Extracellular matrix mineralization in periodontal tissues: Noncollagenous matrix proteins, enzymes, and relationship to hypophosphatasia and X-linked hypophosphatemia.

As broadly demonstrated for the formation of a functional skeleton, proper mineralization of periodontal alveolar bone and teeth - where calcium phosphate crystals are deposited and grow within an extracellular matrix - is essential for dental function. Mineralization defects in tooth dentin and cementum of the periodontium invariably lead to a weak (soft or brittle) dentition in which teeth become loose and prone to infection and are lost prematurely. Mineralization of the extremities of periodontal ligament fibers (Sharpey's fibers) where they insert into tooth cementum and alveolar bone is also essential for the function of the tooth-suspensory apparatus in occlusion and mastication. Molecular determinants of mineralization in these tissues include mineral ion concentrations (phosphate and calcium), pyrophosphate, small integrin-binding ligand N-linked glycoproteins and matrix vesicles. Amongst the enzymes important in regulating these mineralization determinants, two are discussed at length here, with clinical examples given, namely tissue-nonspecific alkaline phosphatase and phosphate-regulating gene with homologies to endopeptidases on the X chromosome. Inactivating mutations in these enzymes in humans and in mouse models lead to the soft bones and teeth characteristic of hypophosphatasia and X-linked hypophosphatemia, respectively, where the levels of local and systemic circulating mineralization determinants are perturbed. In X-linked hypophosphatemia, in addition to renal phosphate wasting causing low circulating phosphate levels, phosphorylated mineralization-regulating small integrin-binding ligand N-linked glycoproteins, such as matrix extracellular phosphoglycoprotein and osteopontin, and the phosphorylated peptides proteolytically released from them, such as the acidic serine- and aspartate-rich-motif peptide, may accumulate locally to impair mineralization in this disease.

Temporal expression of metalloproteinase-8 and -13 and their relationships with extracellular matrix metalloproteinase inducer in the development of ligature-induced periodontitis in rats.

BACKGROUND AND OBJECTIVE: Matrix metalloproteinases (MMPs) play important roles in extracellular matrix degradation and may be regulated by extracellular matrix metalloproteinase inducer (EMMPRIN). The aim of this study was to investigate the temporal expression and localization of MMP-8 and MMP-13 during the development of ligature-induced periodontitis in rats, and to analyze the correlations of EMMPRIN with MMP-8 and MMP-13 in periodontitis. MATERIAL AND METHODS: Periodontitis was simulated in rats by ligaturing the cervix of the lower first molars, as described in our previous method. The rats were killed 0, 3, 5, 7, 11, 15 and 21 d after ligation. Micro-computed tomography examinations were performed to detect alveolar bone loss. Semiquantitative western blotting was used to assess the temporal changes in the levels of MMP-8, MMP-13 and EMMPRIN proteins in gingival tissue. Immunohistochemistry was applied to detect the expression and locations of MMP-8 and MMP-13 in gingival tissue and alveolar bone. RESULTS: Alveolar bone loss showed an exponential increase from days 3 to 11, followed by a slower rate of loss at subsequent study time points. MMP-8 showed a rapid increase of expression from baseline to a peak on day 3, a gradual decrease from days 5 to 7 and then stabilized thereafter. MMP-8 was predominantly located in neutrophil-like cells. Statistically, the expression of MMP-8 was not correlated with the expression of EMMPRIN. The expression of MMP-13 and of EMMRPIN increased from days 3 to 7, and showed a moderate decrease thereafter. The immunoreactivity of MMP-13 was mainly detected in monocytes/macrophages, on the alveolar bone surface, in osteoclasts and in gingival epithelial cells. Statistically, MMP-13 had a strong, positive correlation with EMMPRIN (r = 0.855, p < 0.01). CONCLUSION: The levels of expression of MMP-8 and MMP-13 are temporally varied at different periods during the development of experimental periodontitis. The level of expression of EMMPRIN is closely associated with the expression of MMP-13, but not with the expression of MMP-8. In addition, MMP-13 might be involved in alveolar bone destruction, as well as in physiological bone remodeling.

