IMG-A1: A Novel Immortalized Granulosa Cell Line for Investigating FSH-Dependent Folliculogenesis and Ovarian Pathophysiology

The study of ovarian biology is hampered by the lack of in vitro models that faithfully recapitulate the physiology of granulosa cells (GCs). Primary GCs have a limited lifespan, while most immortalized lines are tumor-derived and exhibit non-physiological hormonal responses. The purpose of this study was to develop and characterize a novel immortalized GC line with a stable, physiologically relevant phenotype. We immortalized primary murine GCs from early antral follicles using lentiviral vector to introduce human telomerase reverse transcriptase (hTERT) gene to create the IMG-A1 cell line. The line was extensively characterized using molecular (qRT-PCR, Western blot), cytogenetic (karyotyping), and functional (hormone stimulation, ELISA, proliferation assays) methods to assess its phenotype and responsiveness to gonadotropins and metabolic stressors. Exhibiting a non-transformed phenotype, IMG-A1 cells retain a stable karyotype and express the follicle-stimulating hormone receptor (FSHR) but not the luteinizing hormone/chorionic gonadotropin receptor (LHCGR). Accordingly, they respond to FSH by upregulating steroidogenic genes like aromatase (<i>Cyp19a1</i>) but are unresponsive to LH/hCG. Furthermore, the line exhibits physiologically relevant responses to hormonal stimulation, including a strong induction of aromatase by FSH and its synergistic upregulation in a hyperandrogenic and hyperinsulinemic milieu. The IMG-A1 cell line is a unique and robust model of early antral granulosa cells, offering a valuable new tool for studying FSH-dependent folliculogenesis, cellular aspects of ovarian pathophysiology, and drug discovery.