Avina Paranjpe

Efficacy of Different Irrigation and Activation Systems on the Penetration of Sodium Hypochlorite into Simulated Lateral Canals and up to Working Length: An In Vitro Study

INTRODUCTION The removal of vital and necrotic pulp tissue, microorganisms, and their toxins is essential for endodontic success. However, the complex anatomy of the root canal system has limited our ability to debride it completely. Hence the purpose of this study was to evaluate the effect of currently used irrigation and activation systems on the penetration of sodium hypochlorite into simulated lateral canals and to working length in a closed system. METHODS One hundred single-rooted teeth were used in this study. A total of 600 simulated lateral canals were created, 6 in each tooth, with 2 lateral canals at 2, 4.5, and 6 mm of working length. To resemble the clinical situation, a closed system was created by coating each root with soft modeling wax. Roots were then randomly assigned to 4 experimental groups: group 1 (n = 20), Endoactivator (sonic activation); group 2 (n = 20), passive ultrasonic (PUI) activation; group 3 (n = 20), F file; group 4 (n = 20), apical negative pressure (ANP) irrigation; and control group 5 (n = 20), positive pressure irrigation. The samples were evaluated by direct observation of the images recorded under the dental operating microscope. RESULTS The results demonstrated that the ANP irrigation group was superior at reaching working length, and PUI was the most effective at lateral canal penetration. CONCLUSIONS The ANP irrigation system demonstrated limited activation of the irrigant into lateral canals but reached the working length significantly more than the other groups tested. In contrast, PUI group demonstrated significantly more penetration of irrigant into lateral canals but not up to the working length.

Effects of mineral trioxide aggregate on human dental pulp cells after pulp-capping procedures.

INTRODUCTION Pulp-capping procedures are routinely performed. The control of infection and biocompatibility of the pulp-capping materials are important factors in determining the treatment outcome. Calcium hydroxide has been considered the gold standard for this procedure. However, previous reports have reported the causes of failures with the use of calcium hydroxide. Mineral trioxide aggregate (MTA) has proved to be effective in the process of pulp capping. METHODS Human dental pulp stromal cells (DPSCs) were cultured on gray MTA, and the levels of gene expression, secretion of vascular endothelial growth factor, and the surface morphology were analyzed. RESULTS MTA promoted cell survival and proliferation, which was significantly different from the controls in human DPSCs. MTA up-regulated the expression of transcription factors like Runx2 and genes like osteocalcin, alkaline phosphatase, and dentin sialoprotein, which are important odontoblastic genes, thereby showing the ability to promote differentiation of the pulpal cells into odontoblast-like cells, which, in turn, are responsible for dentin bridge formation. MTA approximately induced a 1.7-fold increase in the secretion of angiogenic factors like vascular endothelial growth factor, which is important in the process of tissue healing and regeneration. The differences between the control and the MTA groups were statistically significant. Scanning electron microscopic studies revealed the differences in the surface morphologies between the control and the MTA groups. CONCLUSION Overall, this study supports the biocompatible nature of MTA and the possible mechanism of dentin bridge formation along with tissue repair, all of which contribute to a successful treatment outcome.

Resin Monomer 2-Hydroxyethyl Methacrylate (HEMA) is a Potent Inducer of Apoptotic Cell Death in Human and Mouse Cells

Mechanisms by which the resin monomer 2-hydroxyethyl methacrylate (HEMA) induces hypersensitivity reactions in humans are not well-established, nor have the direct effects of HEMA on cell death been fully characterized. The objective of this study was to establish whether HEMA is capable of inducing apoptotic cell death, and whether differences exist in the levels of apoptotic death induced by HEMA in cells obtained from healthy individuals and from patients with established HEMA hypersensitivity. HEMA induced apoptotic death in Peripheral Blood Mononuclear Cells (PBMCs) obtained from both healthy and HEMA-sensitized patients and in the murine RAW cells in a dose-dependent manner. However, induction of cell death by HEMA was lower in PBMCs obtained from patients in comparison with healthy individuals. Studies reported in this paper demonstrate that HEMA induces apoptotic death, and that decreased susceptibility of lymphocytes to HEMA-mediated death might be an important mechanism for the generation and persistence of hypersensitivity reactions in patients.

