N-acetyl cysteine protects TMJ chondrocytes from oxidative stress

Temporomandibular joint (TMJ) inflammation is closely associated with oxidative stress. This study tested the potential of N-acetyl cysteine (NAC), an anti-oxidant amino-acid derivative, in alleviating oxidative stress-related damage in TMJ chondrocytes. The inflammatory condition was simulated by the addition of hydrogen peroxide (H?O?) to TMJ-derived chondrocyte cultures. Exposure to H?O? decreased the cell population by half within 2 days as a result of induced apoptosis and reduced proliferation. Gene expression of aggrecan and collagen II, as well as glycosaminoglycan production, were reduced by more than 70%. These compromised chondrocyte viability and function were fully restored by the addition of NAC to the cultures. NAC reduced the H?O?-elevated intracellular reactive oxygen species to the normal level and increased cellular glutathione reserves. These results indicate that NAC restores oxidative stress-induced cell death and severe functional impairment in TMJ chondrocytes, and warrant in vivo testing to explore its therapeutic potential as an anti-inflammatory agent.     

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.     

Induced migration of dental pulp stem cells for in vivo pulp regeneration

Dental pulp has intrinsic capacity for self-repair. However, it is not clear whether dental pulp cells can be recruited endogenously for regenerating pulp tissues, including mineralizing into dentin. This work is based on a hypothesis that dental pulp stem/progenitor cells can be induced to migrate by chemotactic cytokines and act as endogenous cell sources for regeneration and mineralization. Dental stem cells (DSCs) were isolated from adult human tooth pulp and seeded on the surfaces of 3D collagen gel cylinders that were incubated in chemically defined media with stromal-derived factor-1α (SDF1), basic fibroblast growth factor (bFGF), or bone morphogenetic protein-7 (BMP7). Significantly more cells were recruited into collagen gel by SDF1 or bFGF than without cytokines in 7 days, whereas BMP7 had little effect on cell recruitment. BMP7, however, was highly effective, equally to dexamethasone, in orchestrating mineralization of cultured DSCs. Cell membrane receptors for SDF1, bFGF, and BMP7 were up-regulated in treated DSCs. Upon in vivo delivery, bFGF induced re-cellularization and re-vascularization in endodontically treated human teeth implanted into the dorsum of rats. Thus, endogenous dental pulp cells, including stem/progenitor cells, may be recruited and subsequently differentiated by chemotaxis of selective cytokines in the regeneration of dental pulp.     

Marginal bacterial leakage and pulp reactions in Class II composite resin restorations in vivo

The presence of stainable bacteria under restorations and pulp reactions in 36 teeth restored in vivo with a modified Class II composite resin restoration with two different dentine treatment techniques were studied on three separate follow-up occasions (1-3, 7-10 and 28-32 days). Half of the cavities showed stainable bacteria at the cavity margins and bottoms. Teeth restored with method A (Gluma/Occlusin) showed significantly fewer restorations with stainable bacteria then teeth restored with method B (Life/Occlusin) (p less than 0.05). Significantly more restorations with detectable bacteria were found after 28-32 days and restorative method B (p less than 0.05). There were no differences in occurrence and grade of pulp inflammation for the different dentine treatment techniques and time periods.     

Adhesive resin induces apoptosis and cell-cycle arrest of pulp cells

The application of an adhesive resin near or directly over the pulp was shown to induce pulp inflammation and lack of dentin regeneration. We hypothesize that the absence of dentin bridging is due to adhesive-resin-induced apoptosis of cells responsible for pulp healing and dentin regeneration. Mouse odontoblast-like cells (MDPC-23), undifferentiated pulp cells (OD-21), or macrophages (RAW 264.7) were exposed to SingleBond polymerized for 0-40 seconds. Annexin V and propidium iodide assays demonstrated that SingleBond induced apoptosis of MDPC-23, OD-21, and macrophages. The proportion of apoptotic cells was dependent on the degree of adhesive resin polymerization. Adhesive-resin-induced death of pulp cells was associated with activation of the pro-apoptotic cysteine protease Caspase-3. Interestingly, most cells exposed to adhesive resin that did not undergo apoptosis showed cell-cycle arrest. We conclude that an adhesive resin induces apoptosis and cell-cycle arrest of cells involved in the regeneration of the dentin-pulp complex in vitro.     

