An immunohistochemical study of osteoprotegerin in the human dental pulp.

This study investigated the expression of osteoprotegerin (OPG) in healthy and inflamed dental pulps. Histological sections 7 microm thick of 47 teeth, either caries-free or affected by gross caries, were used. Sections were stained with hematoxylin-eosin, and other sections of the same specimen were subjected to the avidin-biotin peroxidase complex immunohistochemical procedure for detection of OPG. The study focused on the coronal pulp that was divided into peripheral and central regions. In the peripheral pulp healthy and inflamed specimens showed high OPG immunoreactivity of the odontoblastic layer. When no inflammation was present in the central pulp OPG immunoreactivity was light. Fibroblasts and endothelial cells showed immunoreactivity ranging from none to intense. When inflammation was present in the central pulp the chronic inflammatory cells showed intense immunoreactivity.

Tissue reactions after subcutaneous and intraosseous implantation of mineral trioxide aggregate and ethoxybenzoic acid cement.

Biocompatibility of mineral trioxide aggregate and ethoxybenzoic acid cement was investigated by subcutaneous and intraosseous implantation of the materials in rats. Tissue reactions were studied at 15, 30, and 60 days after implantation. Subcutaneous implantation of mineral trioxide aggregate initially elicited severe reactions with coagulation necrosis and dystrophic calcification; the reactions, however, subsided to mostly moderate with time. Subcutaneous implantation of ethoxybenzoic acid cement initially elicited mostly moderate reactions that subsided to mild in time. Osteogenesis was not observed with either material upon subcutaneous implantation indicating that neither material is osteoinductive. Reactions to intraosseous implants of both materials were less intense than with subcutaneous implantation. Osteogenesis occurred in association with intraosseous implants indicating that both materials are osteoconductive.

Effect of hydroxyapatite content on physical properties and connective tissue reactions to a chitosan-hydroxyapatite composite membrane.

The present study investigated the effect on certain physical properties of adding various amounts of hydroxyapatite (HAP) to chitosan sol. Also investigated were connective tissue reactions to a composite membrane that is being developed for possible use in guided tissue regeneration and for the limitation of HA particle migration at sites of implantation. The physical properties evaluated were shrinkage, tensile strength, hardness, calcium ion release, and morphology. Assessment of physical properties indicated that a ratio of HA to chitosan sol of 4/11 by weight is optimal in the preparation of the composite membrane. Subperiosteal implantation of the membranes over rat calvaria revealed that the membranes were well tolerated, with fibrous encapsulation and occasional areas of osteogenesis. Increasing the hydroxyapatite content seems to enhance membrane degradation.

Histopathological and immunohistochemical studies of membranes of deacetylated chitin derivatives implanted over rat calvaria.

Membranes made of 65, 70, 80, 94, and 100% deacetylated chitin (chitosan) were implanted subperiosteally over the calvaria of 100 rats. Reactions were studied at 1, 2, 4, and 8 weeks after implantation. Membranes prepared with 65, 70, and 80% deacetylated chitin initially elicited marked inflammatory reactions that subsided in time with granulation tissue formation and osteogenesis. Osteocalcin-positive cells were detected immunohistochemically in the granulation tissue. On the other hand, membranes made of 94% deacetylated chitin and chitosan showed mild inflammation and minimal osteogenesis. The results indicate that membranes made of 65, 70, and 80% deacetylated chitin enhance osteogenesis at the site of their implantation. However, the initially severe inflammatory reaction associated with these materials needs to be controlled before the materials would be suitable for clinical application.

Oral lichen planus: an immunohistochemical study of heat shock proteins (HSPs) and cytokeratins (CKs) and a unifying hypothesis of pathogenesis.