Increased Lysis of Stem Cells but Not Their Differentiated Cells by Natural Killer Cells; De-Differentiation or Reprogramming Activates NK Cells

The aims of this study are to demonstrate the increased lysis of stem cells but not their differentiated counterparts by the NK cells and to determine whether disturbance in cell differentiation is a cause for increased sensitivity to NK cell mediated cytotoxicity. Increased cytotoxicity and augmented secretion of IFN-γ were both observed when PBMCs or NK cells were co-incubated with primary UCLA oral squamous carcinoma stem cells (UCLA-OSCSCs) when compared to differentiated UCLA oral squamous carcinoma cells (UCLA-OSCCs). In addition, human embryonic stem cells (hESCs) were also lysed greatly by the NK cells. Moreover, NK cells were found to lyse human Mesenchymal Stem Cells (hMSCs), human dental pulp stem cells (hDPSCs) and human induced pluripotent stem cells (hiPSCs) significantly more than their differentiated counterparts or parental lines from which they were derived. It was also found that inhibition of differentiation or reversion of cells to a less-differentiated phenotype by blocking NFκB or targeted knock down of COX2 in monocytes significantly augmented NK cell cytotoxicity and secretion of IFN-γ. Taken together, these results suggest that stem cells are significant targets of the NK cell cytotoxicity. However, to support differentiation of a subset of tumor or healthy untransformed primary stem cells, NK cells may be required to lyse a number of stem cells and/or those which are either defective or incapable of full differentiation in order to lose their cytotoxic function and gain the ability to secrete cytokines (split anergy). Therefore, patients with cancer may benefit from repeated allogeneic NK cell transplantation for specific elimination of cancer stem cells.

Efficacy of sonic and ultrasonic activation for removal of calcium hydroxide from mesial canals of mandibular molars: a microtomographic study.

BACKGROUND The purpose of this study was to use micro-computed tomography (micro-CT) scanning to evaluate the efficacy of sonic and passive ultrasonic irrigation (PUI) on calcium hydroxide (Ca[OH](2)) removal and to measure the volume and percentage of Ca(OH)(2) remaining in the root canal system. METHODOLOGY The root canals of 46 extracted human mandibular molar teeth were prepared with rotary instruments and randomly assigned to two experimental groups (n = 40) as well as positive and negative controls (n = 6). In each experimental group, 20 teeth were assigned to each irrigation protocol, sonic or passive ultrasonic irrigation. All experimental teeth and the positive controls were filled with Ca(OH)(2), whereas the negative control teeth did not receive Ca(OH)(2). All teeth were scanned using micro-CT scanning to determine the dressing volume. After 7 days, the Ca(OH)(2) was removed in the experimental groups using rotary instrumentation only, and the teeth were again scanned using micro-CT scanning to calculate volume and percentage of Ca(OH)(2) removed. Positive control teeth were not subjected to rotary instrumentation. Experimental samples were then irrigated using either sonic or passive ultrasonic and the volume of remaining Ca(OH)(2) was calculated using micro-CT. RESULTS Remnants of Ca(OH)(2) were found in all experimental groups. No Ca(OH)(2) was found in the negative controls, whereas a mean of 8.7 mm(3) of Ca(OH)(2) was recorded in the positive controls. Rotary plus passive ultrasonic irrigation removed significantly more Ca(OH)(2) (85.7%) than rotary plus sonic irrigation (71.5%) (p < 0.001). CONCLUSIONS The combination of rotary instrumentation and passive ultrasonic activation for 3 periods of 20 seconds results in significantly lower amounts of Ca(OH)(2) remnants in the canal compared with sonic irrigation.

Direct contact with Mineral Trioxide Aggregate activates and differentiates Human dental pulp cells

INTRODUCTION Mineral trioxide aggregate (MTA) is routinely used for pulp-capping procedures. However, little is known about its direct interaction with the cells or whether MTA is capable of releasing soluble factors that could help in differentiating cells. There have been no previous studies showing this aspect of MTA. Hence, the aim of this study was to determine whether direct contact of the cells with MTA was necessary to help differentiate the pulp cells into odontoblast-like cells. METHODS Human dental pulp cells were cultured on gray MTA, either in direct contact or away from the cells on a cell culture insert, and the levels of gene expression, secretion of vascular endothelial growth factor (VEGF), and the rates of cell proliferation were analyzed. RESULTS MTA when placed in direct contact with the cells promoted up-regulated the expression of important odontoblastic genes like osteocalcin and dentin sialoprotein, thereby showing that direct contact of the cells with the MTA is necessary to promote differentiation of the pulp cells into odontoblast-like cells, which in turn are responsible for dentin bridge formation. MTA also induced an increase in the secretion of VEGF when placed in direct contact with the cells. CONCLUSIONS Overall, our study supports the fact that direct contact of the cells with the MTA is necessary to help differentiate them into odontoblast-like cells, which in turn will lead to a successful treatment outcome.

N-acetyl Cysteine (NAC)-assisted Detoxification of PMMA Resin:

Despite its proven cytotoxicity, poly-methyl methacrylate (PMMA) resin is one of the most frequently and extensively used materials in dental practice. This study hypothesized that an anti-oxidant amino acid, N-acetyl cysteine (NAC), has the potential to detoxify this material. Ten percent of the rat dental pulp cells were viable when cultured on the PMMA resin for 24 hours, while over 70% of the cells were viable on the NAC-added resin. Nearly all suppressed alkaline phosphatase activity, matrix mineralizing capability, and odontoblastic gene expression, such as dentin sialoprotein, on the untreated control resin was recovered by NAC in a concentration-dependent manner. A Ca/P ratio of 1.65 was found in the extracellular matrix of cultures on NAC-added resin, while that in the untreated resin culture was 0.70. The addition of NAC to PMMA resin significantly ameliorated its cytotoxicity to the dental pulp cells and restored their odontoblast-like cell phenotype to a biologically significant degree.

