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目的探讨环孢素A(cyclosporin A,CsA)作用于人滋养层细胞差异表达的功能基因。方法应用抑制性消减杂交筛选1.0μmol.L-1CsA作用于JAR细胞系前后差异表达的功能基因,经RT-PCR及蛋白质印迹进一步验证CsA作用前后原代分离的人早孕期滋养层细胞及JAR细胞株titin表达的变化。结果CsA作用于人JAR细胞系前后出现6个具有差异表达的基因,其中1个为已知基因titin,2个为功能未知的mRNA,3个为位于16号染色体上的EST。RT-PCR显示,CsA作用于人滋养层细胞24 h后出现titinmRNA的表达,72 h达高峰,并呈现明显的剂量依赖性,1.0μmol.L-1CsA作用最明显。蛋白质印迹显示,CsA可诱导titin的表达。结论CsA可能通过诱导滋养层细胞titin的高表达,改变其生物学行为,从而有利于胚胎着床及早期发育。
Objective To investigate the effect of cyclosporin A (CsA) on human trophoblast cells differentially expressed genes. Methods Inhibitory subtractive hybridization was used to screen the differentially expressed functional genes before and after Jurous cell line treated with 1.0μmol.L-1CsA. RT-PCR and Western blotting were used to further validate the effect of CsA on the early isolation of human trophoblast cells and JAR cells Variation of strain titin expression. Results Six genes with differential expression were found before and after CsA treatment on human JAR cell line. Among them, one was known as titin, two were unknown function mRNAs and three were ESTs located on chromosome 16. The results of RT-PCR showed that titin mRNA was expressed in human trophoblast cells for 24 h, and peaked at 72 h with a dose-dependent manner. The effect of 1.0 μmol·L-1 CsA was the most obvious. Western blotting showed that CsA induced titin expression. Conclusion CsA may induce the high expression of titin in trophoblast cells and change its biological behavior, which is beneficial to embryo implantation and early development.