layer of the alveolar crestal bone showed some destruction and enlargement of the marrow spaces by microradiography (Fig. 2c). The active stage of the destruction of the aiveoiar crest which indicated an accurnuiation of inflammatory cells in the periodontal ligament was histologically distinguished from the other stages. In about 7 % of specimens examined, gingival inflammation was apparent and gingiva! fibers were almost completely lost. Supracrestal fibers were also destroyed, making it easy to prepare alveolar crestal bone free from supracrestal fibers. Light microscopy showed osteociasts lying in Howship's lacunae on the alveolar crestal bone surface (Fig. 3a). Scanning electron microscopy (Fig. 3b-l-2) showed bone destruction with Howship's lacunae which corresponded to the large multi-nucleated osteoclasts recognized by light microscopy. At high magnification, bone salt crystals and denuded bone collagen fibers were observed in the resorbing sites (Fig. 3b-3). The typical scalloped profile af resorption lacunae were evident by microradiography (Fig. 3c). In the resting stage, the resorbing alveolar crest occurred with conversion to a fibrous character. Light microscopy showed new bone formation on the border of the alveolar crestal bone in about 20 % of the specimens examined (Fig. 4a-1). Marrow spaces of alveolar bone showed collagen deposition and demarcation between new and mature bone (Fig. 4a-2), Scanning electron microscopy showed that a collagen fiber plexus covered the typically scalloped profile of the alveolar crest. In some portion, a mineralized-I ike fibrillar network could be seen (Fig. 4b). Microradiography showed
OOYA AND YAMAMOTO
Fig. 1. Healthy alveolar crest, a. Light microscope shews the buccal aspect ot denlo-periosteal fit ^ra crossing over the alveolar crest (AC). (C - cementum, B = alveolar bone), b. Scanning electron microgr ph (SEM) of aiveoiar crestal bone surface covered by supracrestal tibers. x 10,000. c. Microradiograph sh vi(s thin marginal supporting layer (SL) and alveolar bone proper (AP). x 100. Fig. 2. Incipient stage of the destruction of the aiveoiar crest, a. LfghC microscope shows minimal inflam la-
D E S T R U C T I O N OF H U M A N ALVEOLAR
OREST
cresl. a. Light microscope shows osteoclasts (OC) lying in Howship's lacunae on Ihe alveolar crestal bone surface, b-1. SEM feature of figure 3a. prepared by sectioning buccolingually. Note crest (AC) is not covered by supracrestal fib 100. b-2. Alveolar crestal suriace of figur shows numerous resorption iacunae. Althouol ostly correspo to the orptive cells. Resorbing Howship's ot lacunae, the length of which ranged fi mately 30 to 70 ^m. The lacunae of the (arrows) are exposed by these resorptior X 1,000. b-3. High magnification of figure 3b-; bone saft crystals and bone collagen fibei c. Resorbing suriace in microradiograph shows Ihe typical scalloped profile of resorption lacunas (arrows). Lacunae of the osteocyte can be seen. X 500. 'ory 3tate i nd the [c she.5 a re ;oherent appearance in SEM. x 330. c. Mic laye show; slight df Btrudion and enlargemem ol bone marrow (
radiograpfi of figure 2b. Supporting a\v ) spaces. X 100.
OOYA AND Y A M A M O T O
4a-i
Fig. 4. Resting stage of the destruction of the a lar crest, a-1. Light microscope shows new bont mation (arrow) on the border of the alveolar ci bone and collagen deposition of enlarged bone row (BM) spaces, a-2. Light microscope shows bone formation at supracrestai and endosteal \ ivithin boni 'ace with r irregular holes covered wit nective tissues shows a mineralized-like fibrilh work (MF) in SEU. x 3,000. c. Microradiograph shows smoothly defined alveolar crest (arrow) and erilarged bone marrow (BM) spaces, x TOO.
smoother alveolar crestal surface than that of active stage and enlarged marrow spaces (Fig. 4c).
