过表达Gpr3诱导猪颗粒细胞G_1阻滞及凋亡

G蛋白偶联受体3(Gpr3)属于G蛋白偶联受体视紫质家族成员.Gpr3通过激活Gs蛋白介导的下游信号通路,维持卵泡卵母细胞减数分裂的前期阻滞,但在卵泡颗粒细胞中的作用不清.为了明确Gpr3在猪卵泡颗粒细胞中的功能,构建了Gpr3基因的真核表达载体,利用过表达的方式激活其介导的信号通路,并利用MTT、流式细胞术和real-time PCR等方法检测了过表达Gpr3对猪卵泡颗粒细胞增殖及凋亡的影响.结果显示,过表达Gpr3后,猪颗粒细胞的增殖水平显著下调,G0/G1期细胞的百分比增加,S期细胞减少,Cyclin B1和CDK1 mRNA的表达量也显著降低;同时,显著增加了颗粒细胞的凋亡率,在抑制Bcl-2表达的同时,促进了Bax的表达.结果表明:过表达Gpr3在猪颗粒细胞中具有抑增殖促凋亡的作用,丰富了其在调节卵泡发育过程中的生物学功能.

Overexpression of Gpr3 Induces G1-Arrest and Apoptosis in Porcine Granulosa Cells

G protein-coupled receptor 3 (Gpr3) is a member of the G protein-coupled receptor rhodopsin family. Gpr3 is a critical factor for the maintenance of meiotic prophase stability in oocytes via G protein-mediated pathways. In this paper, a Gpr3 overexpression vector was constructed to investigate the roles of Gpr3 in porcine granulosa cells (GCs). The effects on cell proliferation, apoptosis and cell cycle progression were analyzed by MTT, flow cytometry. The expression levels of related genes were determined by real-time PCR. The results showed that Gpr3 overexpression significantly inhibited porcine GCs proliferation, where G0/G1-phase cells were increased and S-phase cells were decreased. The mRNA expression of Cyclin B1 and CDK1 were reduced. Induction of apoptosis was detected following Gpr3 overexpression, where Bcl-2 expression was reduced and Bax expression was increased. These data implied that Gpr3 overexpression inhibited proliferation and promoted apoptosis in porcine GCs.