可乐丽菲露AP-X复合树脂对人牙髓细胞毒性的研究

目的 通过细胞毒性试验，探讨可乐丽菲露AP-X复合树脂及其SEBOND粘接系统在临床上用于活髓牙窝洞直接修复术的安全性。方法选用年轻健康人新鲜离体牙髓，以组织块培养法及酶消化法进行原代培养。同时制备各材料样品，浸入DMEM培养基制取材料浸渍液。分别将各材料的浸渍液与第五代人牙髓细胞共同培养，以MTT法评价材料的细胞毒性。结果可乐丽菲露AP-X复合树脂的细胞毒性小于双组份玻璃离子水门汀Vitremer及复合树脂Z100，其差别具有统计学意义，与复合体F2000的细胞毒性无统计学意义的差别。可乐丽菲露的细胞毒性小于复合体F2000和复合树脂Z100对应粘接剂，且其差别具有统计学意义（P〈0．05）。结论可乐丽菲露AP-X复合树脂及其粘接剂的细胞毒性小于临床上现在常用的其他牙色类材料。     

N-acetyl cysteine alleviates inflammatory reaction of oral epithelial cells to poly (methyl methacrylate) extract

Abstract

Objectives. The purpose of this in vitro study was to determine whether the cytotoxicity of self-curing polymethyl methacrylate (PMMA) dental resin to oral epithelial cells was eliminated by mixing the antioxidant amino acid derivative, N-acetyl cysteine (NAC) with the material. Materials and methods. Rat and human oral epithelial cells cultured on polystyrene were incubated in culture medium with or without extract from self-curing PMMA dental resin, with or without pre-mixing with NAC. On day 1, the cultures were evaluated for cellular damage, intracellular formaldehyde invasion, cellular redox status and pro-inflammatory cytokine production. Formaldehyde content and the amount of released NAC in the extract were evaluated. Results. Rat epithelial cells cultured with PMMA extract showed marked increases in lactate dehydrogenase (LDH) release, intracellular formaldehyde and lysosomal levels and reductions in attached cell number and the amount of E-cadherin compared with those in the culture without the extract; these adverse biological effects were alleviated or prevented by pre-mixing the resin with NAC. In human oral epithelial cells cultured with PMMA extract, the addition of NAC into the resin prevented the intracellular elevation of reactive oxygen species and the reduction in cellular glutathione levels. Human cell cultures with the extract produced higher levels of various pro-inflammatory cytokines than cultures without the extract; this was prevented by mixing the resin with NAC. The extract from PMMA pre-mixed with NAC contained a lower concentration of formaldehyde and a substantial amount of antioxidants. Conclusion. The cytotoxicity of self-curing PMMA dental resin to oral epithelial cells was eliminated by mixing the resin with NAC.     

Cytotoxicity of polymerized resin cements on human dental pulp cells in vitro

ObjectivesThis study was designed to evaluate the cytotoxicity of several resin-based cements (Panavia F, Super-Bond C&B, Chemiace II) after polymerization on cultured human dental pulp cells.MethodsAfter polymerization, specimens from three resin-based cements were eluted with fresh Dulbecco's modified Eagle's medium (DMEM) without serum for 72 h, at 37 °C, using 0.4 g of each substance per milliliter of fresh medium. Elutes obtained during this step were passed through a 0.22-μm filter and diluted with the culture medium by a ratio of 75%, 50%, 25% (v/v). Cytotoxicity of elutes were evaluated by the relative growth rates (RGR) of pulp cells with a modified 3-(4, 5-dimethyl-thiazol-2-yl)-2, 5-diphenyl-tetrazolium bromide (MTT) assay. The RGR of pulp cells were statistic analyzed by the one-way analysis of variance among the groups.ResultsThe RGR of cells exposed to 100% concentration of elution of Panavia F, Super-Bond C&B, and Chemiace II were 74.42%, 85.54%, and 82.39%, respectively. The RGR increased along with the elution of cements diluted. There was significant difference between the Panavia F group and Super-Bond C&B group (p < 0.01), but there was no significant difference in the cytotoxicity between Chemiace II and Super-Bond C&B.ConclusionsAfter polymerization, three resin-based cements (Panavia F, Super-Bond C&B, Chemiace II) induced slight cytotoxicity. The sensitivity of cytotoxicity to human pulp cells depended on the resin-based cements and the concentration of the elution. Super-Bond C&B is the least cytotoxic agent among the three resin-based cements.     

