Comparison of cellular cementum in normal and diseased teeth-a scanning electron microscopic study

JOURNAL OF ENDODONTICS VOL 7, NO 8, AUGUST t981

Comparison of cellular cementum in normal and diseased teeth-a scanning electron microscopic study lames H. S. Simon, DDS; Gary S. Y o n e m o t o , DD5, MS; and Leif K. Bakland, DDS

M a n y investigators have studied the effect of periodontal disease on acellular c e m e n t u m at the ultrastructural level. ' ' However, the effect of i n t l a m m a t o r y disease on t:ellular c e m e n t u m has received m u c h less a t t e n t i o n and is contined to basic histology. Normal cellular c e m e n t u m divested of the overlying organic fibers, seen through the s c a n n i n g electron microscope (SEM), appears as uniform patterns of m o u n d s a n d depressions that produce a pebbled surface appearance. ' ' ' Seen in cross section a n d through transrnission electron microscope (q'EM), the c e m e n t u m appears separated because of small pyramids of calcification at the insertion of each periodontal ligament fibril.: 'Fhese pyramids m a y correspond to the surface m o u n d s or pebbles observed through the SEM. T w o SEM studies investigated the fractured surface of cellular cementum. "Foda and others" identitied Sharpey's fil-x,r bundles a n d classitied the depressions, or c e m e n t u m lacunae, according to the morphology of the lacunae walls. Hansoff' found that the fractured cellular c e m e n t u m had a coarse, uneven appearance, which was contrasted to the regular palisaded structure of acellular c e m e n t u m . However, neither study detailed the n o r m a l crosssectional morphology. A cellular c e m e n t u m exposed to periodontal disease has been

370

extensively studied. Contact microradiographs show that unexposed acellular c e m e n t u m is represented as a h e r n a t i n g bands of high a n d low mineral content, almost lamellar in a p p e a r a n c e ........ Exposed to oral fluids, acellular c e m e n t u m develops a r a d i o p a q u e layer 10 #m wide along the exposed surface, and is referred to as the hypermineralized layer? ....... ': The hypermineralized laver has a greater uniformity in mineral distribution compared with sound, unexposed c e m e n t u m . T h e hydroxyapatite crTstals are closely packed, elongated, a n d of u n i f o r m width; they have been designated as tablet-shaped.: ..... T h e purpose of this study was to investigate the cross sectional morphology of cellular c e m e n t u m usir~g the SEM a n d to compare the characteristics of cellular cementuna morphology in normal and i n t l a m m a t o ry, diseased teeth. 9

MATERIALS A N D M E T H O D S Five normal and five diseased singlerooted human teeth were included in this study. 'fhe teeth were collected from subjects ranging in age l,iom 25 to 55 years old. l,'he criteria ",verc as follows: normal teeth with vital pulps and without any evidence of restoration or periodontal pathology; and diseased teeth with necrotic pulps. Pre-extraction radii)graphs showed apical radiolucent areas, but showed no evidence of periodontal disease affecting the apical lesions. The freshly extracted human teeth

were rinsed in saline solution and immediately placed in either glutaraldehyde (29;) or bul,tered parafi)rmaktel'lyde (.t',::). The teeth were fractured longitudinally bv burring a groove along two thirds of the root surface, then they were split ~,ith a mallet and bibevel chisel. "Fhe specimens were post-tixed for an hour in osmium tetroxide butS-red m a pl I of 72 with .2 molar phosphate buffer (300 m()sm), and then washed m phosphate buffer, and dehydrated in a series of graded alcohol, 5(}':~. 705;..q()';~.. follov,cd by multiple washings in absolute alcohol. They were dried using the critical-point dwing method, with carbon dioxide as the transitional tluid The specimens were mounted, burred side down, on carriers with silver paint, and coated with a thin layer of gold palladium that u s e d a vacuum evaporator to make then) conductive. The specimens were examined with an AMR model 1.000 scanning electron microscope at 20 KV.

