Yonago Acta medica 2000;43:39–46

Three-Dimensional Structure of Connective Tissue Papillae in theHuman Gingiva
Tetsuya Yura, Hitoshi Osatake* and Takao Inoué*
Department of Oral and Maxillofacial Surgery and *Second Department of Anatomy, Faculty ofMedicine, Tottori University, Yonago 683-0826, Japan
Three-dimensional structures of connective tissue papillae of the human gingiva werestudied by scanning electron microscopy. Specimens were obtained from human cadavers fixed with 10% formaldehyde. The free gingiva, attached gingiva and alveolar
mucosa were excised from the incisor, canine and molar regions. They were further
fixed with 2.5% glutaraldehyde and treated with 2 N NaOH for 2 weeks at 20˚C to remove
the epithelium followed by routine specimen preparation methods. Consequently, theconnective tissue papillae were observed three-dimensionally and were classified into
three types as follows. Type I was an elongated papilla with a pointed tip, which wasusually observed in the free gingiva. Type II was a bud-like papilla whose tip wasforked into several tips and usually found in the attached gingiva. Type III was a platelike papilla chiefly located in the alveolar mucosa. In general, pointed elongated papillae(type I) in the free gingiva reduced in height changing into type II papillae towards theattached gingiva, and eventually sparsely distributed changing into type III papillae.
These findings were commonly observed both in the upper and lower jaws. Lots ofsmall holes, measuring 10 to 30 µm in diameter, were observed at the incisor and canineregions. Functional significance and origin of the holes remain unknown.
Key words: connective tissue papillae; human gingiva; scanning electron microscopy; NaOH
maceration method

The gingiva, which covers the neck of the toothand a part of the alveolar bone, is a soft tissue
preventing the invasion of foreign materials andbacteria into underlying tissues. It is grossly
divided into “free gingiva” and “attached gingiva”, which are separated from each other by afine line running parallel to the margin of thegingiva (Orban, 1948; Bollinger and Riethe,1973). Microscopically, it is composed of thestratified squamous epithelium, basal lamina
and underlying lamina propria. The basal lamina is located at the epithelium-connective tissueinterface which is undulated by connective tissue papillae.
The three-dimensional structures of theconnective tissue papillae were reconstructedby wax modeling of serial sections using lightmicroscopy (Karring and Löe, 1970; Löe andKarring, 1971). Since scanning electron
microscopy (SEM) was introduced to examine
the three-dimensional architecture of biologicalmaterials, it has been usefully applied to theobservations of gingival surfaces (Matraversand Tyldesley, 1977, 1978; Cleaton-Jones et al.,
1978; Hodgkins et al., 1978; Kullaa-Mikkonen1986). With the progress of techniques forcleaving the interface between the epithelium
and underlying lamina propria, connective tissue
papillae have been studied three-dimensionally(Klein-Szanto and Schroeder, 1977; Ooya andTooya, 1981). However, there have been no
precise studies which describe the morphological differences in various regions of the gingiva.

Abbreviation: SEM, scanning electron microscopy

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T. Yura et al.

abFig. 1. Scanning electron micrographs showing the surface of the lamina propria (a) and basal surface of theepithelium (b). The interface between the epithelium and lamina propria are cleaved by the NaOH digestion
method. Although the ultrastructures of the basal epithelial surface are well retained, the preservation of theconnective tissue papillae is relatively poor. a: × 40, b: × 40.

In this study, we studied various regions ofhuman connective tissue papillae in themaxillary and mandibular gingiva together withthe alveolar mucosa, by SEM after removing
the epithelium using some chemical digestion
techniques.

ide for 1 h followed by conductive-staining
with 2% tannic acid and 1% osmium tetroxide(Murakami, 1974). The specimens were then dehydrated in a graded series of ethanol, and finally dried in t-butyl alcohol (Inoué and Osatake,1988). After the dried specimens were mounted
on metal specimen stubs with conductive-glue,
they were sputter-coated with platinum at a
thickness of approximately 5 nm. Specimens
were observed and photographed by scanningelectron microscope (HFS-2ST or S-430S,Hitachi, Tokyo, Japan) at 25 kV.

