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Inductions of granulosa cell luteinization and cumulus expansion are dependent on the fibronectin-integrin pathway during ovulation process in mice.

ABSTRACT: It has been known that EGF-like factor secreted from LH-stimulated granuloma cells acts on granulosa cells and cumulus cells to induce ovulation process. Granulosa cells are changed the morphology with differentiating cell functions to produce progesterone. Cumulus cells are detached to make a space between the cells to accumulate hyaluronan rich matrix. LH also changes extracellular matrix (ECM) components including fibronectin in the follicular walls and granulosa cell layers. EGF like factor and fibronectin synergistically play important roles in numerous cell functions, especially cancer cell migration, estimating that fibronectin would impact on granulosa cells and cumulus cells. To clear this hypothesis, the localizations of fibronectin and its receptor integrin were observed by immunofluorescence technique. The functions were monitored by the detection of downstream signaling pathway, focal adhesion kinase (FAK). The pharmacological approach in both in vivo and in vitro were used for analyzing the physiological roles of FAK during ovulation process. The immunofluorescence staining revealed that fibronectin and integrin were observed in granulosa cells, cumulus cells and the space between cumulus cells and oocyte at 4 and 8 h after hCG injection. Concomitantly with the changes of fibronectin-integrin localization, FAK was phosphorylated in periovulatory follicles. The injection of FAK inhibitor suppressed not only ovulation but also luteinization of granulosa cells and cumulus expansion. In cultured-granulosa cells, fibronectin-integrin synergistically activated FAK with amphiregulin (AREG). Such cooperative stimulations induced a morphological change in granulosa cells, which resulted in the maximum level of progesterone production via the induction of Hsd3b. When cumulus-oocyte complexes (COCs) were cultured with AREG in the presence of serum, the maximum level of cumulus expansion was observed. The AREG-induced cumulus expansion was also suppressed by FAK inhibitor. Thus, it is concluded that fibronectin and AREG synergistically activate FAK not only in granulosa cells and cumulus cells to induce successful ovulation process.

SUBMITTER: Kitasaka H

PROVIDER: S-EPMC5805282 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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Inductions of granulosa cell luteinization and cumulus expansion are dependent on the fibronectin-integrin pathway during ovulation process in mice.

Kitasaka Hiroya H Kawai Tomoko T Hoque S A Masudul SAM Umehara Takashi T Fujita Youko Y Shimada Masayuki M

PloS one 20180208 2

It has been known that EGF-like factor secreted from LH-stimulated granuloma cells acts on granulosa cells and cumulus cells to induce ovulation process. Granulosa cells are changed the morphology with differentiating cell functions to produce progesterone. Cumulus cells are detached to make a space between the cells to accumulate hyaluronan rich matrix. LH also changes extracellular matrix (ECM) components including fibronectin in the follicular walls and granulosa cell layers. EGF like factor ...[more]

PMID: 29420611

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Project description:During follicle maturation, oxygen levels continuously decrease in the follicular fluid and reach lowest levels in the preovulatory follicle. The current study was designed to comprehensively understand effects of low oxygen levels on bovine granulosa cells (GC) using our established estrogen active GC culture model. As evident from flow cytometry analysis the viability of GC was not found to be affected at severely low oxygen condition (1% O2) compared to normal (atmospheric) oxygen condition (21% O2). Estimations of hormone concentrations using competitive radioimmunoassay revealed that the production of estradiol and progesterone was significantly reduced at low oxygen condition. To understand the genome-wide changes of gene expression, mRNA microarray analysis was performed using Affymetrix's Bovine Gene 1.0 ST Arrays. This resulted in the identification of 1104 differentially regulated genes of which 505 were up- and 599 down-regulated under low oxygen conditions. Pathway analysis using Ingenuity pathway analyzer (IPA) identified 36 significantly affected (p < 0.05) canonical pathways. Importantly, pathways like "Estrogen-mediated S-phase Entry" and "Cyclins and Cell Cycle Regulation" were found to be greatly down-regulated at low oxygen levels. This was experimentally validated using flow cytometry based cell cycle analysis. Up-regulation of critical genes associated with angiogenesis, inflammation, and glucose metabolism, and down-regulation of FSH signaling, steroidogenesis and cell proliferation indicated that low oxygen levels induced early luteinization associated changes in granulosa cells. Identification of unmethylated CpG sites in the CYP19A1 promoter region suggests that granulosa cells were not completely transformed into luteal cells under the present low oxygen in vitro condition. In addition, the comparison with earlier published in vivo microarray data indicated that 1107 genes showed a similar expression pattern in granulosa cells at low oxygen levels (in vitro) as found in preovulatory follicles after the LH surge (in vivo). Overall, our findings demonstrate for the first time that low oxygen levels in preovulatory follicles may play an important role in supporting early events of luteinization in granulosa cells.

Dataset Information

Inductions of granulosa cell luteinization and cumulus expansion are dependent on the fibronectin-integrin pathway during ovulation process in mice.

Publications

Inductions of granulosa cell luteinization and cumulus expansion are dependent on the fibronectin-integrin pathway during ovulation process in mice.

Similar Datasets

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