1)eveloprnent of the observatmn aitcria. There ',,,ere tv,'o areas of study located on the apical root surface covered with cellular cementurn (Fig 1). Area 1 was a rectangular area, bound bv tile edge of the foramen, and bound apically by the tip of Ill(: fractured portion of tile root. Area 2 was a rectangular area, along the periphery of tile fractured cmnental surlace. Specifying the areas to be examined allo~ed tile study of comparal,)le areas among various specimens, regardless of tl,ae inherent dil,l,erences in root nmrphol" ogy. St{M photomicrographs of the cellular cementum from areas l, and 2 were taken at magnifications Hf 500 X, 1,000 X, and 2,000 • of all the specimens. There were three consistently identifiable surlaCe characteristics common to every pk rare.

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k'tg 2 Top, fractured surface of normal ~ellular cementum showmg random(y arranged uncalc~ed fibers (ortg mag x 2,000, area 2). Bottom, dt.[]erent specimen showing norreal cellular cementum morphologr (onlr mag • 2,000, area 2). P. projectwn; D, depre.,sion; F. fiber.

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:,-," There were labeled as projections, depressions, and fibers (Fig 2); they were counted on the picture magnified 2,00O x . To facilitate the counting, a transparent grid, 90 mm by 115 mm, and consisting of 88 2 by 2 mm crosses, was superimposed over all the SEM photomicrographs magnilied 2,000• Whenever any portion of a cross coincided with the characteristic being investigated, the cross was counted. Energy' di.q~erswe X-ray spectrometo'. In addition, two of the ten specimens, one from each of the two test groups, were analyzed for their relative calcium and phosphorus mineral content using energy dispersive X-ray spectrometry'. Focal points along the fractured surface border, located in areas 1 and 2, ,,,,'ere analyzed. The teeth were examined using an ARM Model 1,000 scanning electron microscope with a Kevex 5,100 system X-ray energy spectrometer. Photomicrographs were taken of the resulting elemental distribution maps for comparison purposes. Stattstical amdysis of the data (MannWhither test). The data were treated as ordinal data and the Mann-Whitney test was used to determine significant differences between normal and diseased cellular cementum, when evaluated according to the three morphologic characteristics.

RESULTS W h e n viewed in cross section, the n o r m a l cellular c e m e n t u m morphology' consiste(t of calcitied projections a n d corresponding depressions. Interspersed on this irregular surface >,'ere some depressions that have been identified as cementocyte lacun a e / Small, r a n d o m l y arranged, uncalcified fibers were e m b e d d e d in the calcified matrix (Fig 2). Visual dissimilarities were obse~'ed in the morphology of the norrnal specimens compared with the diseased ones (Fig 3). T h e n o r m a l specimens had consistent surface patterns of regularly arranged projections, depressions, and fibers, in contrast, the diseased specimens were more haphazardly arranged and contained more calcified projections a n d c e m e n t u m lacunas, but they had

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fewer fibers than the normal specimens. These observations were q u a n tified a n d statistically e v a l u a w d with the Mann-~A'hitney test. These resuhs were found Io be statistically significant at alpha = .05 (Table).

Res,rption. A consistent observation in the diseased specimens was surface resorption. T h e resorptive areas appeared at or near the apex and ahmg the periphery of the fractured cemental surface that produced scalloped, bay-like areas. W i t h i n some of these resorpti(m bays, n u m e r o u s small m o u n d s were seen (Fig 4). Amorphous later or zone. In all tile diseased specimens, a distinctive layer or zone appeared on the surface of tile c e m e n t u m . This layqr had a (lark, uniformly a m o r p h o u s appear371

JOURNAL OF ENDODONTICS I VOL 7, NO 8, AUGUST 1981

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Table * Summary of m o r p h o l o g i c comparison results. Comparison of normal and diseased cellular c e m e n t u m morphology according to the observed surface characteristics. T h e differences in the n u m b e r of projections and fibers b e t w e e n the two groups were statistically significant.