The expression of heat shock proteins HSP60 and HSP70 and cytokeratins CK1/10 and CK7/18 were compared in epithelium of oral lichen planus (OLP) lesions and oral fibromas using an avidin-biotin-peroxidase complex (ABC) immunohistochemical method. An immunostaining intensity distribution (IID) index was developed to assess staining intensity and the proportion of positively stained cells in different layers of the epithelium. The expression of HSP60 in the basal layer was significantly higher in OLP than in fibromas. No difference in HSP70 expression was evident between OLP and fibromas. The expression of CK1/10 in the epithelial basal and suprabasal layers was significantly higher in OLP than in fibromas. There was no demonstrable staining for CK7/18 in either OLP or fibromas. A significant correlation was evident between the expression of HSP60 and CK1/10 in the basal epithelial cells in OLP. The findings support a role for HSP60 in the pathogenesis of OLP. A unifying hypothesis of the pathogenesis of OLP, involving two sequential immune reactions, is proposed.

Immunohistochemical study of the expression of human chorionic gonadotropin-beta in oral squamous cell carcinoma.

BACKGROUND: Human chorionic gonadotropin (hCG) is a glycoprotein hormone comprised of two dissimilar subunits (alpha and beta) and normally is synthesized by trophoblastic tissue. Although hCG expression has been identified in a variety of neoplastic tissues, to the authors' knowledge no investigation has centered on tumors of oral origin. METHODS: Oral squamous cell carcinomas (OSCC) were studied in comparison with oral fibromas for the presence of hCGbeta using the avidin-biotin-peroxidase complex immunohistochemical technique. RESULTS: hCGbeta immunoreactivity was identified in 29 of 45 OSCC (64%). The positively staining cells in each tumor specimen were few (range, 0.5-5%) and were scattered throughout the tumor. When tumors were classified according to grade, it was found that hCGbeta staining was positive in 5 of 15 well differentiated OSCC (33%), in 12 of 15 moderately differentiated OSCC (80%), and in 12 of 15 moderately to poorly differentiated OSCC (80%). hCGbeta immunoreactivity could not be demonstrated in any of the oral fibromas. CONCLUSIONS: The presence of hCGbeta positive tumor cells appears potentially to reflect a malignant behavior of OSCC.

Biocompatibility of two apatite cements.

Biocompatibility and osteogenic potential of two calcium phosphate cements (G-5 and G-6) and Super-EBA were investigated by subcutaneous and intraosseous implantation in 90 rats. Reactions were studied microscopically at 15, 30, and 60 days after implantation. Super-EBA was well tolerated by both soft and hard tissues. G-5 was highly biocompatible with resorption and bone replacement at intraosseous implantation sites. G-6 promoted moderate inflammation and a foreign body giant cell response over the 60-day study period. None of the materials elicited osteogenesis or dystrophic calcification at the subcutaneous implantation sites.

An in vitro comparison between laser fluorescence and visual examination for detection of demineralization in occlusal pits and fissures.

It has been demonstrated that when excited by laser light carious enamel appears dark compared to luminescent sound enamel. The aim of this study was to compare the sensitivity and specificity of visual exams (V), laser fluorescence (LF) and dye-enhanced LF (DELF) for detecting demineralization in occlusal pits and fissures. The actual presence of lesions was determined by subsequent confocal laser microscopy (CM), which was compared to histology (H). Independent clinical examiners visually graded three sites on occlusal surfaces of extracted, human premolars as sound or carious and also rated the color of each graded site as: 0 = same as surrounding enamel; 1 = white; 2 = light brown, or 3 = brown/dark brown. An argon laser was used to illuminate the teeth for LF and DELF; the images were captured with a CCD camera and then analyzed. DELF images were captured after the teeth had been exposed to 0.075% sodium fluorescein. Sections were then cut from each specimen and analyzed by CM and H for the presence or absence of caries. Results showed that DELF (0.72) was significantly more sensitive (p<0.05) than LF (0.49) and V (0.03) for detecting caries, but there were no significant differences among the methods in specificity (V 1.00; LF 0.67; DELF 0.60). When color was used as an indication of caries in V (VC, sensitivity 0.47; specificity 0.70), V exams were not different from LF. The area under the ROC curve, using H as the gold standard and CM as the test, was 0.78. Results indicated that DELF was the best diagnostic tool and that VC and LF were equally effective as diagnostic methods, when color of fissures was included as an indication of demineralization in the visual exam.

