Alteration of biological properties of bacterial lipopolysaccharide by calcium hydroxide treatment.

Bacterial lipopolysaccharide (LPS) plays a major role in the development of periapical bone resorption. Although the chemical properties of LPS are altered by treatment with an alkali such as calcium hydroxide, the effects of calcium hydroxide on the biological properties of LPS are not known. The purpose of this study was to investigate whether treatment of LPS with calcium hydroxide alters its biological action as measured by human monocyte secretion of prostaglandin E2. Monocyte cell cultures were stimulated with LPS or calcium hydroxide-treated LPS and culture supernatants were analyzed for prostaglandin E2 content using gas chromatography-mass spectrometry. Prostaglandin E2 was identified in supernatants of LPS-stimulated monocytes but not in those stimulated with calcium hydroxide-treated LPS. It was concluded that the treatment with calcium hydroxide may alter biological properties of bacterial LPS.

Effects of a bacterial cell wall fragment on monocyte inflammatory function.

Gram-negative bacteria recovered from necrotic pulps of teeth with periapical lesions have been shown to promote bone resorption through the effects of their lipopolysaccharide (LPS). Recently, it was shown that microflora of root-filled teeth with persisting periapical lesions consisted mainly of single species of Gram-positive bacteria. But Gram-positive bacteria do not contain LPS and their role in the development of periapical lesions is not clearly understood. The purpose of this study was to evaluate the effects of muramyl dipeptide (MDP), a cell wall component of both Gram-negative and Gram-positive bacteria, on cytokine release from monocytes. Human monocyte cultures were treated with MDP or LPS and interleukin-1 beta, and tumor necrosis factor-alpha levels in the supernatants were estimated. MDP and LPS stimulated cytokine release, but the effect of MDP was significantly less than that of LPS.

Tumor necrosis factor identified in periapical tissue exudates of teeth with apical periodontitis.

Root canal samples, taken from periapical tissue exudates during routine root canal treatment procedures, were processed for identification of tumor necrosis factor using a mouse anti-human monoclonal antibody and enzyme-linked immunosorbent assay. Detectable levels of tumor necrosis factor were identified in periapical tissue exudates in chronic apical periodontitis.

Effect of calcium hydroxide on bacterial lipopolysaccharide.

Apical periodontitis and its concomitant periapical osteolysis is caused by pulpal infection, and bacterial lipopolysaccharide (LPS) is known to play a major role in the bone resorption process. Little is known concerning the effect of root canal intervisit dressings on residual LPS in root canals after bacterial cell lysis. The purpose of this study was to evaluate the effects of calcium hydroxide on bacterial LPS. Free hydroxy fatty acids were quantified in samples of LPS treated with calcium hydroxide. Calcium hydroxide treatment of LPS was shown to release elevated quantities of hydroxy fatty acids. It was concluded that calcium hydroxide hydrolyzed the lipid moiety of bacterial LPS, resulting in the release of free hydroxy fatty acids. This result suggests that calcium hydroxide-mediated degradation of LPS may be an important reason for the beneficial effects obtained with calcium hydroxide use in clinical endodontics.

Root canal dentinal tubule disinfection.

Dentinal tubules of the root canal walls of human teeth were infected in vitro with a known bacterial isolate. The roots were exposed to either calcium hydroxide or iodine potassium-iodide for various periods of time and the viability of microorganisms was determined by incubation of entire root samples in a culture medium. The effects of the two agents on microbial viability were evaluated and compared. Iodine potassium-iodide disinfected dentin effectively. In contrast, bacteria remained viable in the dentin after relatively extended periods of calcium hydroxide treatment.

Cytotoxic evaluation of root-end filling materials in cultures of human osteoblast-like cells and periodontal ligament cells.

The cytotoxicity of three root-end filling materials (amalgam, IRM, and Super-EBA) was evaluated in cultures of human periodontal ligament cells and human osteoblast-like cells. Ten-millimeter-long plastic test tubes were filled with 3 mm of freshly mixed root-end filling materials at one end (1.5 mm diameter). The opposite end was sealed and attached by heat to a 35-mm cell culture dish. Human periodontal ligament cells and human osteoblast-like cells were seeded in the dishes. The size of cell-free zones around the root-end filling materials and the total cell number per dish were calculated after 3 and 7 days. Empty test tubes used as controls did not influence the growth and distribution of the cultured cells. Cell density increased in all groups in the test period. Amalgam had a larger cell-free zone, compared with IRM and Super-EBA and showed a reduction in total cell number per dish for both tested cell types. IRM and Super-EBA also had a cell-free inhibition zone for both cell types, but no significant reduction in total cell number per dish. This study showed that amalgam had a higher cell toxicity to human periodontal ligament cells and human osteoblast-like cells than IRM and Super-EBA.