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ance (fig 5). In the majority of the specimens, the layer extended from the apex, gradually diminishing in a coronal direction along the fractured root surface. It was interrupted only by areas of resorption and seemed to occur subjacent to the lesion associated with the tooth. In one specimen, cellular cementum appeared to be deposited over the zone (Fig 6). "['he amorphous layer varied in thickness, but the widest measurement made was 4.5 p.m. In some specimens, Sharpey's fibers from the peridontal ligament were seen inserting into the layer (Fig 7).

372

Diseased teeth

Energi' dispersive x-ray spectrometiI" results. Analysis of the energy dispersive X-ray spectrometry photomicrographs showed a difference in the relative proportional content of calcium and phosphorus. The normal cellular cementum had relatively equal contents of calcium and phosphorus; whereas, the diseased cellular c e m e n t u m showed a relatively higher calcium content c o m p a r e d with phosphorus. The a m o r p h o u s layer showed a lower a m o u n t of calcium compared with phosphorus content and, tinally, compared with its underlying diseased cementum, the diseased cementum showed a much higher phosphorous content and less calcium.

DISCUSSION

Periapical intlammation may be produced by irritants located in the root canal system and in tile apical region. T h e effects of inflammation on c e m e n t u m are resorption and necrosis. T h e specitic role cementum plays in the periapical repair process remains unclear. O u r findings of increased projections and depressions in diseased c e m e n t u m seem to indicate an increase in calcification in the presence of inflammation. Further, the decrease in n u m b e r of fibers supports the observations of Boyde and Lester, '~ that the more fibrous the fractured surface, the less calci-

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Fig. 4 - Top, chseased specimen fractured through the apex showing resorptzon into canal (orig mag • 50). M*ddle, higher magmJicatwn of resorptzon area shou'mg small cemental mounds. (ortg mag • 500) Bottom, higher magn~catton of the mound area (or(e mag • ZOO0). C, cellular cementum; D, dentin; S, canal space; L, lateral canal, 17, resorption; M, eemental mounds.

Ftg. 5 - Top, fractured sur]ace through root canal qf diseased specimen sho~t:mg amorphous zone or la~.er (orzi~ mag X 50)..~ltddle, htgher magmfcatwn of amorphous I(oer m area 2 (ortg mag • Bottom, d(~erent specimen shmcmg amorphous ~'one or lq),er (ortie mae • 1,000, area 2). S, canal space; C, cellular cementum; A, amorphous zone or layer. 373

Fig. 6-Left..fractured surface q/&seased .7~ecmzen showing amorphou, zone u'uh apparent cellular cementum depo.~zted on its surface (oft R mag • 500, area 2). Rticht, hzgher ma~mfication of same area (or~,r ma.~ • 2,000, area 2). A. amorphous zone or layer, C, cellular cementum.

Fzg. 7-Another diseased speczmen showtug periodontal hgamentfbers insertmg into amorphoua zone (or~ mag • 5,000, area 2). A, amorphous zone: C, cellular ~ementum, F, ji~)gr.s

lied the collagenous matrix. T h e energy dispersive X-ray spectrometry showed an increase m the calcium and a decrease in phosphorous levels of diseased cellular cementum, and also supports the conclusion of increased calcification when cellular c e m e n t u m is exposed to the inflammatory environment. This may be comparable to the tindings associated with acellular cementuna exposed to periodontal disease where an increase in calcilication has linen reported in the form of a surface hypermineralized zone? ......... However, cellular c e m e n t u m is not exposed to the same factors, and the healing potential is very different. T h e observation o1" cemental resorption in all our diseased cellular c e m e n t u m specimens is in actor374

dance with the usual findings that in the presence of inflammation, resorption occurs.' "~: T h e small m o u n d s seen oil the resorptive areas were interpreted as deposition of new c e m e n t u m in an attempt at healing (Fig 4). Most resorption areas are apparently self-limiting and repair bv cemental deposition?"" T h e dense amorphous layer found in all the diseased specimens was not anticipated. Artifact due to processing was dismissed tx:cause the layer was not found on any of the normal specimens, Furthermore, Sharpey's fibers were seen inserting into the layer (Fig 7), and in one specimen, cellular c e m e n t u m was deposited on the layer (Fig 6). T h e possibility that this layer m a y represent aeellular cementum or pre-

