不同浓度GnIH对鸭颗粒细胞周期和增殖的影响

为了研究不同浓度GnIH对鸭颗粒细胞周期、增殖及相关基因表达的影响。本研究分别用不同浓度GnIH（0、0.1、1、10和100 ng·mL^-1）处理体外培养的鸭颗粒细胞24 h（n=3），观察细胞的生长状态，通过流式细胞术和EdU方法检测细胞周期和细胞增殖，并用qRT-PCR方法检测增殖相关基因CDK6、CyclinD1、IGF-2、IGFBP-2、p27^kip1的表达。结果显示，各浓度GnIH处理组的细胞生长状态良好，形态正常，细胞轮廓清晰，组间死亡细胞数差异不显著（P>0.05）；在0.1和1 ng·mL^-1 GnIH处理组，细胞周期阻滞在G2期的比例显著上升（P<0.05）；随着GnIH处理浓度的增加，EdU阳性细胞数所占的百分比降低；在0.1和1 ng·mL^-1 GnIH处理组，颗粒细胞中CDK6、CyclinD1、IGF-2、IGFBP-2、p27^kip1基因的相对表达量均下降，在10和100 ng·mL^-1 GnIH处理组中这些基因的相对表达量则有所上升。研究表明，在体外培养的鸭颗粒细胞中，GnIH能使细胞周期阻滞在G2期，并降低EdU阳性细胞所占百分比和下调增殖相关基因的表达水平，从而抑制颗粒细胞增殖来影响动物繁殖性能。

Effects of Different Concentrations of GnIH on Cell Cycle and Proliferation of Duck Granulosa Cells

The study aimed to explore the effects of different concentrations of GnIH on cell cycle, proliferation and related gene expression of duck granulosa cells. Primary duck granulosa cells were cultured in vitro and treated with different concentrations of GnIH (0, 0.1, 1, 10 and 100 ng·mL^-1) for 24 h (n=3). The cell growth status was observed, and the cell cycle and proliferation were detected by flow cytometry and EdU method, and the expression of proliferation-related genes CDK6, CyclinD1, IGF-2, IGFBP-2, p27^kip1 were detected by qRT-PCR. The results showed that the growth status of the cells in the GnIH treatment group at each concentration was good, the morphology was normal, the cell outline was clear, and the number of dead cells were not significantly different between groups (P>0.05). In the 0.1 and 1 ng·mL^-1 GnIH treatment groups, the proportion of cells in the G2 phase increased significantly (P<0.05). With the increase of the concentration of GnIH, the percentage of EdU-positive cells were decreased. In the 0.1 and 1 ng·mL^-1 GnIH treatment groups, the relative expression of CDK6, CyclinD1, IGF-2, IGFBP-2, p27^kip1 genes were downregulated in granulosa cells. The relative expression of these genes were increased in the 10 and 100 ng·mL^-1 GnIH treatment groups. Studies have shown that in the duck granulosa cells cultured in vitro, GnIH could hinder the cell cycle in G2 phase, meanwhile the percentage of EdU-positive cells were decreased and the expre-ssion of proliferation-related genes were downregulated, which could inhibit the proliferation of granulosa cells to affect the reproductive performance of animals.