Effects of overexpressing <i>SAV1</i> on expression of <i>FSHR</i>, <i>StAR</i>, <i>CCND2</i> and <i>GDF9</i> mRNA examined and cell proliferation.

<p>Granulosa cells were transfected with reconstructed pFLAG-SAV1 plasmids, pFLAG CMV-2 empty vector (negative control) and no plasmid (blank control). (A) The expression of <i>SAV1</i> gene before and after the GCs transfected with pFLAG-SAV1expression vector for 24 h was examined by qRT-PCR. Quantification of mRNA expression as normalized to the <i>18S rRNA</i> gene, the values on the bar graphs are the mean ± SEM of 10 hens (n = 10) from a representative experiment. (B) Expression levels of SAV1 protein in the GCs before and after the transfection with with pFLAG-SAV1 vector was detected by western blotting. The β-actin was used as the loading control. (C) The influence of <i>SAV1</i> overexpression on <i>FSHR</i>, <i>StAR</i>, <i>CCND2</i> and <i>GDF9</i> mRNA abundances in the granulosa cells from prehierarchichal follicles (6 to 8 mm in diameter) was examined. (D) The effects of SAV1 overexpression on the GC proliferation were detected by BrdU incorporation assay (original magnification × 200). Of which panel a, blank control; panel b, negative control; panel c-d, RNAi group. (E) The number of BrdU⁺ cells was expressed as percentage and calculated relative to the total number of cells counted in the microscope fields observed in the negative control. For each group, bars with superscript symbol indicates that difference was significant compared to the control group ** P<0.01,* P<0.05.