| ObjectivesTo observe the clinical efficacy of Xuling Jiangu prescription in the treatment of postmenopausal osteoporosis of kidney deficiency and blood stasis type.The effect of Xuling Jiangu prescription on IL-17 RA 、 Act1 、 FOSB in the IL-17 signaling pathway was observed through transcriptome analysis and in vivo and in vitro experiments to explore its mechanism.Methods1.Postmenopausal osteoporosis patients with kidney deficiency and blood stasis were randomly divided into experimental group of 35 cases and control group of 32 cases,which were treated with Xuling Jiangu granules and calcitriol capsule respectively on the basis of taking calcium gluconate tablets orally.The course of treatment was 6 months.The TCM syndrome scores,bone mineral density,bone metabolism and safety indexes before and after treatment were recorded.2.The peripheral blood samples of 6 patients in the experimental group before and after treatment and 6 subjects in the normal group without osteoporosis were collected,and the differentially expressed genes were screened by high-throughput sequencing technology of transcriptomics and functional enrichment analysis was performed.Quantitative real-time PCR was used to detect the m RNA expression of IL-17 RA 、 Act1 、 FOSB in the IL-17 signaling pathway in the peripheral blood of 27 patients in the experimental group before and after treatment and 27 subjects in the normal group without osteoporosis.3.Three months old male SD rats were selected for preparation of drug-containing serum;osteoclast precursor cell line Raw264.7 was used to induce osteoclast differentiation,and quantitative real-time PCR was used to detect osteoclast differentiation marker genes CTSK,TRAP,MMP-9 m RNA expression.Drug-containing serum and pathway inhibitor(17-AGG)were used for intervention.They were divided into control group,inhibitor group,drug-containing serum+inhibitor group,blank serum group,drug-containing serum group.After 24 h,48h and 96 h of intervention,RNA and protein of five groups of cells were extracted.The m RNA and protein expressions of IL-17 RA,Act1,and FOSB in the IL-17 signaling pathway were detected by quantitative real-time PCR and Western-blot.Results1.In the experimental group,35 cases were enrolled,8 cases were dropped out,and 27 cases were completed;32 cases were enrolled in the control group,5 cases were dropped out,and 27 cases were completed.Compared with the total effective rate of TCM syndromes,the experimental group was significantly better than the control group,and the difference was statistically significant(P<0.05).After treatment,TCM syndrome scores in both groups were significantly decreased(P<0.05),but there was no significant difference between the two groups(P>0.05).There was no significant difference in BMD between groups and within the group(P>0.05).There was no significant difference in bone metabolism indexes between groups and within the experimental group(P>0.05),but the experimental group increased PINP and decreased CTX after treatment.There were no serious adverse reactions in either group.2.A total of 598 differentially expressed genes were screened in the experimental group before treatment and in the normal group without osteoporosis,and 319 differentially expressed genes were screened before and after treatment.Comparing 598 and 319 differentially expressed genes,a total of 19 common differentially expressed genes were found including FOSB,H4C2,RAMP3,etc.When pathway analysis was performed,it was found that only FOSB genes were enriched in the IL-17 signaling pathway.Compared with patients,the normal group without osteoporosis had lower IL-17 RA m RNA expression,but the difference was not statistically significant(P>0.05),Act1 m RNA expression was significantly lower,while FOSB m RNA expression was significantly higher(P<0.05).Compared with before treatment,the expressions of IL-17 RA and Act1 m RNA were decreased after Xuling Jiangu treatment(P>0.05),while the expression of FOSB m RNA was significantly increased(P<0.05).3.Compared with the control group,the number of osteoclasts differentiated and the m RNA expressions of differentiation marker genes CTSK,TRAP and MMP-9 in the drug-containing serum group were significantly reduced after intervention(P<0.05).Compared with the control group,the expressions of IL-17 RA,Act1 m RNA and protein in the inhibitor group and the drug-containing serum group decreased(P<0.05);Compared with the blank serum group,the m RNA and protein expressions of IL-17 RA and Act1 were lower after the drug-containing serum intervention,but the difference was not statistically significant(P>0.05).Compared with the drug-containing serum group,the expressions of IL-17 RA and Act1 proteins in the drug-containing serum +inhibitor group decreased,but the difference was not statistically significant(P>0.05).Similarly,compared with the control group,the expression of FOSB m RNA and protein in the inhibitor group also decreased(P<0.05),and the m RNA expression in the drug-containing serum group increased,but the change in protein expression was not statistically significant(P>0.05).Conclusions1.Xuling Jiangu prescription can relieve the clinical symptoms of postmenopausal osteoporosis patients with kidney deficiency and blood stasis syndrome.The prescription has the similar effect on bone mineral density as calcitriol,can improve bone metabolism markers,and it is safe.2.The results of transcriptomic analysis suggested that the therapeutic mechanism of Xuling Jiangu prescription might be related to the regulation of the expression of genes related to the IL-17 signaling pathway.3.Xuling Jiangu prescription affects the expression of IL-17 RA,Act1 and FOSB m RNA on the IL-17 signaling pathway and Xuling Jiangu prescription-containing serum can inhibit the differentiation of osteoclasts,and its mechanism of action is related to regulating the expression of genes related to the IL-17 signaling pathway and inhibiting the transmission of downstream signals. |
PDF Full Text Request