Molecular mechanism and functional role of macrophage colony‑stimulating factor in follicular granulosa cells

Our previous demonstrated that macrophage colony‑stimulating factor (M‑CSF) stimulated the production of estradiol (E2) and progesterone (P) in luteinized granulosa cells (GCs), and that its secretion may be regulated by follicle‑stimulating hormone (FSH). The present study aimed to examine the effect of M‑CSF alone or with Letrozole treatment on the function of non‑luteinizing granulosa cells, using the COV434 cell line, and its interaction with FSH. Human luteinized granulosa cells (LGC) were isolated from the follicular fluid of superovulated infertile patients (average age, 30.8±2.1 years) undergoing an intracytoplasmic sperm injection. The LGC were cultured with various concentrations of recombinant human macrophage colony stimulating factor (rhM‑CSF; 0, 10, 25, 50 or 100 ng/ml), rhM‑CSF+Letrozole (10‑6 mol/l), rhFSH (0, 10, 25, 50 or 100 IU/ml), or rhFSH+Letrozole (10‑6 mol/l). E2 concentrations in the media were measured using ELISA. The expression levels of the FSH receptor and the M‑CSF receptor were detected via reverse transcription‑quantitative polymerase chain reaction. Following COV434 cell treatment with M‑CSF, cell proliferation was quantified using the MTS assay and protein expression was detected by western blotting. It was demonstrated that M‑CSF and FSH stimulated the production of E2. The production of FSH receptors was enhanced by rhM‑CSF or rhM‑CSF+Letrozole in vitro in a dose‑dependent manner. It was observed that rhFSH promoted the expression of the M‑CSF receptor, at a certain concentration. Proliferation of COV434 cells increased in a dose‑dependent manner following treatment with RhM‑CSF. Furthermore, M‑CSF induced the phosphorylation of c‑Jun N‑terminal kinase (JNK) and p38; however, the level of E2 in the medium was not altered when the cells were pretreated with the JNK inhibitor SP600125 or the p38 inhibitor SB203580. The present study suggested that M‑CSF may be important in regulating the response of GCs to gonadotropin and may have a promotive effect in the early phase of follicular development. The biological effects of M‑CSF may partially be attributed to activation of the JNK and p38 signaling pathways. M‑CSF may represent a novel follicular development regulator agent in the future.