ligament while others thought that the inflammation spread from the gingiva into alveolar bone and that rarely, if ever, extension was directly into the periodontal ligament (Kronfeld 1935, Thoma & Goldman Discussion 1937). On the other hand, coUagenase activFor many years, opinion has differed re- ity was demonstrated in inflamed gingi-al connective tissue (Fullmer & Gibson 19(-6) garding the pathway followed by the extension of gingival inflammation into the sup- and the collagen content of the infiltraied porting periodontium. Coolidge (1931) and portion of the gingival connective tissue \ ^is Noyes (1937) held that inflammation ex- shown to be markedly reduced by morp otended along the fibers of the periodontal metric and biochemical analysis (Schroe er
DESTRUOTION OF H U M A ^4 ALVEOLAR CREST et al. 1973). The results of this study illustrate the predominance of periosteal resorption of the alveolar crest and the extension of gingival inflammation into the supporting periodontal tissues directly. It is also suggested that an almost continuous lining of resorption lacunae along the periosteal aspect of the alveolar crest might compromise the anchorage of the supracrestal fibers. Glickman (1972) suggested that the bone destruction caused by inflammation was necessarily a continuous process and that bone loss in periodontal disease was not simply a destruction process since the response of alveolar bone to inflammation included bone formation as well as resorption. In this study, the cresta! bone was often destroyed by osteoclastic resorption. Deposition of collagen and mineralization of the fibers on the alveolar crest during the resting stage may be considered a defense mechanism against further destruction of the alveolar crestal bone due to inflammation. Scanning electron microscopy provided reliable identification of the functional state and overall activity of the alveolar bone surface.
Acknowledgement
This study was supported by a Grant-in-Aid for Scientific Research (No. 47038) from the Ministry of Education, Science and Culture of Japan. Address: Oi'partmcnt of Oral Pathology Siliool of Dentistry T^ohoku University Stiryo, Sendai, 980
References
CooUdge, E. D. 1931. Inflammatory changes in the gingiva! tissue due to local irritation. /. Am. Dent, Assoc. 18: 2255. Fullmer, H. M. & Gibson, W. A. 1966. CoUagenolytic activity in gingivae of man. Nature. 209: 728-729. Glickman, 1. 1972. Bone loss and patterns of bone destruction in periodontal disease. In: Clinical periodontology, &1. Glickman, I., pp. 218-232. Philadelphia, London, Toronto: W.B. Saunders Co. Glickman, 1. & Wood, H. 1942. Bone histology in periodontal disease. /. Dent. Res. 21: 3554. Hancox, N. M. & Boothroyd, B. 1963. Structure-function relationships in the osteoclast. In: Mechanisms of hard tissue destruction. ed. Sognnaes, R. F., pp. 497-514. Washington. D. C : Am. Assoc. Adv. Sci. Kronfeid, R. 1935. The condition of the alveolar bone underlying periodonta! pockets. J. Periodontol. 6: 22-29. Noyes, F. B. 1937. A review of the work in lymphatics of dental origin, /. Am. Dent. Assoc. 14: 714. Reichhorn-Kjennerud, I. 1963. Dento-alveolar resorption in periodontal disorders. Tn: Mechanisms of hard tissue destruction, ed. Sognnaes, R.F., pp. 297-319. Washington. D.C.: Am. Assoc. Adv. Sci. Schroeder, H. E., Mlinzel-Pedrazzoli. S. & Page, R. 1973. Correlated morphometric and biochemical analysis of gingival tissue in early chronic gingivitis in man. Arch. Oral Biol. 18: 899-923. Thoma, K. H. & Goldman, H. M. 1937. Classification and histopathoiogy of parodontal disease. J. Am. Dent. Assoc. 24: 1915-1928. Weinmann, J.P. 1941. Progress of gingival inflammation into the supporting structures of the teeth. /. PeriodontoL 12: 71-82.
