3-硝基丙酸对鸡等级卵泡颗粒细胞孕酮分泌的影响

Effects of osthole on progesterone secretion in chicken preovulatory follicles granulosa cells

鸡等级卵泡颗粒细胞培养24 h后,在培养基中加入10、5、2.5 mmol/L的3-硝基丙酸(3-NPA)继续培养24 h。利用活性氧(ROS)试剂盒检测细胞中ROS的含量;采用ELISA检测细胞上清中孕酮(P4)含量,并采用qRT-PCR技术检测类固醇生成急性调节蛋白(StAR)、细胞色素P450胆固醇侧链裂解酶(P450scc)及3β-类固醇脱氢酶(3β-HSD)mRNA的相对表达量,以检测3-NPA对颗粒细胞孕酮分泌的影响。结果,加入3-NPA培养24 h后,10、5、2.5 mmol/L组细胞中ROS含量相对于细胞对照组均显著升高(P<0.05);各组细胞上清的孕酮分泌量并无显著差异,而10、5、2.5 mmol/L组细胞中StAR、P450scc、3β-HSD mRNA的相对表达量相对于细胞对照组均极显著降低(P<0.001)。上述结果说明,3-NPA会导致鸡等级卵泡颗粒细胞的ROS生成并能显著降低StAR、P450scc、3β-HSD mRNA的相对表达量。

After 24 h of culture of hen preovulatory follicles granulosa cells,10,5,and 2.5 mmol/L 3-nitropropionic acid(3-NPA)were added to the culture medium and then cultured for another 24 h.The effect of 3-NPA on the progesterone secretion of granulosa cells were assessed by reactive oxygen species(ROS)content detected by ROS kit,progesterone(P4)content in supernatant of cells by ELISA and the relative expression of steroidogenic acute regulatory protein(StAR),cytochrome P450 cholesterol side chain cleavage(P450scc)and 3-hydroxysteroid dehydrogenase(3β-HSD)gene mRNA by qRT-PCR.In result,after 24-hour culture with 3-NPA,the ROS content in the cells of the 10,5,and 2.5 mmol/L groups were significantly increased compared to the control group(P<0.05);the gene expression of StAR,P450scc,and 3β-HSD of the10,5,and 2.5 mmol/L groups were significantly decreased(P<0.001)than control group though there was no significant difference in the progesterone secretion.In conclusion,3-NPA could induce ROS production and significantly decrease the gene expression of StAR,P450scc,3β-HSD in hen preovulatory follicles granulosa cells.