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通过抑制骨形态发生蛋白/信号转导和转录激活因子信号通路对小鼠卵巢颗粒细胞功能及相关基因表达的影响

The effects of mouse ovarian granulosa cell function and related gene expression by suppressing BMP/Smad signaling pathway.

作者信息

Zhang Li, Wang Hejian, Yu Daolun, Chen Jie, Xing Chaofeng, Li Jie, Li Jun, Cai Yafei

机构信息

College of Life Science, Anhui provincial Key Lab of the Conservation and Exploitation of Biological Resources, Anhui Normal University, Wuhu, People's Republic of China.

College of Animal Science and Technology, Nanjing Agricultural University, Nanjing, People's Republic of China.

出版信息

Anim Cells Syst (Seoul). 2018 Sep 19;22(5):317-323. doi: 10.1080/19768354.2018.1497706. eCollection 2018.

DOI:10.1080/19768354.2018.1497706
PMID:30460113
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC6171428/
Abstract

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 E 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.

摘要

BMP I型受体抑制剂可选择性抑制BMP/Smad信号通路，主要通过抑制BMP I型受体活性来阻止Smad1、Smad5和Smad9的磷酸化。本研究的目的是通过用LDN-193189（一种BMP I型受体抑制剂）抑制BMP/Smad信号通路，探讨其对小鼠卵巢颗粒细胞功能及相关基因表达的影响。在本研究中，我们培养原代小鼠卵巢颗粒细胞，处理后收集细胞和细胞培养基。通过免疫荧光（IF）研究小鼠卵巢颗粒细胞中Smad9和P-smad9蛋白的细胞定位和表达；通过QRT-PCR和蛋白质免疫印迹法检测相关基因的mRNA和蛋白表达；使用ELISA试剂盒检测相关激素的浓度；最后，用CCK-8法绘制细胞生长曲线分析细胞生长情况。结果表明，抑制BMP/Smad信号通路可抑制LHR和FSHR的表达，抑制细胞增殖并减少E的分泌，作用机制可能是降低smad9的表达，同时，我们发现smad9的反馈调节可能影响FSHR的表达和细胞增殖。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/c5629a0c651f/TACS_A_1497706_F0006_OB.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/c61f36ddc977/TACS_A_1497706_F0001_OC.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/784c40086003/TACS_A_1497706_F0002_OC.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/5182b443a410/TACS_A_1497706_F0003_OB.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/5219a02fd4ca/TACS_A_1497706_F0004_OB.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/ed6b66d68f98/TACS_A_1497706_F0005_OB.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/c5629a0c651f/TACS_A_1497706_F0006_OB.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/c61f36ddc977/TACS_A_1497706_F0001_OC.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/784c40086003/TACS_A_1497706_F0002_OC.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/5182b443a410/TACS_A_1497706_F0003_OB.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/5219a02fd4ca/TACS_A_1497706_F0004_OB.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/ed6b66d68f98/TACS_A_1497706_F0005_OB.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/21e1/6171428/c5629a0c651f/TACS_A_1497706_F0006_OB.jpg

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