Matrix metalloproteinase-9 expression in alveolar extraction sockets of Zoledronic acid-treated rats.

PURPOSE: The use of nitrogen-containing bisphosphonates (n-bis) is associated with necrosis of the jaws, also known as bisphosphonate-related osteonecrosis of the jaws (BRONJ); however, the pathophysiology is unknown. Matrix metalloproteinase-9 (MMP-9) expression is essential for normal bone healing and is also required for angiogenesis. N-bis alters MMP-9 expression in vitro and in vivo; therefore, we hypothesized that n-bis alters MMP-9 expression during oral wound healing after tooth extraction. MATERIALS AND METHODS: A total accumulated dose of 2.25 mg/kg (n = 20) of Zoledronic acid (ZA) Zometa or saline (control, n = 20) was administered to Sprague-Dawley male rats. Next, both groups had maxillary molar teeth extracted. Rats were sacrificed at postoperative day 1, 3, 7, or 21. Western blotting or multiplex ELISA was used to evaluate proteins of interest. Real-time polymerase chain reaction was used to assess the relative quantities of target gene mRNA. MMP-9 enzymatic activity was assessed by zymography. RESULTS: The ZA group showed a statistically significant reduction in bone mineralization rate 21 days after tooth extraction compared with the control group (Student t test, P = .005). Moreover, ZA-treated animals showed a statistically significant increase in MMP-9-specific mRNA at postoperative days 3 (P = .003), 7 (P < .0001), and 21 (P < .0001) and protein on postoperative days 3 (P = .005) and 7 (P < .0001). MMP-9 enzymatic activity was also increased in ZA-treated rats compared with control animals (Student t test, P = .014). We also evaluated the extraction sockets for the presence of tissue inhibitor of MMP-1 (TIMP1), which is an inhibitor of MMP-9 enzymatic activity. TIMP1-specific mRNA and protein were not significantly altered by ZA treatment at the times tested (P > .05). Receptor of NF-κB ligand (RANKL) is known to regulate the expression of MMP-9; we therefore assessed the RANKL expression in our experimental oral wound-healing model. The ZA-treated animals had significantly increased RANKL mRNA at postoperative days 3 (P = .02) and 21 (P = .004), while the protein expression was significantly increased at postoperative days 1 (P < .0001), 7 (P = .02), and 21 (P = .03) compared with the control group. CONCLUSIONS: ZA reduced bone mineralization within tooth extraction sockets, suggesting aberrant bone healing. ZA increases the amount and enzymatic activity of MMP-9, while apparently not altering the amount of TIMP1 within extraction sockets. RANKL is increased in ZA-treated rats, which suggests that increased MMP-9 expression is due, in part, to augmented RANKL expression.

Healing of extraction sockets in collagenase-2 (matrix metalloproteinase-8)-deficient mice.

Matrix metalloproteinase-8 (MMP-8) participates in skin wound healing and inflammation. We hypothesized that MMP-8 plays a role in wound healing after tooth extraction and in periapical inflammation. Bone formation, collagen metabolism, and inflammation in tooth extraction socket and in periapical lesions were analyzed in wild-type mice and in MMP-8-deficient (MMP-8(-/-)) mice. New trabecular bone area in the extraction sockets and in periapical lesions were similar in both groups. In extraction sockets significantly more type III procollagen was synthesized, and the neutrophil and MMP-9 levels were lower in MMP-8(-/-) mice. The amount of Fas ligand, identified as a substrate for MMP-8, was lower in alveolar mucosa but higher in alveolar bone of MMP-8(-/-) mice. These results indicate that MMP-8 can modulate inflammation and collagen metabolism of alveolar bone and mucosa.

MMP-3 response to compressive forces in vitro and in vivo.

