Background： Estrogen, as an important hormone in human physiological process, is closely related to bone metabolism. The aim of this study was to investigate the mechanism of estrogen on osteoblasts metabolisrn in MC...Background： Estrogen, as an important hormone in human physiological process, is closely related to bone metabolism. The aim of this study was to investigate the mechanism of estrogen on osteoblasts metabolisrn in MC3T3-EI cells. Methods： We treated the MC3T3-EI cells with different concentrations of β-estradiol （0.01, 0.1, 1, and 10 nmol/L）, observed the morphological changes of the cells, and detected the cell＇s proliferation and apoptosis of MC3T3-EI cells. Two transcriptome libraries were constructed and sequenced. Quantitative real-time polymerase chain reaction （qRT-PCR） was performed to confirm the differentially expressed genes （DEGs）, and then treated the MC3T3-EI cells with estrogen receptor （ER） inhibitors α and β, respectively, and then examined the expression of Tgfbr1 and Bmprla genes. The promoter of Tg/brl and Bmprla gene was analyzed, and the ER response elements were identified. Finally, Chip was used to verity the binding of ER to Tgfbr1 and Bmprla promoter. Results： In the high-concentration β-estradiol treatment group （1 nmol/L and 10 nmol/L）, there was no significant difference in the morphology of the cells under the microscope, 1 nmol/L and 10 nmol/L treated group appeared statistically significant difference in cell apoptosis and proliferation （P 〈 0.05 and P 〈 0.01, respectively）. We found 460 DEGs compared with the control group. Among the DEGs, there were 66 upregulated genes and 394 downregulated genes. Gene ontology classification and Kyoto Encyclopedia of Genes and Genornes pathway analysis revealed that many bone metabolism-related biological processes and cell signaling pathways were disordered. The qRT-PCR verification showed that the expressions of Tgfbrl- and Bmprl a-related genes in bone metabolism pathway in the 10 nmol/L treatment group were significantly decreased （P 〈 0.05）. ER β was involved in the inhibitory effect of Tgfbr1 and Bmprla genes. The bioinformatics of the promoter found that there were three ER response elements in the promoter of Tgfbr1, and there were two ER response elements in Bmprla promoter regions. Chip experiments showed that estrogen could enhance the binding of ERs to Tgfbr1 and Bmprla genes. Conclusions： Estrogen can promote the apoptosis and proliferation of osteoblasts simultaneously, and the mechanism may be the joint action of transforming growth factor-beta, Wnt, mitogen-activated protein kinase, and nuclear factor-kappaB bone metabolism-related signaling pathway. Estrogen inhibits the expression of Tgfhr1 and Bmrla genes through ER β and affects the metabolism of MC3T3-E 1 osteoblasts.展开更多
文摘Background： Estrogen, as an important hormone in human physiological process, is closely related to bone metabolism. The aim of this study was to investigate the mechanism of estrogen on osteoblasts metabolisrn in MC3T3-EI cells. Methods： We treated the MC3T3-EI cells with different concentrations of β-estradiol （0.01, 0.1, 1, and 10 nmol/L）, observed the morphological changes of the cells, and detected the cell＇s proliferation and apoptosis of MC3T3-EI cells. Two transcriptome libraries were constructed and sequenced. Quantitative real-time polymerase chain reaction （qRT-PCR） was performed to confirm the differentially expressed genes （DEGs）, and then treated the MC3T3-EI cells with estrogen receptor （ER） inhibitors α and β, respectively, and then examined the expression of Tgfbr1 and Bmprla genes. The promoter of Tg/brl and Bmprla gene was analyzed, and the ER response elements were identified. Finally, Chip was used to verity the binding of ER to Tgfbr1 and Bmprla promoter. Results： In the high-concentration β-estradiol treatment group （1 nmol/L and 10 nmol/L）, there was no significant difference in the morphology of the cells under the microscope, 1 nmol/L and 10 nmol/L treated group appeared statistically significant difference in cell apoptosis and proliferation （P 〈 0.05 and P 〈 0.01, respectively）. We found 460 DEGs compared with the control group. Among the DEGs, there were 66 upregulated genes and 394 downregulated genes. Gene ontology classification and Kyoto Encyclopedia of Genes and Genornes pathway analysis revealed that many bone metabolism-related biological processes and cell signaling pathways were disordered. The qRT-PCR verification showed that the expressions of Tgfbrl- and Bmprl a-related genes in bone metabolism pathway in the 10 nmol/L treatment group were significantly decreased （P 〈 0.05）. ER β was involved in the inhibitory effect of Tgfbr1 and Bmprla genes. The bioinformatics of the promoter found that there were three ER response elements in the promoter of Tgfbr1, and there were two ER response elements in Bmprla promoter regions. Chip experiments showed that estrogen could enhance the binding of ERs to Tgfbr1 and Bmprla genes. Conclusions： Estrogen can promote the apoptosis and proliferation of osteoblasts simultaneously, and the mechanism may be the joint action of transforming growth factor-beta, Wnt, mitogen-activated protein kinase, and nuclear factor-kappaB bone metabolism-related signaling pathway. Estrogen inhibits the expression of Tgfhr1 and Bmrla genes through ER β and affects the metabolism of MC3T3-E 1 osteoblasts.