Integrin expression in human dental pulp cells and their role in cell attachment on extracellular matrix proteins.

Integrins are a family of heterodimeric glycoproteins consisting of alpha and beta subunits that noncovalently interact to form cell surface adhesion receptors. The objective of this study was to identify integrins in human dental pulp cells and determine their role in human dental pulp cell attachment to the biological active molecules, laminin and fibronectin. Integrin expression was studied by immunoblot and immunoprecipitation using monoclonal integrin antibodies. The role of integrin in human dental pulp cell adhesion on laminin and fibronectin was determined by inhibition of cell adhesion with those antibodies. This study found human dental pulp cells expressed alpha 1, alpha 3, alpha 5, alpha 6, alpha v, and beta 1 integrin subunits. The adhesion of human dental pulp cells to laminin and fibronectin was not inhibited by monoclonal antibody to any subunit, except that anti-beta 1 antibody inhibited pulp cells adhesion on laminin. These data provide information for further studying the role of integrins in dental pulp cell biological function.

An in vitro method for longitudinal evaluation of toxicity of endodontic sealers.

An in vitro method for longitudinal evaluation of root canal sealers was developed and applied. A newly introduced cell culture chamber was used to evaluate the cytotoxic effects of test samples immediately after mixing and for an extended period of time thereafter. A ranking of the test materials, based on their cytotoxicity, was allowed by the method.

Adhesion of human osteoblasts on root-end filling materials.

Adhesion of human osteoblasts to root-end filling materials (mineral trioxide aggregate (MTA), IRM, composite, and amalgam) was observed by scanning electron microscopy. Root-end filling materials were inserted into 96-well flat-bottomed plates and condensed to disks of approximately 1 mm thick and the same diameter as the wells. After the disks were set, they were placed in the bottom of Nunc four-well culture plates at one disk per well. Then human osteoblasts were seeded into the wells at 1.5 x 10(5) cells per well. After 1 day in culture the disks of root-end filling materials along with cells grown on their surface were examined with a scanning electron microscopy. Results showed that osteoblasts attached and spread on MTA and composite by forming a monolayer. Osteoblasts also attached on amalgam, but with few cells spreading. In the presence of IRM, osteoblasts appeared rounded with no spreading. These results indicate that osteoblasts have a favorable response to MTA and composite resin compared with IRM and amalgam.

Enamel matrix derivative prolongs primary osteoblast growth.

Enamel matrix derivative (EMD) secreted by cells of the epithelial root sheath plays an important role in cementogenesis and periodontal tissue formation. The mechanisms by which EMD influences cell function are not known. The purpose of this study was to determine the effect of EMD on cell growth of primary mouse osteoblasts. Osteoblasts were digested from 6- to 8-day-old mouse calvaria and plated into 6-well cell culture plates at an initial density of 5000 cells/cm2. After 24-h incubation with Dulbecco's modified eagle medium (DMEM) supplemented with 10% fetal bovine serum, cells were incubated in three different groups of media: DMEM only as control, DMEM with 25 microg/ml EMD, and DMEM with 100 microg/ml EMD. At days 3, 7, 10, and 14, the total cell number per well was calculated, and cell morphology was examined. At each observation period the number of cells in the EMD groups was significantly greater (ANOVA, p < 0.01) than that in the control group. EMD had a greater effect on osteoblast survivor in the higher concentration than in the lower concentration. Furthermore normal morphology of the primary osteoblasts was maintained in the EMD groups. These results suggest that EMD prolongs primary osteoblast growth and may have an effect on osteoblasts during periodontal regeneration.

Antifungal effects of sodium hypochlorite and chlorhexidine in root canals.

The purpose of this study was to evaluate the antifungal properties of 0.12% chlorhexidine, 1% NaOCl, and 5% NaOCl. Root sections were enlarged and the smear layer was removed in half of the specimens. The specimens were fixed in the wells of tissue culture plates. Each root canal was dispensed with an inoculum of Candida albicans. After 10 days, the root sections were treated with 3 ml of either disinfectant solution for 1 min, 5 min, 30 min, and 1 h. Then, root sections were incubated in test tubes having Sabouraud's Dextrose Broth at 37 degrees C for 24 h. In the presence of the smear layer, antifungal activity was observed only in 1-h treatment groups for all solutions. However, in the absence of the smear layer, 5% NaOCl alone started to show antifungal activity after 30 min. The antimicrobial effectiveness of irrigating solutions should be re-evaluated, particularly in patients predisposed to oral candidiasis.