A scanning electron microscopic study of the destruction of human alveolar crest in periodontal disease K. OoYA AND H. YAMAMOTO
Department of Oral Pathology, School of Dentistry, Tohoku University. Japan The crestal surfaces of human alveolar bone obtained at autopsy were studied by scanning electron microscopy. Healthy alveolar crests v/ere covered by supracrestal fibers, making it difficult to prepare and visualize collagen-free crestal bone. The incipient stage of the iXcstruction o( Ihe aJveolar crestal bone was associated with a loosening of the supracresta! fibers. Bone surfaces free of supracrestal fibers were easily prepared from the active inflammatory lesions. These denuded bone surfaces were characterized by numerous Howship's lacunae. In the resting stage, the alveolar crest had a honeycomb appearance with a covering plexus of collagen fibers and mineralized-like collagen matrix. It was concluded that the destruction of supracrestal fibers was important in the process of the destruction of the alveolar cresta! bone, which proceeded by osteoclastic resorption. (Accepted for publication February 23, 1978)
Introduction
Bone destruction takes place in a wide variety of pathological conditions. Alveolar bone destruction is an essential phenomenon in periodontal disease. Many theories have been proposed regarding the etiology of bone destruction in periodonJal disease (Weinmann 1941, Glickman & Wood 1942, Reichborn-Kjennerud 1963, Hancox & Boothroyd 1963). Bone destruction in periodontal disease is not necessarily a continuous process. The destruction of the alveolar bone in periodontal disease is determined by the balance of resorption and regeneration of bone (Thoma & Goldman 1937). Alveolar bone is often destroyed by osteoclastic resorption adjacent to sites of inflamed connective tissue. The purpose of this investigation was to observe the changes
on the surface of the alveolar crest by means of light microscopy, microradiography and scanning electron microscopy. Special attention was paid to the fine structural alteration of the connections between supracrestal fibers and the surface of the alveolar crestal bone.
Materials and Methods
The materials used were obtained within five hours after death from one hundred and twenty-two cadavers aged from 15 ;o 75 years which were secured at autopsy. Two hundred upper and lower jaws were fixed in 10 % neutral, buffered formalin. One hundred specimen blocks were pri^ pared by cutting in a buccolingual pla e parallel to the long axis of the teeth. H: If of the block was decalcified for 4S hrs a
DESTRUCTION OF H U M A N ALVEOLAR CREST 5.2 % nitric acid, dehydrated in graded alcohols, embedded in paraffin, serially sectioned at 8 microns and stained with haematoxylin-eosin stain and silver impregnation. The other half was intended for scanning ekctTon microscopy and the connective tissue of the gingiva was removed mechanically under a binocular microscope from the alveolar crest. Twenty specimens for scanning electron microscopy were dehydrated in acetone, cleaned by ultrasonic means to remove bone dust and other debris prior to coating with carbon and gold under continuous tilting and rotation, and examined in the JSM U-3 scanning electron microscope. The same specimens were examined by microradiography. They were cut on a Giliing.s Bronwill thin sectioning machine and the thickness reduced by grinding to 80 ^m. Microradiographs were produced on Kodak spectroscopic plates 649-0 with a Softex CMR type soft x-ray machine operated at 7 kV and 3 mA.