Materials and Methods
Specimens were obtained from human cadavers(two males: 63 and 80 years old, one female: 50years old). These cadavers were initially fixedwith a 10% formalin solution through the rightfemoral artery. The gingiva was excised fromthe incisor, canine and molar regions together
with the alveolar mucosa in the left upper andlower jaws. The specimens were further cut
into three parts: free gingiva, attached gingiva
and alveolar mucosa. The size of eachspecimen was approximately 5 × 10 mm. They
were fixed again for an additional 24 h in 2.5%
glutaraldehyde buffered with 0.15 mol/L
phosphate buffer (pH 7.4) and rinsed for 1 h in
the same buffer. Some specimens were digested in 6 N NaOH at 60˚C for 10 min to cleave theepithelium-connective tissue interface (Inouéand Gabella, 1992). Other specimens weremacerated in 2 N NaOH for 2 weeks at 20˚C(Ohtani et al., 1988). After rinsing in the samebuffer, they were postfixed in 1% osmium tetrox-Results
When specimens were digested with 6 NNaOH, the epithelium-connective tissue
interface was cleaved. The superficial surfaceof the lamina propria was characterized by lotsof villous protrusions of connective tissue
papillae (Fig. 1a), but their preservation wasrelatively poor. The corresponding basal surface of the epithelium had lots of large holes
which had housed the connective tissue papillae(Fig. 1b).
When specimens were macerated with 2 N
NaOH, the epithelium was effectively removed
and the underlying connective tissue papillae
were demonstrated in three dimensions by SEM(Figs. 2 to 4).
40Connective tissue papillae of the gingiva

acb

Fig. 2. Scanning electron micrographs showing the three types of connective tissue papillae of the humangingiva prepared by the NaOH maceration method. The ultrastructure of the connective tissue papillae iswell preserved. a: Elongated papilla with pointed tip (type I). b: Bud-like papillae whose tips are forked into
several tips (type II ). c: Plate-like papillae whose tips are serrated (type III). a: × 160, b: × 95, c: × 90.

The connective tissue papillae showed
different shapes and sizes in their locations.
They were classified into types I, II and III as
follows (Fig. 2). Type I papillae were elongatedpapillae whose tips were usually pointed, but
occasionally bifurcated or three-forked (Fig.
2a). Type II papillae were lower than type I.
They appeared as bud-like configurations because their tips were usually forked into two to
five sections (Fig. 2b). Type III papillae exhibited plate-like elevations (Fig. 2c). Their freemargins were usually serrated, showing forklike configurations. They were sometimes
highly serrated, changing into several solitaryrod-like papillae.
Both in the upper and lower jaw, a pointed
configuration (type I) in the free gingiva, reduced in height, could be seen showing the bud-like
configuration of type II the farther they werefrom the gingival margin. The plate-like papillae of type III were usually found in the alveolar
mucosa.

area far from the free gingival margin, theirheight tended to be reduced and their tips became bifurcated and twisted. In the molar
region, the papillae were arranged in parallelrows along the gingival margin. They weredistributed at a density of 250 to 270/mm2 andno significant differences in density were noted
between the incisor, canine and molar regions.
In the attached gingiva, the papillae weremostly short (type II), showing a bud-likeappearance (Figs. 3b, e and h). Their width andheight measured 30 to 80 µm and 50 to 100 µm,respectively. They were distributed at thedensity of 90 to 150/mm2. In the incisor region,
the papillae tended to be arranged in rows
perpendicular to the gingival margin (Fig. 3b).
In the canine region, a cluster of type I papillae
was intercalated among the type II papillae(Fig. 3e).
In the alveolar mucosa, connective tissuepapillae were poorly developed. Type III papillae were scattered on a relatively flat superficial
surface of the lamina propria (Figs. 3a, d and g).
In the incisor region, typical connective tissue
papillae were not visible. Instead, ridge-likeelevations of the superficial layer of the lamina
propria were observed.
Small openings were sometimes visible
among the connective tissue papillae. They
usually occurred in the attached gingiva of theConnective tissue papillae in the upperjaw
In the free gingiva, connective tissue papillae
were slender, and pointed (type I) (Figs. 3c, fand i). Their width and height measured 30 to60 µm and 200 to 250 µm, respectively. In the41