氯丙嗪对大鼠离体培养卵巢颗粒细胞凋亡的影响

Effect of Chlorpromazine on Apoptosis of Rat Ovary Granulose Cells in Vitro

本试验旨在通过氯丙嗪对颗粒细胞凋亡影响的研究,探讨氯丙嗪对雌性大鼠性腺毒性的作用机制。对未成熟的Wistar大鼠卵巢颗粒细胞进行原代培养,用不同浓度的氯丙嗪(0、0.1、1、10μmol/L)染毒细胞,细胞培养24h。染毒结束后采用MTT法检测细胞相对活力,荧光染料Hoechest 33258检测颗粒细胞的凋亡变化,RT-PCR检测凋亡调控基因Bax、Bcl-2和P53mRNA的表达。在本试验所设置的剂量范围内,与对照组比较,氯丙嗪能极显著促进颗粒细胞凋亡(P<0.01),呈浓度依赖关系;RT-PCR检测则显示氯丙嗪能引起Bcl-2、Bax、P53mRNA表达水平和Bax mRNA/Bcl-2mRNA值升高,除低剂量组的Bax mRNA表达水平和Bax mRNA/Bcl-2mRNA值无明显改变外,其余各组与对照组相比均差异极显著(P<0.01)。氯丙嗪可显著抑制大鼠卵巢颗粒细胞活力,诱导颗粒细胞凋亡。

The aim is to investigate the relationship between female gonad toxicity of chlorpromazine on rat and the apoptosis of granulosa cells.Immature ovarian granulosa cell of Wistar rat was primary cultured,cells were cultured and treated with different concentration chlorpromazine（0,0.1,1,10 μmol/L） for 24 h.After exposured to chlorpromazine 24 h,cell viability was detected by MTT assay,apoptotic cells were identified microscopically by chromatin condensation and nuclear fragmentation with Hoechest 33258,mRNA expressions of Bax,Bcl-2 and P53 were determined by reverse transcriptase-polymerase Chain reaction（RT-PCR）.Compared to control group,chlorpromazine could inhibit the proliferation of ovarian granulosa cell（P0.01） in a concentration-dependent way.RT-PCR detection showed that chlorpromazine upregulated the mRNA expression of Bax,P53 and the mRNA ratio of Bax/Bcl-2.Compared with control group,the differences between relevant indicators was statistically significant（P0.01） except Bax and Bax/Bcl-2 of low group.Chlorpromazine may serve as an important endocrine disrupting medicine,mediating rat granulose cell apoptosis in vitro.The effect is bidirectional and concentration dependent.