ResuHs
In healthy aiveoiar bone, the alveolar crest was difficult to prepare free from supracrestal fibers. The alveolar crest was covered by supracrestal fibers which ran from cementum to the crestal bone (Fig. la). The principal fibers on the alveolar crestal bone appeared to be the predominant constituent and were made up of fibrils 700 A to 1300 A in diameter, as seen by scanning electron microscopy (Fig. lb). Microradiography showed that the healthy alveolar crestal bone consisted of two lamina: a thin marginal supporting layer and an alveolar bone proper enclosing small marrow spaces (Fig. lc). In the incipient stage of the destruction of the alveolar crest which indicated the in'iimal nature of the inflammatory state, th suptacrestal fibers revealed slight irregul. r arrangement and loosening of the con-
499
nections with the alveolar crestal bone (Fig. 2a). Individual supracrestal fibers were less distinct and shown as bundles of coherent coUagenous fibers, in contrast to the normal supracrestal fibers (Fig. 2b). The marginal supporting layer of the alveolar crestal bone showed some destruction and enlargement of the marrow spaces by microradiography (Fig. 2c). The active stage of the destruction of the aiveoiar crest which indicated an accurnuiation of inflammatory cells in the periodontal ligament was histologically distinguished from the other stages. In about 7 % of specimens examined, gingival inflammation was apparent and gingiva! fibers were almost completely lost. Supracrestal fibers were also destroyed, making it easy to prepare alveolar crestal bone free from supracrestal fibers. Light microscopy showed osteociasts lying in Howship's lacunae on the alveolar crestal bone surface (Fig. 3a). Scanning electron microscopy (Fig. 3b-l-2) showed bone destruction with Howship's lacunae which corresponded to the large multi-nucleated osteoclasts recognized by light microscopy. At high magnification, bone salt crystals and denuded bone collagen fibers were observed in the resorbing sites (Fig. 3b-3). The typical scalloped profile af resorption lacunae were evident by microradiography (Fig. 3c). In the resting stage, the resorbing alveolar crest occurred with conversion to a fibrous character. Light microscopy showed new bone formation on the border of the alveolar crestal bone in about 20 % of the specimens examined (Fig. 4a-1). Marrow spaces of alveolar bone showed collagen deposition and demarcation between new and mature bone (Fig. 4a-2), Scanning electron microscopy showed that a collagen fiber plexus covered the typically scalloped profile of the alveolar crest. In some portion, a mineralized-I ike fibrillar network could be seen (Fig. 4b). Microradiography showed
OOYA AND YAMAMOTO
Fig. 1. Healthy alveolar crest, a. Light microscope shews the buccal aspect ot denlo-periosteal fit ^ra crossing over the alveolar crest (AC). (C - cementum, B = alveolar bone), b. Scanning electron microgr ph (SEM) of aiveoiar crestal bone surface covered by supracrestal tibers. x 10,000. c. Microradiograph sh vi(s thin marginal supporting layer (SL) and alveolar bone proper (AP). x 100. Fig. 2. Incipient stage of the destruction of the aiveoiar crest, a. LfghC microscope shows minimal inflam la-
D E S T R U C T I O N OF H U M A N ALVEOLAR
OREST
cresl. a. Light microscope shows osteoclasts (OC) lying in Howship's lacunae on Ihe alveolar crestal bone surface, b-1. SEM feature of figure 3a. prepared by sectioning buccolingually. Note crest (AC) is not covered by supracrestal fib 100. b-2. Alveolar crestal suriace of figur shows numerous resorption iacunae. Althouol ostly correspo to the orptive cells. Resorbing Howship's ot lacunae, the length of which ranged fi mately 30 to 70 ^m. The lacunae of the (arrows) are exposed by these resorptior X 1,000. b-3. High magnification of figure 3b-; bone saft crystals and bone collagen fibei c. Resorbing suriace in microradiograph shows Ihe typical scalloped profile of resorption lacunas (arrows). Lacunae of the osteocyte can be seen. X 500. 'ory 3tate i nd the [c she.5 a re ;oherent appearance in SEM. x 330. c. Mic laye show; slight df Btrudion and enlargemem ol bone marrow (
radiograpfi of figure 2b. Supporting a\v ) spaces. X 100.
OOYA AND Y A M A M O T O
4a-i
Fig. 4. Resting stage of the destruction of the a lar crest, a-1. Light microscope shows new bont mation (arrow) on the border of the alveolar ci bone and collagen deposition of enlarged bone row (BM) spaces, a-2. Light microscope shows bone formation at supracrestai and endosteal \ ivithin boni 'ace with r irregular holes covered wit nective tissues shows a mineralized-like fibrilh work (MF) in SEU. x 3,000. c. Microradiograph shows smoothly defined alveolar crest (arrow) and erilarged bone marrow (BM) spaces, x TOO.