During orthodontic tooth movement, bone resorption occurs at the compression site. However, the mechanism underlying resorption remains unclear. Applying compressive force to human osteoblast-like cells grown in a 3D collagen gel, we examined gene induction by using microarray and RT-PCR analysis. Among 43 genes exhibiting significant changes, cyclo-oxygenase-2, ornithine decarboxylase, and matrix metalloproteinase-3 (MMP-3) were up-regulated, whereas membrane-bound interleukin-1 receptor accessory protein was down-regulated. The MMP-3 protein increases were further confirmed by Western blot. To ascertain whether MMP-3 is up-regulated in vivo by orthodontic force, we examined human bone samples at the compressive site by realigning the angulated molars. Immunohistochemical staining revealed MMP-3 distributed along the compressive site of the bony region within 3 days of compression. Since MMP-3 participates in degradation of a wide range of extracellular matrix molecules, we propose that MMP-3 plays an important role in bone resorption during orthodontic tooth movement.

Histological and histomorphometrical changes in rat alveolar bone following antagonistic tooth extraction and/or ovariectomy.

OBJECTIVE: To examine changes appearing in the alveolar bone following the removal of the mechanical stress of occlusal loading, as well as the added influence of estrogen deficiency on such changes. DESIGN: The right mandibular molars of female rats were extracted. After 8 weeks, 12 animals were ovariectomized (OVX), and the other 12 were subjected to sham surgery (sham). Four weeks after surgery, all rats were sacrificed. The left-half and right-half maxillas of the sham group (the sham-occluded side and the sham-extruded side, respectively) and right-half maxilla of OVX group (the OVX-extruded side) were examined by histological observation and bone histomorphometry. RESULTS: The vertical height of alveolar bone in the sham-extruded and the OVX-extruded sides increased as compared with that of the sham-occluded side. In both extruded sides, active bone formation occurred on the surface of the alveolar bone facing the periodontal ligament, but the bone marrow was expanded and the bone volume had decreased in the internal area of the alveolar bone. In the OVX-extruded side, the bone marrow expanded more remarkably than that of the sham-extruded side, and the highest percentage of osteoclast surface was detected. CONCLUSIONS: Around the extruded teeth, there were regional differences in bone dynamics between the internal area of the alveolar bone and the bone surface facing the periodontal ligament, and estrogen deficiency seems to have caused further loss of bone volume in the interior of the alveolar bone supporting the extruded tooth.

Tartrate-resistant acid phosphatase in mononuclear and multinuclear cells during the bone resorption of tooth eruption.

Tartrate-resistant acid phosphatase (TRAP) has been used as a cytochemical marker for the cell mediators of bone resorption, osteoclasts and their mononuclear precursors. We have applied a cytochemical method for TRAP to study the dependence of the osteoclast-mediated bone resorption of tooth eruption on the dental follicle, a connective tissue investment of the developing tooth, by analyzing the TRAP activity of mononuclear cells in the dental follicle before and during pre-molar eruption in dogs. The percentage of TRAP-positive monocyte cells increases until mid-eruption, slightly preceding a previously demonstrated rise in numbers of osteoclasts on adjacent bone surfaces. These data suggest an ontogenetic relationship between follicular mononuclear cells and osteoclasts on adjacent alveolar bone surfaces during tooth eruption. However, because TRAP occurs in other tissues and is not an exclusive indicator of pre-osteoclasts, proof of their relationship will have to await application of more definitive techniques.

A quantitative enzyme histochemical analysis of the distribution of alkaline phosphatase activity in the periodontal ligament of the rat incisor.

The spatial distribution of alkaline phosphatase (ALP) activity was examined in the periodontal ligament of the continuously growing rat incisor. With the indoxyl-tetrazolium salt method, enzyme activity was demonstrated in undecalcified cryosections, and the amount of reaction product was quantified. ALP activity appeared to be distributed heterogeneously. Its highest activity was found in the bone-related compartment of the ligament. In the tooth-related compartment and the supracrestal extension of the ligament, enzyme activity was significantly lower, but still higher than in the lamina propria of the gingiva. In the part of the ligament bordering the cementum, highest activity was found in the apical region just occlusal to Hertwig's epithelial root sheath, where formation of acellular cementum begins. From there toward the incisal edge, the activity of the enzyme gradually decreased. It is suggested that differences among the various parts of the periodontal ligament are related to local variations in phosphate metabolism and cementum deposition.

