小鼠胎盘周细胞的原代培养及其在胎盘血管毒物筛查中的初步应用

目的:建立小鼠胎盘周细胞的分离、培养方法并应用于胎盘血管毒物筛查研究。方法:将C57小鼠雌雄合笼受孕后,在小鼠妊娠的第18.5天,取胎盘,通过机械分离和组织消化法分离、培养小鼠胎盘周细胞;采用免疫荧光法鉴定细胞来源与纯度;采用MTS比色法检测胎盘周细胞的生长曲线。将胎盘周细胞分别暴露于0、6.25、12.50、25.00、50.00和100.00 μmol/L的醋酸铅24 h后,用MTS法检测染毒后的细胞活力。结果:利用机械分离和组织消化法分离、培养后,小鼠胎盘周细胞可贴壁生长,呈多边形,表达特异性标志α-平滑肌肌动蛋白(α-SMA)。在暴露于6.25~100.00 μmol/L醋酸铅24 h后,随着染毒剂量的增加,细胞的存活率逐渐下降,且各个剂量组之间的差异具有统计学意义(P<0.05)。结论:采用机械分离和组织块消化法可以简单、快捷地获得高纯度的小鼠胎盘周细胞,并可以应用于铅等胎盘血管毒物的检测。

Usefulness of primary mouse placental pericyte culture to screen for placental vascular toxicants

OBJECTIVE:To establish the method of isolation and culture of mouse placental pericytes and to use them for screening of placental vascular toxicants. METHODS:Placental pericytes were isolated from day 18.5 pregnant mice by mechanical separation and tissue digestion, and cultured in vitro. Immunocytochemistry tests revealed the cell characteristics and purity of placental pericytes. Cell proliferation was measured by using the Pomega MTS Cell Proliferation Colorimetric Assay Kit. Pericytes were treated with 6.25-100.00 μmol/L lead acetate and cell toxicity of each group was determined by MTS assay. RESULTS:Mouse placenta pericytes were successfully isolated by mechanical and tissue digestion method and could grow well on culture plates. The pericytes exhibited typical polygonal morphology and positive staining for α-SMA. After 6.25-100.00 μmol/L lead acetate treatment for 24 h, a gradual decline in cell viability was observed, and the difference was statistically significant. CONCLUSION:High purity placental pericytes from mice can be successfully collected by mechanical separation and tissue digestion, and can be used to screen placental vascular toxicants.