smoother alveolar crestal surface than that of active stage and enlarged marrow spaces (Fig. 4c).
ligament while others thought that the inflammation spread from the gingiva into alveolar bone and that rarely, if ever, extension was directly into the periodontal ligament (Kronfeld 1935, Thoma & Goldman Discussion 1937). On the other hand, coUagenase activFor many years, opinion has differed re- ity was demonstrated in inflamed gingi-al connective tissue (Fullmer & Gibson 19(-6) garding the pathway followed by the extension of gingival inflammation into the sup- and the collagen content of the infiltraied porting periodontium. Coolidge (1931) and portion of the gingival connective tissue \ ^is Noyes (1937) held that inflammation ex- shown to be markedly reduced by morp otended along the fibers of the periodontal metric and biochemical analysis (Schroe er
DESTRUOTION OF H U M A ^4 ALVEOLAR CREST et al. 1973). The results of this study illustrate the predominance of periosteal resorption of the alveolar crest and the extension of gingival inflammation into the supporting periodontal tissues directly. It is also suggested that an almost continuous lining of resorption lacunae along the periosteal aspect of the alveolar crest might compromise the anchorage of the supracrestal fibers. Glickman (1972) suggested that the bone destruction caused by inflammation was necessarily a continuous process and that bone loss in periodontal disease was not simply a destruction process since the response of alveolar bone to inflammation included bone formation as well as resorption. In this study, the cresta! bone was often destroyed by osteoclastic resorption. Deposition of collagen and mineralization of the fibers on the alveolar crest during the resting stage may be considered a defense mechanism against further destruction of the alveolar crestal bone due to inflammation. Scanning electron microscopy provided reliable identification of the functional state and overall activity of the alveolar bone surface.
Acknowledgement
This study was supported by a Grant-in-Aid for Scientific Research (No. 47038) from the Ministry of Education, Science and Culture of Japan. Address: Oi'partmcnt of Oral Pathology Siliool of Dentistry T^ohoku University Stiryo, Sendai, 980
References
CooUdge, E. D. 1931. Inflammatory changes in the gingiva! tissue due to local irritation. /. Am. Dent, Assoc. 18: 2255. Fullmer, H. M. & Gibson, W. A. 1966. CoUagenolytic activity in gingivae of man. Nature. 209: 728-729. Glickman, 1. 1972. Bone loss and patterns of bone destruction in periodontal disease. In: Clinical periodontology, &1. Glickman, I., pp. 218-232. Philadelphia, London, Toronto: W.B. Saunders Co. Glickman, 1. & Wood, H. 1942. Bone histology in periodontal disease. /. Dent. Res. 21: 3554. Hancox, N. M. & Boothroyd, B. 1963. Structure-function relationships in the osteoclast. In: Mechanisms of hard tissue destruction. ed. Sognnaes, R. F., pp. 497-514. Washington. D. C : Am. Assoc. Adv. Sci. Kronfeid, R. 1935. The condition of the alveolar bone underlying periodonta! pockets. J. Periodontol. 6: 22-29. Noyes, F. B. 1937. A review of the work in lymphatics of dental origin, /. Am. Dent. Assoc. 14: 714. Reichhorn-Kjennerud, I. 1963. Dento-alveolar resorption in periodontal disorders. Tn: Mechanisms of hard tissue destruction, ed. Sognnaes, R.F., pp. 297-319. Washington. D.C.: Am. Assoc. Adv. Sci. Schroeder, H. E., Mlinzel-Pedrazzoli. S. & Page, R. 1973. Correlated morphometric and biochemical analysis of gingival tissue in early chronic gingivitis in man. Arch. Oral Biol. 18: 899-923. Thoma, K. H. & Goldman, H. M. 1937. Classification and histopathoiogy of parodontal disease. J. Am. Dent. Assoc. 24: 1915-1928. Weinmann, J.P. 1941. Progress of gingival inflammation into the supporting structures of the teeth. /. PeriodontoL 12: 71-82.