Changes in the tartrate-resistant acid phosphatase cell population in dental follicles and bony crypts of rat molars during tooth eruption.

It was the aim of this study to determine the cellular changes that occur in the enamel organ, dental follicle, and surrounding bony crypt of the rat molar prior to and during tooth eruption. By use of light microscope histochemistry to detect cells containing tartrate-resistant acid phosphatase (TRAP), it was seen that TRAP-positive mononuclear cells were present in the dental follicle prior to the onset of eruption (e.g., three days postnatal age) and then declined in number during eruption. Concurrently, TRAP-positive osteoclasts were initially present in large numbers on the surface of the bony crypts surrounding the molars (three days postnatal age) and then declined in number as eruption progressed. Electron microscopy confirmed that these were mononuclear cells and osteoclasts. The results suggest that the mononuclear cells are either precursors of the osteoclasts or perhaps release cytokines that affect osteoclast formation or activity. Staining for alkaline phosphatase (ALP) activity indicated that at an early postnatal age (secretory stage of amelogenesis), ALP was detected only in the stratum intermedium of the enamel organ, whereas at a later age (maturation phase of amelogenesis), it was present only in the ameloblasts. These results, combined with a survey of the literature, strongly suggest that ALP moves from the base of the enamel organ to the enamel itself over a period of time ranging from pre- to post-eruption. Rat molars are teeth of limited eruption, and the cellular events that occur in eruption appear comparable with what is seen in dog and human dentition, especially in terms of the cellular events seen in the dental follicle prior to and during eruption.(ABSTRACT TRUNCATED AT 250 WORDS)

Cyclic AMP in dental and periodontal tissues during tooth eruption in kittens.

Cyclic adenosine 3',5'-monophosphate (cAMP) concentrations and cellular distribution were studied in dental and periodontal tissues during tooth eruption in kittens. Although the mean levels of cAMP around developing teeth were similar in all the tissue samples, there were marked differences in cAMP stainability of tissues apical and occlusal to the erupting teeth.

Distributional changes of osteoclasts and pre-osteoclastic cells in periodontal tissues during experimental tooth movement as revealed by quantitative immunohistochemistry of H(+)-ATPase.

To investigate the mechanism of alveolar bone remodeling in response to orthodontic force application, we examined the distribution of osteoclasts and pre-osteoclastic cells using quantitative immunohistochemistry of vacuolar type H(+)-ATPase. For orthodontic force to be produced by the Waldo method, an orthodontic elastic band was inserted between the upper first and second molars of rats. The observed areas of periodontal tissues around second molars were the distal surfaces of mesial roots, as the pressure side, and the mesial surfaces of distal roots, as the tension side. Specific expression of vacuolar-type H(+)-ATPase at the ultrastructural level was detected in mononuclear and multinucleated pre-osteoclastic cells, as well as osteoclasts with ruffled borders on bone surfaces. At 6 hrs after orthodontic force application, many osteoclasts and pre-osteoclastic cells with H(+)-ATPase expression were first observed in vascular canals of the alveolar bone crest near the pressure side of the periodontal ligament, but the number of osteoclasts was not increased in the periodontal ligament. On day 1 after tooth movement, osteoclasts were increased in number in the periodontal ligament and in adjacent alveolar bones on the pressure side, but were seldom observed in corresponding areas on the tension side. The number of osteoclasts increased until day 7, but had decreased by day 14. These results suggest that, in bone remodeling during experimental tooth movement, (1) osteoclasts and pre-osteoclastic cells can be identified by H(+)-ATPase immunohistochemistry, (2) osteoclasts and pre-osteoclastic cells are rapidly induced after force application, (3) osteoclast induction first occurs in vascular canals of the alveolar bone crest on the pressure side, and then, (4) the number of osteoclasts increases in the periodontal ligament on the pressure side.