人牙髓细胞在离体牙髓腔内向成牙本质细胞样细胞分化的研究

目的观察人牙髓细胞在离体牙髓腔内向成牙本质细胞样细胞分化的潜能。方法将原代培养的人牙髓细胞接种至处理过的离体牙髓腔内,2周后固定、脱钙、包埋、切片、染色,显微镜下观察其生长及分化情况。结果牙髓细胞在牙本质表面生长良好,部分细胞伸出胞质突伸入牙本质小管中,表现出成牙本质细胞样细胞的形态。结论牙本质可以诱导牙髓细胞向成牙本质细胞样细胞分化,可为组织工程化牙髓的研制提供实验依据。     

复合树指单体对人牙髓细胞毒性的研究

目的：评价复合树脂单体对人牙髓的毒性作用,探讨不同细胞系对检测敏感性的影响。方法：选用人牙髓细胞（ＬＳＣ）为实验细胞,以Ｌ－９２９小鼠成纤维细胞作为对照细胞系,采用ＭＴＴ比色分析法,对两种牙科用复合树脂单体（ＴＥＧＤＭＡ和ＵＤＭＡ）进行体外细胞毒性研究。结果：在低于ＩＣ５０的各浓度组中,ＬＳＣ细胞的生存率高于Ｌ－９２９细胞;ＭＴＴ比色法作为一种能定量检测牙科材料细胞毒性的有效方法,实验时应注重考虑     

Response of pulp cells to resin infiltration of enamel white spot-like lesions

ObjectivesTo investigate the trans-enamel and trans-dentinal biological effects of treating enamel white spot-like lesions (EWSLs) with resin infiltration components (RICs) on odontoblast-like cells (MDPC-23) and human dental pulp cells (HDPCs).MethodsEWSLs were induced in 60 enamel/dentin discs (4.0 ± 0.2 mm thick) using S. mutans. The discs were adapted into artificial pulp chambers and MDPC-23 were seeded on the dentin surface. The components of a resin infiltration system (Icon) were applied individually or in combination on the enamel surface as following (n = 10/treatment): Etch, Infiltrant, Etch+Infiltrant, or Etch+Dry+Infiltrant. The application of water or hydrogen peroxide served as negative and positive controls, respectively. After 72 h, MDPC-23 viability was evaluated. The extracts were exposed for 72 h to pre-cultured MDPC-23 and HDPCs in 96-well plates to evaluate cell viability, alkaline phosphatase activity (ALP), mineralized nodule formation (MN), and the expression of inflammatory cytokines (ICs) and mineralization-related genes (MRs). Data were analyzed by ANOVA complemented with Tukey or Games-Howell post-hocs (α = 5%).ResultsCell viability, ALP activity, and MN formation were significantly reduced in response to the RICs, presenting intermediate values compared to positive and negative controls. Likewise, ICs were upregulated, whereas MRs were downregulated. Among the RICs, the Etch component caused the most notorious detrimental effects.SignificanceResin infiltration of EWSLs negatively affected the metabolism of pulp cells in vitro. Therefore, even though resin infiltration is a micro-invasive therapy for non-cavitated caries in enamel, it should be closely followed up seen that components may diffuse and unbalance pulp homeostasis.     

In vivo and in vitro glycosaminoglycans from human dental pulp.

A qualitative assessment was made of the type of glycosaminoglycans (GAG) present in normal human dental pulp using electrophoresis on cellulose-acetate plates. A comparison was also made between the GAG derived directly from the dental pulp (in vivo) and those derived from cultured pulp fibroblasts from the same individual (in vitro). The results of this study showed four main types of GAG in normal human dental pulp tissue, which were dermatan sulfate, heparan sulfate, hyaluronic acid, and chondroitin sulfate. GAG synthesis from cultured pulp fibroblasts in vitro was different from the GAG present in the dental pulp (in vivo). Extracellular GAG, as well as pericellular GAG consisted of dermatan sulfate, hyaluronic acid, chondroitin sulfate, and heparin. Cellular GAG, however, contained only dermatan sulfate, hyaluronic acid, and chondroitin sulfate. There was no difference in type of GAG from the second and fourth passaged pulp fibroblasts.     