Immunohistochemical study of gamma-aminobutyric acid and bombesin/gastrin releasing peptide in human dental pulp.

This study investigated the presence of the putative peripheral neuromodulators Gamma-aminobutyric Acid (GABA) and Bombesin/Gastrin-Releasing Peptide (BN/GRP) in the human tooth pulp. Caries free and asymptomatic carious teeth were processed for paraffin embedding and sectioned at six microns. From each specimen, sections were stained with hematoxylin and eosin; other sections were subjected to Avidin-Biotin-Peroxidase Complex immunohistochemistry for GABA and BN/GRP. Sections of rat brain and small cell lung carcinoma served as positive controls. Results indicate the presence of specific GABA-like and BN/GRP-like immunoreactivity within the pulps of both normal and carious teeth. Overall staining for both ligands was significantly more intense within inflamed pulps. Based on their actions elsewhere, GABA and BN/GRP may play a role in the dental pulp as peripheral neuromodulators or as growth factors.

In vitro evaluation of effects of ultrasonic root-end preparation on resected root surfaces.

This in vitro study investigated structural alterations in resected roots that had root-end preparations made with a conventional microhead handpiece and ultrasonics at two intensity levels. Root ends were examined with fluorescence confocal microscopy. Serial histologic sections to the 2 mm levels and then at the level of 3 mm and 4 mm from the resected surface were examined. Statistical analysis of the confocal data indicated no significant difference between the groups in the number and length of the fractures. Results of the histologic data indicated that root ends prepared by ultrasonics had a statistically greater number of fractures than both the control and the conventionally prepared groups. The latter did not differ significantly from each other.

In vitro properties of a chitosan-bonded bone-filling paste: studies on solubility of calcium phosphate compounds.

The present study investigated properties of various mixtures of organic acids (malic and malonic) and calcium phosphate compounds (beta-tricalcium phosphate, ashed bovine bone, and synthetic hydroxyapatite) with the objective of determining the optimum combination of organic acid and calcium phosphate compound for components of a chitosan-bonded bone-filling paste. beta-tricalcium phosphate was decomposed by malic acid and malonic acid, but these two acids did not decompose synthetic hydroxyapatite and ashed bovine bone. Assessment of ion release from a set paste containing either synthetic hydroxyapatite or ashed bovine bone indicated that only calcium ions were appreciably released after storing and stirring the set paste in physiologic saline for 7 days.

Absence of detrimental effects of fluoride exposure in diabetic rats.

This study is part of a comprehensive programme to investigate fluoride toxicity and the hypothesis that fluoride ingested by "medically compromised' animals will result in altered physiological function. Its objectives were to monitor fluoride retention, tissue fluoride concentrations and genetic variables in diabetic and control rats chronically exposed to fluoride, and to determine whether or not adverse effects occurred. Male, Zucker fatty diabetic rats and Zucker age-matched lean controls were fed a low-fluoride diet ( < 1.2 parts/10(6) F-) ad libitum and received 0, 5, 15 or 50 parts/10(6) fluoride in their drinking water for 3 or 6 months. Fluoride metabolic balance was determined for 4 days before the end of each study phase. Plasma and urine were analysed for biochemical markers of tissue function, and plasma, urine, faeces and tissues were analysed for fluoride. Bone marrow cells from animals killed after 6 months of treatment were examined for frequency of sister chromatid exchange, a marker of genetic damage. The diabetic rats consumed, excreted and retained significantly greater amounts of fluoride than the controls (p < 0.05). There were dose-related increases in fluoride excretion, retention and tissue concentrations in both classes of animals, which were significantly greater in the diabetic rats. In spite of greater amounts of fluoride in the tissues of diabetic animals, there was no evidence, under these experimental conditions, that any of the fluoride exposures tested caused measurable adverse effects on the physiological, biochemical or genetic variables that were monitored.