Effect of a pulsing electromagnetic field on demineralized bone-matrix-induced bone formation in a bony defect in the premaxilla of rats.

A 2-mm non-healing bony defect was prepared in the premaxilla of male Wistar rats weighing about 180 g as a simulation of an alveolar cleft, for determination of whether a pulsing electromagnetic field (PEMF) could promote regeneration of bone induced by demineralized bone matrix (DBM). The defect was either treated with 7 mg DBM or was left as a non-grafted control. The rats were exposed to a PEMF with a frequency of 100 Hz, a 10-ms-wide burst with 100 microseconds-wide quasi-rectangular pulses, repeating at 15 Hz, and magnetic field strength of 1.5-1.8 G. Alkaline phosphatase activity increased significantly from day 7 in the DBM-graft-plus-PEMF group and from day 10 in the DBM-graft group, reaching a maximum on day 14. A greater-than-two-fold rise in alkaline phosphatase activity and a three-fold rise in the amount of 45Ca incorporation in the DBM-graft-plus-PEMF group were attained compared with those of the DBM-graft group. The DBM-graft-plus-PEMF group produced more bone with almost complete osseous bridging in the defect sites than did the group treated with DBM only on day 35. The findings indicate that PEMF had an enhancing effect on the bone-inductive properties of the DBM through the stimulation of osteoblast differentiation induced by DBM.

Localization of the NO-cGMP signaling pathway molecules, NOS III-phosphorylation sites, ERK1/2, and Akt/PKB in osteoclasts.

BACKGROUND: Nitric oxide (NO) mediates different cellular functions by activating soluble guanylate cyclase (sGC) that converts guanosine-5'-triphosphate (GTP) to cyclic guanosine-3',5'-monophosphate (cGMP). Membrane-bound GCs produce cGMP in response to natriuretic peptides in osteoblasts, but neither the NO-target enzyme sGC, nor the phosphorylation sites of NOS III, nor their regulation by extracellular signal-regulated kinases 1 and 2 (ERK1/2) and Akt/protein kinase B (Akt/PKB) in osteoclasts have been established. METHODS: Rat molars with periodontium were perfusion- and post-fixed, decalcified, and frozen-sectioned. Free-floating sections were stained using nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) and tartrate-resistant acid phosphatase (TRAP) histochemical techniques and immunoreacted with antisera against NO-synthase (NOS) I-III, NOS III phoshorylated at Thr495, NOS III phoshorylated at Serine1177 (Ser1177), ERK1/2, phosphorylated ERK1/2, Akt/PKB, phosphorylated Akt/PKB, sGC (alpha2/beta1), and cGMP. RESULTS: NADPH-d staining and immunostaining of NOS I-III, NOS III phosphorylated at Ser1177, ERK1/2, Akt/PKB, phosphorylated Akt/PKB, sGC (alpha2 and beta1-subunits), and cGMP were detected in osteoclasts. Immunohistochemical reaction products for NOS III phosphorylated at threonine495 (Thr495) and phosphorylated ERK1/2 could not be identified in osteoclasts. Comparison of TRAP activity and immunostaining for sGC beta1-subunit revealed that sGC beta1-subunit is only expressed in a sub-population of osteoclasts. CONCLUSIONS: NO is likely to be generated by NOS I and NOS III in osteoclasts. The inconstant expression of NOS II in some osteoclasts may be explained with inducible expression of NOS II upon physiological cell activation. Localization of the sGC alpha2- and beta1-subunits and cGMP in osteoclasts is compatible with an involvement of NO-sGC signaling in the homeostasis of osteoclasts. The phosphorylation site of NOS III at Ser1177 and phosphorylated Akt/PKB are involved in regulation of NO production by NOS III in osteoclasts under basal conditions.

