hCG Improves Luteal Function and Promotes Progesterone Output through the Activation of JNK Pathway in the Luteal Granulosa Cells of the Stimulated IVF Cycles†

Abstract

Human chorionic gonadotropin (hCG) is a luteotropic hormone that promotes the survival and steroidogenic activity of corpus luteum (CL) by acting through luteinizing hormone receptors (LHRs) expressed on luteinized theca and granulosa cells (GCs). Therefore, it is used to support luteal phase in in vitro fertilization (IVF) cycles to improve clinical pregnancy rates and prevent miscarriage. However, the molecular mechanism underlying this action of hCG is not well characterized. To address this question, we designed an in vitro translational research study on the luteal GCs obtained from 58 IVF patients. hCG treatment at different concentrations and time points activated c-Jun N-terminal kinase (JNK) pathway and significantly increased its endogenous kinase activity along with upregulated expression of steroidogenic enzymes (steroidogenic acute regulatory protein (stAR), 3β-Hydroxysteroid dehydrogenase (3β-HSD)) in a dose-dependent manner in the luteal GCs. As a result, in vitro P production of the cells was significantly enhanced after hCG. When JNK pathway was inhibited pharmacologically or knocked-down with small interfering RNA luteal function was compromised, P4 production was declined along with the expression of stAR and 3β-HSD in the cells. Further, hCG treatment after JNK inhibition failed to correct the luteal defect and promote P4 output. Similar to hCG, luteinizing hormone (LH) treatment improved luteal function as well and this action of LH was associated with JNK activation in the luteal GCs. These findings could be important from the perspective of CL biology and luteal phase in human because we for the first time identify a critical role for JNK signaling pathway downstream LHR activation by hCG/LH in luteal GCs. Summary Sentence JNK signaling pathway plays a central role in the upregulated expression of the steroidogenic enzymes StAR and 3b-HSD and augmented progesterone production by hCG/LH in human luteal granulosa cells.

Terminal differentiation of human granulosa cells as luteinization is reversed by activin-A through silencing of Jnk pathway

“…We recently showed that hCG-induced improvement in luteal function is associated with activation of JNK pathway. When JNK pathway was inhibited pharmacologically or knocked-down with siRNA luteal function was compromised and P 4 production declined along with the expression of StAR and 3β-HSD in the cells 24 . It is worthwhile to note that the observed changes in the steroidogenic function of luteal GCs after JNK inhibition either pharmacologically or via siRNA are similar to those we observed after activin-A treatment.…”

“…Limited data is available on the biological role of this pathway in human ovary. We previously documented the critical roles of JNK pathway in the regulation of FSH induced follicle growth and cell cycle progression of granulosa cells in mouse model 29,30 , in the pathogenesis of granulosa cell tumor in human 31 , and very recently in the hCG-induced improvement of luteal function in human luteal GCs 24 . Albeit not human a study on rat ovary and granulosa cells demonstrated that the expression patterns of Jun and Fos family members in response to FSH and LH were distinct.…”

“…The samples size required for statistical significance and proper interpretation of the results was calculated according to the experimental methodologies used in the study as we previously described 24 . mRNA levels of the target genes used in the qRT-PCR assay (steroidogenic enzymes, LH and FSH receptor) and hormone levels are continuous variables therefore, expressed as the mean ± SD.…”

“…It was previously documented that human luteal granulosa cells (GCs) continue to have steroidogenic activity in vitro [16][17][18][19][20][21][22] . In agreement, we first confirm that these cells have a certain basal steroidogenic activity that produce detectable amounts of E 2 and P 4 even under serumfree culture condition.…”

“…Luteal GCs were isolated during oocyte retrieval procedure from follicular aspirates and cultured according to the procedures we previously described 16,24 . The cells were pooled for the experiment for most of the time only when the patients were comparable for age, infertility etiology, and stimulation protocol.…”