cementum was considered. Acellular cementum, viewed under the SEM, is composed of numerous perpendicular fitxers or tibrils (Sharpcy's fibers) in a palisading uniform pattern? ~ The layer at similar magnilications did not reveal similar fibers or fibrils. No similarity was found in the photomicrographs of the laver compared with those of the precementum described by B o y d e " and Jones. '~ Precementum is seen as a laver uniformly composed of many collagen fibrils; whereas, while the laver in our study appears completely amorphous. ] ' h e amorphous layer is not analagous to the hypermiueralized zone found in acellular c e m e n t u m pathosis ........ because it is a hypomineralized zone. Fiber reattachment in periodontal therapy may be hindered by the hypermineralized layer. ltowever, periapical healing may be enhanced by allowing fiber reattachment into the hypomineralized layer. The layer may result from the complex inflammatory, enzymatic, and molecular influences that acc o m p a n y periapical inflammatory disease. These physical or chemical changes are reflected in the morphology of the apical cellular cementum.'"" "Whether these chemical changes produce an uptake of calcium in the underlying cellular cementum or a leaching out of cal-

J O U R N A l . OF E N D O I ) O N I I C 5

cium into the surrounding inflammatory lesion is unknown. In contrasl Io periodontal disease, apical inflammatory disease is a predictably reversible process. If dw root canal system can }xe successfully cleaned and obturatcd, healing can be anticipated. Because the layer appears to be a specific response by cellular cem('ntun~ to an intlammatory' environment, its signiticance may contribute to our understanding of disease and repair involving the root apex. SUMMARY

lesion associated with the tootil. The (]iseased cellular cementun! con,ained a higher Ca and lower P level than the normal specimens, as seen thru the energy dispersive X-ray Sl)cctror:mtry. T h e a m o r p h o u s laver or zone had higher P and lower Ca levels than its underlying diseased cellular c e m e n t u m . This atnorph<)us layer is a pathologic entity, and its signilicance at this time is unkllown.

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ative of normal and periapically diseased teeth, ',,,'as examined under the scanning electron microscope. A l l specimens were fractured and left untreated with the organic component intact to obtain a crt)ss sectiona[ view t)t" tile cellular c e n l e n t u m . ()ur findings were as fi)lh~ws: The diseased c e l l u l a r c e n l c n t illn contained more projections, more cementum lacunae, and fiev,'er tibers compared with normal cellular cementum, which indicated increased calcification. Ccmental tvsorf)tion w a s als() a c ( ) n s i s t e n I o b s e r v a t i o n . interrupted,

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layer or zone was seen in all the diseased specimens. It was located along the periphery of the fractured cellular (enleDtuiu and seemed to follow the outline of the per)apical

References 1 Rutx'n. M.. and Shapiro. A. An anal',sis tlf H,t)t sutl'acc changes in [x'rio(lt'tit)tlontal t:[is('ast" ,J IX.st R(.s 48:H-H)-HS:'). 1!)1i9 3..":,cl'.'i~. K.. and llal~,, E. l'('riodontall', db.t'a,,<:d C('llP,'tlttlm stt+t(Ji<'d b`. <+x)tlcLat,vd nlhrc-~radil)s }. elcrtron prt)lw anal,~sis, and (,l(.ctron mh'to'-,COl)~ .] Pcrh)dontal Res 12:,I I!)- t2!L 1977. 4 Boyd(', A., and Jones. s. ~.('annm~ eh'cIron nli('tosc()py Of c(~rtl('fltUlII arid +~llaflJe ) fibre b
V O L 7, N O 8, A U G U S t

1981

nficmscopc. Z Z c l l t ; . s c h 13():318-337. 1!)7:2. +i SMvi~, K An uhrastru(tural study of (t'il)('t)ilHll

t]Drn~ittil)li.

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22: 103-120. I!li>t. 7. S('l,,ig, K T h e tim" structure of human <-cmcntum .'k
375