Should cementoblasts express alkaline phosphatase activity? Preliminary study of rat cementoblasts in vitro.

BACKGROUND: A well-characterized cell culture model for cementoblasts is essential to understand the mechanisms of periodontal ligament (PDL) reattachment and regeneration. Whether cementoblasts express alkaline phosphatase (ALP) activity in vivo and in vitro remains to be determined. METHODS: Using a 2-step method of enzyme digestion/explant culture, osteoblasts, gingival/PDL fibroblasts, and cementoblasts were obtained from alveolar bone, gingiva, and the root surface of rat first molars and cultured. Initially, bone sialoprotein (BSP) was immunolocalized on tissue sections of periodontium and on cultured cells to distinguish mineral-forming cells from fibroblasts. Proteins were extracted from these cells to assess ALP activity by using an enzyme assay. RNA was extracted from the same cell source to detect ALP mRNA by reverse transcriptase polymerase chain reaction (RT-PCR). RESULTS: Cultured PDL/gingival fibroblasts were spindle shaped. Osteoblasts were irregularly shaped, and cell clusters/nodules were observed as they approached confluence. The cementoblasts manifested a polygonal shape and had two morphotypes: osteoblast-like and cuboidal or stratified. BSP was localized within the mineralized tissues and in osteoblasts and cementoblasts in culture and in tissue sections. The highest level of ALP activity was found in osteoblasts, a moderate level in PDL fibroblasts, and the lowest level in gingival fibroblasts. The cementoblasts lacked ALP activity, and this was reflected by a very weak signal (or no signal at all) for ALP mRNA in the cementoblasts. CONCLUSIONS: These studies indicate that cells consistent with a cementoblast-like phenotype may be successfully cultured, and that they lack ALP activity.

Alkaline phosphatase in the periodontal ligament of the rabbit and macaque monkey.

This enzyme's specific activity was high in the periodontal ligament with other tissues in both species, but much higher in the rabbit. The alkaline phosphatase in the rabbit ligament is a liver/kidney/bone form and not an intestinal one.

Expression of cathepsin K mRNA during experimental tooth movement in rat as revealed by in situ hybridization.

The expression of cathepsin K. a novel collagenolytic enzyme specifically expressed in osteoclasts, was investigated in the rat maxillary dentoalveolar unit during experimental tooth movement by in situ hybridization histochemistry with a non-radioisotopic cRNA probe for rat cathepsin K. Orthodontic elastics were inserted into the interproximal space between the maxillary first and second molars of 7-week-old male SD rats according to Waldo's method and sections prepared from tissues obtained at 12 hr, 1, 2, 3, 4, 7, and 12 days after orthodontic force application. Cathepsin K mRNA expression was detected in the mono- and multinuclear osteoclasts on the pressure side of the alveolar bone at 12 hr after force application, and the distribution and number of cathepsin K mRNA-positive osteoclasts increased time-dependently on the pressure side. At 3-4 days, a marked increase in cathepsin K mRNA-positive osteoclasts was found not only on the pressure side but also on the tension side of the alveolar bone in response to tooth movement. At 7-12 days, the cathepsin K mRNA-positive osteoclasts on both sides had disappeared. These findings suggest that the recruitment of osteoclasts on the pressure side begins during the initial stage of orthodontic tooth movement and the site-specific early induction of cathepsin K mRNA may cause an imbalance in the relative resorption activities on the pressure and tension side incident to such movement.

Histochemical demonstration of acid phosphatase activity in terminal Schwann cells associated with Ruffini endings in the periodontal ligament of rat incisors.