Strategies to Rescue Mesenchymal Stem Cells (MSCs) and Dental Pulp Stem Cells (DPSCs) from NK Cell Mediated Cytotoxicity

Background The aim of this paper is to study the function of allogeneic and autologous NK cells against Dental Pulp Stem Cells (DPSCs) and Mesenchymal Stem Cells (MSCs) and to determine the function of NK cells in a three way interaction with monocytes and stem cells. Methodology/Principal Findings We demonstrate here that freshly isolated untreated or IL-2 treated NK cells are potent inducers of cell death in DPSCs and MSCs, and that anti-CD16 antibody which induces functional split anergy and apoptosis in NK cells inhibits NK cell mediated lysis of DPSCs and MSCs. Monocytes co-cultured with either DPSCs or MSCs decrease lysis of stem cells by untreated or IL-2 treated NK cells. Monocytes also prevent NK cell apoptosis thereby raising the overall survival and function of NK cells, DPSCs or MSCs. Both total population of monocytes and those depleted of CD16+ subsets were able to prevent NK cell mediated lysis of MSCs and DPSCs, and to trigger an increased secretion of IFN-γ by IL-2 treated NK cells. Protection of stem cells from NK cell mediated lysis was also seen when monocytes were sorted out from stem cells before they were added to NK cells. However, this effect was not specific to monocytes since the addition of T and B cells to stem cells also protected stem cells from NK cell mediated lysis. NK cells were found to lyse monocytes, as well as T and B cells. Conclusion/Significance By increasing the release of IFN-γ and decreasing the cytotoxic function of NK cells monocytes are able to shield stem cells from killing by the NK cells, resulting in an increased protection and differentiation of stem cells. More importantly studies reported in this paper indicate that anti-CD16 antibody can be used to prevent NK cell induced rejection of stem cells.

Fusobacterium nucleatum Outer Membrane Proteins Fap2 and RadD Induce Cell Death in Human Lymphocytes

ABSTRACT Bacterially induced cell death in human lymphocytes is an important virulence factor for pathogenic bacteria. Previously discovered mechanisms of bacterially induced cell death are predominantly based on the transfer of bacterial proteins to the target host cell, such as the toxins secreted through type I, II, and VI secretion systems or effector proteins injected through type III, IV, and Vb secretion systems. Here, we report a mechanism employed by the Gram-negative oral pathogen Fusobacterium nucleatum for cell death induction of human lymphocytes via two outer membrane proteins (OMPs), Fap2 and RadD, which share regions homologous to autotransporter secretion systems (type Va secretion systems). Genetic and physiological studies established that inactivation of the two OMPs led to significantly reduced ability to trigger cell death in Jurkat cells, while the corresponding double mutant was almost completely attenuated. Additional biochemical and molecular analyses demonstrated that cell-free F. nucleatum membranes are sufficient to induce cell death in Jurkat cells, suggesting that no active process or effector protein transfer was necessary to induce eukaryotic cell death.

Restored viability and function of dental pulp cells on poly-methylmethacrylate (PMMA)-based dental resin supplemented with N-acetyl cysteine (NAC)

This study examines cytotoxicity of poly-methylmethacrylate (PMMA)-based dental temporary filling resin to dental pulp cells, and the potential amelioration of the toxicity with an anti-oxidant amino-acid, N-acetyl cysteine (NAC). Dental pulp cells extracted from rat maxillary incisors were cultured on the resin material with or without NAC incorporation, or on the polystyrene. The cultures were supplied with osteoblastic media, containing dexamethasone. Forty five percent of cells on the PMMA dental resin were necrotic at 24h after seeding. However, this percentage was reduced to 27% by incorporating NAC in the resin, which was the level equivalent to that in the culture on polystyrene. The culture on the untreated resin was found to be negative for alkaline phosphate (ALP) activity at days 5 and 10 or von Kossa mineralized nodule formation at day 20. In contrast, some areas of the cultures on NAC-incorporated resin substrates were ALP and von Kossa positive. Collagen I and dentin sialoprotein genes were barely expressed in day 7 culture on the untreated resin. However, those genes were expressed in the culture on the resin with NAC. These results suggest that the decreased cell viability and the nearly completely suppressed odontoblast-like cell phenotype of dental pulp cells cultured on PMMA dental resin can be salvaged to a biologically significant degree by the incorporation of NAC in the resin.