The effect of Bushen Huoxue prescription on proliferation and extracellular matrix synthesis of nucleus pulposus cells by regulating circ_0036763/miR-583 axis
Objective To explore the possible effects of Bushen Huoxue Formula(the kidney tonifying and blood activating prescription)on the proliferation and extracellular matrix synthesis of nucleus pulposus cells by regulating the circ_0036763/miR-583 axis.Methods Real time fluorescence quantitative polymerase chain reaction(qRT-PCR)was used to detect the expression levels of circ0036763 and miR-583 in normal and intervertebral disc degeneration(IDD)nucleus pulposus cells;IDD nucleus pulposus cells were divided into pcDNA group,pcDNA circ_0036763 group,pcDNA circ_0036763+mimic NC group,and pcDNA circ_0036763+miR-583 mimic group.qRT-PCR was used to detect the expression levels of circ_0036763 and miR-583 in nucleus pulposus cells in each group,methyl thiazolyl tetrazolium(MTT)was used to detect cell proliferation(A value),and Western blot was used to detect the expression of proliferating cell nuclear antigen(PCNA),collagen Ⅰ,and collagen Ⅱ proteins in nucleus pulposus cells,The dual luciferase assay reported experimental validation of the targeting relationship between circ_0036763 and miR-583.27 mice were divided into sham surgery group,IDD group,and kidney tonifying and blood activating formula group.IDD models were established in all groups except for the sham surgery group.After successful modeling,the sham surgery group and IDD group were given physiological saline by gavage,while the kidney tonifying and blood activating formula group was given 1.5 g/ml of kidney tonifying and blood activating formula by gavage for 3 consecutive weeks.QRT-PCR was used to detect the expression levels of circ0036763 and miR-583 in the nucleus pulposus cells of mice in each group,MTT was used to detect cell proliferation,and Western blot was used to detect the expression of PCNA,collagen Ⅰ,and collagen Ⅱ proteins.Results The expression level of circ_0036763 in IDD nucleus pulposus cells decreased,while the expression level of miR-583 increased(all P＜0.05);Overexpression of circ_0036763 can promote proliferation and extracellular matrix synthesis of nucleus pulposus cells(all P＜0.05);Circ_0036763 targets miR-583 and upregulates miR-583 reversible overexpression.Circ_0036763 enhances the proliferation and extracellular matrix synthesis ability of IDD nucleus pulposus cells.Compared with the sham surgery group,the IDD group showed an increase in collagen Ⅰ protein expression and miR-583 expression levels(all P＜0.05),while the cell A value,PCNA and collagen Ⅱ protein expression,and circ_0036763 expression levels decreased(all P＜0.05);Compared with the IDD group,the Kidney Tonifying and Blood Activating Formula group showed a decrease in collagen Ⅰ protein expression and miR-583 expression levels(all P＜0.05),while the cell A value,PCNA and collagen Ⅱprotein expression,and circ_0036763 expression levels increased(all P＜0.05).Conclusions The kidney tonifying and blood activating formula(Bushen Huoxue)may induce proliferation and extracellular matrix synthesis of nucleus pulposus cells by regulating the circ_0036763/miR-583 axis.
维普
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