Turner's syndrome in association with hemangioma of the parotid gland: report of case.

We report a case of Turner's syndrome with hemangioma of the parotid gland. The symptom complex of this syndrome are infantilism, pterygium colli and unusual carrying angle of the elbow. Orodental alterations and multisystem afflictions have been reported in these patients. Diagnosis is based on a combination of the clinical features, buccal smear finding of the absence of barr body and confirmed by karyotyping which can detect mosaic cell lines of the X chromosome.

Detection of mutans streptococci in secondary carious lesions using immunofluorescent techniques and confocal laser scanning microscopy.

Secondary caries is one of the major reasons causing restoration failure; however, little is known of its microbial etiology, mainly because of the difficulties in eliminating bacterial contamination during collection and sample preparation. Therefore, the purpose of this study was to investigate the feasibility of immunofluorescent techniques and confocal laser scanning microscopy for identification and quantification of bacteria in secondary carious lesions. Thirty-six extracted human teeth, clinically diagnosed as having secondary caries, were used in the study. The teeth were sectioned in half across the secondary carious lesion. One half of each tooth was processed using the Brown and Hopps histologic staining method for bacterial detection. Sections (100 microns thick) were obtained from the other half of each tooth for immunofluorescence labeling to detect and identify mutans streptococci in the subsurface of the lesion using confocal imaging techniques. Mutans streptococci were detected in 88.9% of the samples analyzed with the immunofluorescent technique. The Brown and Hopps histopathologic examination demonstrated evidence of bacteria in 94.4% of the samples. In addition, quantification of bacteria was conducted by digitalization of confocal images. The results indicated that the immunofluorescence confocal laser scanning microscopy technique was sensitive and specific for detection and quantification of mutans streptococci in secondary carious lesions.

Pulp reactions to light-cured glass ionomer cements.

This study compared the pulp responses to two light-cured glass ionomer liners with a chemically set glass ionomer lining cement and zinc oxide-eugenol. Deep Class V cavities were prepared at high-speed with carbide burs under a water coolant in 96 teeth of three young adult beagle dogs. The smear layer was left intact according to the manufacturers' recommendations. The cavities in each quadrant were lined with either Vitrebond light-cured glass ionomer liner, X-R Ionomer light-cured liner, GC lining cement or ZOE which served as a control. All cavities were restored with Silux Plus resin composite. The ZOE liner was covered with a varnish prior to the resin composite placement. The animals were terminated at 1, 4 and 12 weeks. Jaws were excised, fixed in 10% formalin and decalcified in 5% formic acid. Teeth were sectioned at 7 microns, and stained with hematoxylin and eosin. Selected slides were stained with the Brown and Hopps method for histologic demonstration of bacteria. The microscopic findings indicated that pulpal reactions to the two light-cured glass ionomer liners were minimal and comparable to those elicited by the chemically-cured glass ionomer lining cement and ZOE. Bacteria were observed in nine specimens but only one of these revealed a moderate pulp reaction.

Vascular neoplasms of the parotid gland. Parotid vascular tumors.

Vascular neoplasms of the parotid gland are common in early childhood, particularly in females. We reviewed the clinical, histologic, and treatment details of 10 cases of hemangiomas and one case of lymphangioma that involved the parotid gland. Histologically, the cellularity and increased division figures in these lesions should not be interpreted as a sign of a malignant condition. A watchful expectancy for spontaneous regression and preservation of the facial nerve at surgery are advocated.

Histological examination of paraformaldehyde-exposed gutta-percha implanted in rats.

