The effects of mouse ovarian granulosa cell function and related gene expression by suppressing BMP/Smad signaling
Li Zhang1, Hejian Wang1, Daolun Yu1
1College of Life Science, Anhui provincial Key Lab of the Conservation and Exploitation of Biological Resources, Anhui Normal University, Wuhu, People's Republic of China.
BMP I type receptor inhibitor can selectively inhibit BMP/Smad signaling pathways, mainly by inhibiting the BMP I type receptor activity to prevent phosphorylation of Smad1, Smad5 and Smad9. The aim of the present study was to explore the effects of mouse ovarian granulosa cell function and related gene expression by suppressing BMP/Smad signaling pathway with LDN-193189(A type of BMP I type receptor inhibitor). In this study, we cultivate the original generation of mouse ovarian granular cells then collect cells and cell culture medium after treatment. Cellular localization and expression of Smad9 and P-smad9 proteins was studied by immunofluorescence (IF) in the ovarian granulosa cells of mouse; Related genes mRNA and proteins expression was checked by QRT-PCR and Western blot; Detected the concentration of related hormones by using ELISA kit; finally, the growth of the cells was analyzed by plotting cell growth curve with CCK-8 assay. The results indicate that, suppression of BMP/Smad signaling pathway can inhibit the expression of LHR and FSHR, inhibit cell proliferation and decrease E2 secretion, the mechanism of action maybe reduce the expression of smad9, at the same time, we found that the feedback regulation of smad9 may affect the expression of FSHR and cell proliferation.
BMP I type receptor inhibitor can selectively inhibit BMP/Smad signaling pathways, mainly by inhibiting the BMP I type receptor activity to prevent phosphorylation of Smad1, Smad5 and Smad9. The aim of the present study was to explore the effects of mouse ovarian granulosa cell function and related gene expression by suppressing BMP/Smad signaling pathway with LDN-193189(A type of BMP I type receptor inhibitor). In this study, we cultivate the original generation of mouse ovarian granular cells then collect cells and cell culture medium after treatment. Cellular localization and expression of Smad9 and P-smad9 proteins was studied by immunofluorescence (IF) in the ovarian granulosa cells of mouse; Related genes mRNA and proteins expression was checked by QRT-PCR and Western blot; Detected the concentration of related hormones by using ELISA kit; finally, the growth of the cells was analyzed by plotting cell growth curve with CCK-8 assay. The results indicate that, suppression of BMP/Smad signaling pathway can inhibit the expression of LHR and FSHR, inhibit cell proliferation and decrease E2 secretion, the mechanism of action maybe reduce the expression of smad9, at the same time, we found that the feedback regulation of smad9 may affect the expression of FSHR and cell proliferation.
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