Histochemical staining for acid phosphatase, a marker for lysosomal elements, distinguished rounded, intensely reactive cells from less reactive fibroblasts and osteoblasts in the lingual periodontal ligament. The highly reactive cells were located exclusively in the alveolar half of the ligament. Double staining for acid phosphatase and S-100 protein confirmed that these reactive cells were identical with the terminal Schwann cells associated with periodontal Ruffini endings. Electron microscopically, reaction products for acid phosphatase were observed in the lysosomes and Golgi apparatus in the paranuclear cytoplasm of the terminal Schwann cells. As the terminal Schwann cells associated with the Ruffini endings are assumed to be capable of synthesizing exportable proteins, acid phosphatase in this type of cell may be involved in the processing of macromolecules in synthetic and/or secretory pathways.

Histomorphometric and biochemical study of osteoclasts at orthodontic compression sites in the rat during indomethacin inhibition.

Prostaglandins affect the number of osteoclasts at compression sites in orthodontic tooth movement. They may also have a role in tooth movement and influence the extent of root resorption. The purpose was to examine the effect of indomethacin on the activity of resident osteoclasts, recruitment of new osteoclasts and root resorption at orthodontic compression sites. Two separate populations of osteoclasts were studied: those resident at the sites after initial appliance activation and those recruited by a subsequent activation. Orthodontic appliances were activated to provide mesially directed forces of 40 g on the maxillary molars of rats. The appliances were activated with the same force after 4 days. The rats were killed at 1, 3, 6 and 10 days after initial activation. Half of the rats were injected with indomethacin. Tooth movement was measured cephalometrically; osteoclast numbers, sizes, numbers of nuclei per osteoclast and root resorption were assessed histomorphometrically; tartrate-resistant acid phosphatase (TRAP) in alveolar bone was measured biochemically. Indomethacin inhibited both initial tooth displacement and that following the delay. It also reduced the increase in osteoclast numbers, total osteoclast surface and alveolar bone TRAP at day 10. It had no effect on the surface area of each individual osteoclast or number of nuclei in each osteoclast. Root resorption increased in both groups but it was enhanced at day 10 in the indomethacin group. These data suggest that orthodontic tooth movement after appliance activation requires the recruitment of osteoclasts to sites of compression and that this is indomethacin-sensitive. Furthermore, indomethacin enhances root resorption at compression sites 10 days after appliance reactivation.

[Localization of beta-glucuronidase in human mouth mucosal epithelium]. / Lokalisation der beta-Glucuronidase im Mundschleimhautepithel des Menschen.

The authors investigated the activity of beta-glucuronidase in the mucous membrane of the alveolar process and in the buccal mucous membrane under light and electron microscopes. Demonstration of the enzyme was performed by means of a modification of the method of Hayashi. Under the light microscope enzymatic activity was demonstrated in both types of epithelium, particularly in the fibroblasts of connective tissue, higher activity being found in vascular walls, inflammatory infiltrates and particularly in macrophages. Findings in the epithelium were positive only in the stratum basale and the lower layers of the stratum spinosum. Submicroscopically activity of the enzyme was apparent in all layers of the epithelium, particularly in primary and secondary lysosomes and in cisternae of the granular endoplasmic reticulum, and only rarely in Golgi complex. There were only a few positive structures in the surface layers of the epithelium. Activity was lower in the buccal mucosa than in the alveolar epithelium. In the mucosa of the oral cavity the enzyme probably has a role in the degradation of proteoglycans.

Quality of periodontal healing. IV: Enzyme histochemical evidence for an osteoblast origin of reparative cementum.

The aim of the present study was to map the distribution of healing gingival and periodontal connective tissue well as osteoblastic activity in marginal periodontal wound healing using activity of alkaline phosphatases as markers. Activity of non-specific alkaline phosphatase (NSAP) was abundant in periodontal but not in gingival connective tissue. Activity of vanadate-resistant alkaline phosphatase (VRAP) had a preferential localisation to periosteal and endosteal bone surfaces but was not detectable in cementoblasts along root surfaces. However, following spontaneous marginal healing of exposed dentine surfaces, activity of VRAP was also found on healing dentine surfaces. It was concluded that osteoblasts rather than cementoblasts form reparative cementum on exposed dentine surfaces during marginal periodontal healing.