Characterization and establishment of an immortalized rabbit ovary granulosa cell line for reproductive biology experiments

Granulosa cells (GCs) are the key components of ovarian follicles for regulating oocyte maturation. communicating with oocytes through complex gap junctions and regulate the growth and development of oocytes. In addition to being considered to have great potential for human therapeutic model development and livestock breeding. Here, we transformed lentivirus-mediated simian virus 40 Large T (SV40LT) into primary rabbit granulosa cells (Pri RGCs) to establish an immortalized cell line. Morphologically, the immortalized RGCs (Im RGCs) were indistinguishable from the Pri RGCs, cell structure was intact following H&E staining. No significant difference in cell proliferation viability and growth between Im RGCs and Pri RGCs. Based on GCs-specific markers, the expression of FSHR , StAR , CYP11A1 and CYP17A1 were identified by PCR, immunofluorescence, and western blot analysis. ELISA shows that ImRGCs can synthesize steroid hormones normally. Karyotype analysis showed that the number of chromosomes remained normal during the process of infinite passage, soft-agar cloning experiment and nude mice tumorigenic experiment Indicates that the ImRGCs were not malignantly transformed. In conclusion, the immortalized granulosa cell line of rabbit follicle was established and can be used as a tool cell for future studies on biological research, animal husbandry and female reproductive disease.