两种牙科材料对人胚胎干细胞来源成纤维细胞的细胞毒性研究

目的:观察口腔常用材料氢氧化钙(calcium hydroxide,CH)和复合树脂对人胚胎干细胞来源成纤维细胞(embryo body fibroblasts-H9,EBf-H9)、原代培养人牙髓细胞(human dental pulp cells,hDPCs)及小鼠成纤维细胞L929的细胞毒性,探讨使用EBf-H9评价牙科材料生物安全性的可行性。方法:诱导人胚胎干细胞(H9)分化为EBf-H9,原代培养hDPCs,并经免疫细胞化学染色鉴定。用细胞计试剂盒-8(cell counting kit-8,CCK-8)检测比较梯度浓度CH和复合树脂浸提液对EBf-H9、hDPCs和L929细胞的细胞毒性。结果:CH或复合树脂分别作用24 h和48 h(或72 h)之后,L929细胞的存活率显著低于EBf-H9和hDPCs(P<0.05),但EBf-H9和hDPCs的细胞存活率没有统计学差异(P>0.05)。结论:对于CH和复合树脂的毒性反应,L929细胞与hDPCs有较大差异,EBf-H9比L929细胞更接近hDPCs的反应,提示在牙科材料生物安全性评价中,人胚胎干细胞来源成纤维细胞具有替代现有细胞检测模型的潜力。

Cytotoxicity of two dental materials on fibroblasts derived from human embryonic stem cells

Objective: To investigate the cytotoxicity of calcium hydroxide(CH) and composite resin on fibroblasts derived from human embryo body fibroblasts-H9(EBf-H9),human dental pulp cells(hDPCs) and immortalized fibroblasts L929;and to evaluate the use of EBf-H9 as a cellular model for cytotoxicity screening of dental materials.Methods: The EBf-H9 cells were derived from human embryonic stem cells(H9) via outgrowth of embryonic body(EB);hDPCs were isolated from healthy dental pulp,and identified by immunochemical staining.Cell Counting Kit-8(CCK-8) assay was applied to analyze the cytotoxicity of CH and composite resin with serial concentrations on the 3 kinds of cells.Results: Following 24 h and 48 h(or 72 h) post-treatment of CH and composite resin,the viability of L929 cells was significantly lower than that of EBf-H9 and hDPCs(P<0.05),and there was no significant difference between the last two groups(P>0.05).Conclusion: Immortalized fibroblasts L929 cells exhibited different response to CH and composite resin compared with EBf-H9 and hDPCs,and the last two cell types were similar to each other.This study indicated that fibroblasts derived from human embryonic stem cells were a potential cellular model instead of traditional immortalized murine cell line for cytotoxicity screening assay.