早孕绒毛外细胞滋养细胞的体外培养方法

目的:探讨改良人早孕绒毛外细胞滋养细胞原代分离培养方法。方法:建立改良的复合酶消化梯度离心法,分离培养绒毛外细胞滋养细胞,通过免疫细胞化学、Giemsa染色和细胞侵袭实验检测细胞纯度、数量及生物学活性(侵袭性),并与低浓度胰蛋白酶消化方法进行比较。结果:低浓度胰蛋白酶消化方法和改良的复合酶消化梯度离心法所得贴壁细胞量分别为(11.69±1.26)×104和(17.21±0.51)×104,两者比较有统计学差异(P<0.05);绒毛外细胞滋养细胞纯度分别为(63.20±7.12)%和(92.10±5.11)%,两者比较有统计学差异(P<0.05),穿膜细胞数分别为(36.10±3.28)和(66.20±5.17),两者比较有统计学差异(P<0.05),提示该实验室改良的复合酶消化梯度离心法所得细胞总量、绒毛外细胞滋养细胞纯度及数量均显著高于低浓度胰蛋白酶消化方法。结论:该实验室改良的复合酶消化梯度离心法获得绒毛外细胞滋养细胞纯度和数量显著提高。

The method for culture in vitro of extravillous cytotrophoblast cells in early pregnancy

Objective:To explore an improved method for isolation and cultivation of human extravillous cytotrophoblast cells in early pregnancy.Methods:An improved compound enzyme digestion gradient centrifugation method was established,extravillous cytotrophoblast cells were isolated and cultured;the cell purify,amount and biological activity(invasiveness)were detected by immunocytochemistry,Giemsa staining and cell invasion test,then the results were compared with low dose trypsinase digestion method.Results:The amounts of adherent cells derived from low dose trypsinase digestion method and improved compound enzyme digestion gradient centrifugation method were(11.69±1.26)×104 and(17.21±0.51)×104,respectively,there was significant difference between the two groups(P<0.05);the purities of extravillous cytotrophoblast cells were(63.20±7.12)% and(92.10±5.11)%,respectively,there was significant difference between the two groups(P<0.05);the amounts of permeable cells were(36.10±3.28)and(66.20±5.17),respectively,there was significant difference between the two groups(P<0.05),the results indicated that the amount of obtained cells,purify and amount of extravillous cytotrophoblast cells derived from improved compound enzyme digestion gradient centrifugation method were significantly higher than those derived from low dose trypsinase digestion method.Conclusion:The purify and amount of extravillous cytotrophoblast cells derived from improved compound enzyme digestion gradient centrifugation method increase significantly.