Early in vivo and in vitro effects of amelogenin gene splice products on pulp cells

Recombinant amelogenin gene splice products A+4 and A-4, implanted in the pulp, induce the recruitment, proliferation, and differentiation of reparative cells. Our aim was to investigate the precocious events occurring in the pulp 1 d and 3 d after implantation of agarose beads alone or loaded with A+4 or A-4. Proliferation and cell recruitment towards an odonto/osteogenic phenotype were visualized by detection of the proliferation cell nuclear antigen (PCNA) and RP59. After implantation of beads alone or loaded with A+4, at day 3, pulp cells were moderately immunopositive for osteopontin (OP), whereas labeling was strongly positive upon treatment with A-4. Dentin sialoprotein (DSP) labeling was not detectable. Parallel in vitro studies were carried out on odontoblastic and mesenchymal progenitor cells in order to evaluate the effect of the amelogenin peptides on the expression of a series of marker genes involved in the odontoblastic/osteogenic/chondrogenic differentiation pathways. Altogether, our results suggest that the 'signaling' effects of the amelogenin peptides A+4 and A-4 may differ according to the type of target cells, their stage of differentiation, the time of treatment, and the type of amelogenin peptide (A+4 or A-4).     

Proliferative ability and alkaline phosphatase activity with in vivo cellular aging in human pulp cells

Little is known about the effect of aging on characteristic functions of pulp cells. When damaged pulp is recovered and mineralized tissue is formed to protect remaining pulp tissue, the general responses of pulp tissue after adequate stimuli (pulp cell proliferation and activation of alkaline phosphatase [ALPase]) are thought to be essential. In this study, we compared proliferative ability and ALPase activity between cultures of human pulp (HP) cells obtained from young and aged donors. The in vitro proliferative lifespan of HP cells from young donors was longer than HP cells from aged donors. Growth rates and ALPase activity of HP cells decreased with increasing donor age. These findings suggest that impaired repair of pulp and dentin in aged patients is partly due to a decrease in the proliferative ability and ALPase activity in aged pulp cells.     

Dental pulp stem cells for in vivo bone regeneration: A systematic review of literature

ObjectiveThis review of literature was aimed to assess in vivo experiments which have evaluated the efficacy of dental pulp stem cells (DPSCs) for bone regeneration.DesignAn electronic search of English-language papers was conducted on PubMed database. Studies that assessed the use of DPSCs in bone regeneration in vivo were included and experiments evaluating regeneration of hard tissues other than bone were excluded. The retrieved articles were thoroughly reviewed according to the source of stem cell, cell carrier, the in vivo experimental model, defect type, method of evaluating bone regeneration, and the obtained results. Further assessment of the results was conducted by classifying the studies based on the defect type.ResultsSeventeen papers formed the basis of this systematic review. Sixteen out of 17 experiments were performed on animal models with mouse and rat being the most frequently used animal models. Seven out of 17 animal studies, contained subcutaneous pockets on back of the animal for stem cell implantation. In only one study hard tissue formation was not observed. Other types of defects used in the retrieved studies, included cranial defects and mandibular bone defects, in all of which bone formation was reported.ConclusionWhen applied in actual bone defects, DPSCs were capable of regenerating bone. Nevertheless, a precise conclusion regarding the efficiency of DPSCs for bone regeneration is yet to be made, considering the limited number of the in vivo experiments and the heterogeneity within their methods.     

rhBMP-7对牙髓细胞影响的体内、外研究

目的:观察rhBMP-7对体外培养的人牙髓细胞生物学的影响,及其诱导大鼠牙髓组织异位成骨作用.方法:体外人牙髓细胞连续培养,MTT法、碱性磷酸酶(ATP)检测法比较不同浓度BMP-7对体外人牙髓细胞增殖及ALP活性的影响.将BMP-7处理过的大鼠牙髓组织移植到同种异体大鼠的肾被膜下和头部皮下,分别在术后10、20 d观察移植组织的钙化情况.结果:60μg/L的rhBMP-7可以显著地刺激体外培养的人牙髓细胞增殖,同时提高细胞ALP活性.将用此浓度处理过的大鼠牙髓组织移植后数天,移植组织会在其诱导下产生不同程度的钙化.结论:BMP-7可诱导大鼠牙髓组织异位成骨.