Colonization of Candida albicans on cleaned human dental hard tissues.

Candida albicans is a fungus that commonly infects oral mucosal surfaces. Limited data exist on biofilm formation by C. albicans on dental surfaces. Human premolar teeth were infected with C. albicans for 10 days and hard-tissue surfaces were examined with a scanning electron microscope. Enamel, cementum and dentine, in the absence or presence of a smear layer, were readily colonized by this micro-organism. Hyphae penetrated into cracks, followed the ridges of the cavities and migrated into dentinal tubules. Blastospores and hyphae were embedded in an extracellular material. These findings suggest that dental hard tissues may be invaded by C. albicans and thus can potentially present a reservoir for disseminating candidal infections.

The role of integrin beta 1 in human dental pulp cell adhesion on laminin and fibronectin.

OBJECTIVE: This study is to identify the expression of integrin beta 1 in human dental pulp cells and the role of integrin beta 1 in pulp cell adhesion on extracellular matrix protein laminin and fibronectin. STUDY DESIGN: Immunoblot detection of integrin beta 1 in human pulp cells was with the use of monoclonal anti-beta 1 antibody. Dental pulp cell adhesion assay on extracellular matrix protein laminin and fibronectin and blocking cell adhesion was performed with monoclonal anti-beta 1 antibody. RESULT: Integrin beta 1 was identified in human dental pulp cells. Pulp cells adhered and spread on both laminin and fibronectin. Monoclonal anti-beta 1 antibody inhibited human dental pulp cells adhesion on laminin but not on fibronectin. CONCLUSIONS: Integrin beta 1 was expressed on human dental pulp cells and mediated cell adhesion on laminin. Human dental pulp cells also adhered on fibronectin but the adhesion was not regulated by beta 1 integrin.

Growth patterns of Candida albicans in relation to radicular dentin.

UNLABELLED: Candida albicans is the most common fungal pathogen isolated from the oral cavity. The role of this organism as an endodontic pathogen is poorly understood. OBJECTIVES: The aim of this study was to observe the interaction of C. albicans with root canal walls and the growth patterns of this microorganism in relation to radicular dentin. STUDY DESIGN: Fifteen root sections were infected with C. albicans grown in calf serum and incubated for various periods. The sections were fixed in glutaraldehyde, split into two halves, and evaluated by scanning electron microscopy. RESULTS: Blastospores and hyphal structures were observed on the root canal walls of all specimens. Filamentous hyphal form was dominant in 5-day specimens. Most of the hyphae and blastospores showed penetration into dentinal tubules. The body of germinating mother cells and hyphae demonstrated collapsed cell walls as a result of vacuole formation. CONCLUSIONS: With this invasive affinity to dentinal structures, C. albicans may be considered a dentinophilic microorganism.

Effects of enamel matrix derivative on gene expression of primary osteoblasts.

OBJECTIVE: The aim of this study was to investigate the effects of enamel matrix derivative (EMD) on gene expression of collagen alpha1 (I), osteocalcin, prostaglandin G/H synthase 2 (PGHS-2), interleukin-6, and insulin-like growth factor I in primary mouse osteoblasts. STUDY DESIGN: Primary osteoblasts were digested from 6- to 8-day-old mouse calvaria. Cells were divided into 4 groups and cultured for 24, 48, and 72 hours with a serum-free modified Eagle medium as negative control, modified Eagle medium with 25 microg/mL EMD, modified Eagle medium with 100 microg/mL EMD, and modified Eagle medium plus 10% fetal bovine serum as positive control. Gene expression was determined by Northern blot and reverse transcription polymerase chain reaction analysis. RESULT: EMD enhanced collagen I, interleukin-6, and PGHS-2 expression and did not stimulate the expression of osteocalcin and IGF-I. CONCLUSION: These results indicate that EMD might regulate certain gene expression during periodontal tissue regeneration.

Endodontic considerations in restoration of partial overdenture abutments.

Overdenture abutment teeth often require endodontic treatment. Various factors, such as status of the pulp, periodontal state of the tooth, and the sequence of overdenture treatment, influence endodontic management of the patient. After completion of endodontic treatment, the coronal part of the root filling is removed, leaving an adequate amount of the root filling in the apical part of the root canal undisturbed. The abutment tooth is then permanently restored with a filling material, or is prepared for a cast restoration.