Adsorption of some paraformaldehyde was noted in a previous study evaluating its sterilizing effect on gutta-percha (GP). This study examined histologically the effect of this adsorption when formaldehyde-exposed GP was implanted into the subcutaneous connective tissue of rats. GP implants were prepared in cylinder shape using a template designed to standardize size. Fifty GP cylinders were exposed to paraformaldehyde for 7 days before being implanted, while 50 others were implanted without exposure. Fifty rats had two implant sites prepared, at dorsal-interscapular and dorsal-caudal regions. Sham operations were performed on 10 rats to examine the effect of the surgery itself. The animals were killed at 4, 7, 14, 28, and 56 days. There was a significant difference between the two categories of implants only at 7 days, with the GP specimens without paraformaldehyde exposure showing more inflammation than GP with paraformaldehyde specimens (p = 0.043). Although the GP-alone specimens showed greater initial inflammation, both groups recovered in the same time period. One of the GP specimens with paraformaldehyde still showed a moderate/severe response at 56 days, whereas all of the GP-alone specimens showed only none/mild responses. The GP examined appeared to cause more inflammation than was expected.

Connective tissue reactions to glass ionomer cements and resin composites.

This study evaluated the histopathologic reactions of rat connective tissue to two glass-ionomer cements (Fuji Cap II, Fuji Ionomer Type III) and two microfilled light-cured composite resins (Helio-molar Radiopaque and Helioprogress). IRM (zinc oxide-eugenol cement) was used as a control. Discs of the materials, 5 mm in diameter and 2 mm thick, that had set for 15 minutes were implanted under the dorsal skin of 75 Sprague-Dawley rats. There were 15 rats in each group and each animal received two identical implants. Five rats from each group were terminated at 7, 28 and 85 days after implantation. Histologic sections of the implant sites were stained with hematoxylin and eosin. Findings at all study periods indicated that Fuji Ionomer Type III elicited more intense reactions than the other materials. Reactions to Fuji Cap II, Heliomolar Radiopaque and Helioprogress, at all study periods, were comparable to each other and to IRM.

X-linked (recessive) hypomaturation amelogenesis imperfecta: a prosthodontic, genetic, and histopathologic report.

A 16-year-old white girl requested esthetic restorations for her teeth. She was a manifesting heterozygote for the X-linked recessive form of amelogenesis imperfecta with hypomaturation defect. Lyonization theory states that in the somatic cells of female mammals, one of the two X chromosomes is randomly inactivated early in development. Therefore, females who are heterozygous for a given X-linked gene will be mosaic with varying proportions of cells in which only one of a particular pair of alleles is active. This mosaicism produced by lyonization ensures considerable phenotypic variability in the clinical expression of X-linked disorders. Histologic examination of the patient's extracted third molars demonstrated the expected lyonization effect. The patient received six porcelain laminate veneered crowns to restore her maxillary incisor and canine teeth. This article represents the first reported use of such restorations to alleviate the cosmetic and functional handicaps that accompany this genetic disease.

Human saliva coronal microleakage in obturated root canals: an in vitro study.

This study assessed salivary penetration through obturated root canals as related to time by using two methods of analysis--histological examination and dye penetrations. A total of 160 human maxillary anterior teeth were instrumented to size 60 Hedstrom file. Ten of the teeth were not obturated; 150 teeth were obturated by lateral condensation of gutta-percha and Roth's root canal sealer. Fifty of these teeth received intermediate restorative material temporaries to a thickness of approximately 3 mm. All teeth were immersed in 50 ml of whole human saliva and kept at 37 degrees C and 100% humidity. The saliva was changed daily. At 2, 7, 14, 28, and 90 days, 32 teeth were removed from the saliva. Of these, two were unobturated and were examined for bacterial penetration by culturing of the apical one-third. Ten teeth without temporaries were immersed in Pelikan ink for 2 days to demonstrate the extent of salivary penetration. These teeth were decalcified and cleared to allow direct measurement of dye penetration. Decalcified serial 7-microns-thick sections were prepared from the remaining 20 teeth, 10 with and 10 without intermediate restorative material temporaries. The sections were stained with hematoxylin and eosin stain and Brown and Hopps stain. Saliva penetration assessed in histological sections was significantly less than was visualized with dye analysis. Salivary penetration at 3 months was significantly greater than at the four earlier study periods. This amount of salivary penetration was considered to be clinically significant. The results strongly suggest retreatment of obturated root canals that have been exposed to the oral cavity for at least 3 months.