| Purpose: Based on the theory of “the kidney stores the essence,produces the marrow and controls the bones” in traditional Chinese medicine,the effect of Zuogui pill drug-containing serum on proliferation and differentiation of rat BMSCs into osteoblasts in vitro was observed through experimental research,and the changes of SHH,SMO,GLI1 and BMP2,GPR48 and other related factor protein expressions in Hedgehog signaling pathway of cells were also observed,so as to explore the relevant utility mechanism of Zuogui pill in preventing and treating OP,and provide a relatively novel experimental basis for clinical treatment of OP.Material and method: Thirty-two SPF female barren rats weighing about 200-230g(3months old)were randomly divided into 4 groups: normal group,induction liquid group,fosamax group and Zuogui pill group,with 8 rats in each group.After adaptive feeding of the rats for 3 days,the experimental rats were continuously fed with corresponding drugs in each group once a day for 5 days to prepare corresponding drug-containing serum.After 5 days,the rats were decapitated and killed,and the materials were obtained,and the drug-containing serum of 4 groups of rats was retained.The purchased Wistar rat BMSCs were resuscitated,propagated and passaged,and the cells were divided into 4 groups.Filtering the drug-containing clear liquid,and then culturing rat BMSCs in vitro with a culture medium containing drug-containing serum of corresponding groups of drugs.One group is added with induction liquid,and the other three groups are respectively added with the prepared drug-containing clear liquid.ELISA was used to detect the protein expression of SHH,SMO,GLI1,ALP,GPR48,BMP2 and other related factors in Hedgehog signal pathway of each group of cells.SPSS20.0 software was used for statistical analysis,and the experimental data were compared and analyzed,so as to compare the effect of drug-containing serum of each group on BMSCs differentiation into osteoblasts,and to explore the mechanism of Zuogui pill in preventing and treating OP and the difference in the degree of influence.Results:1.the detection results of ALP concentration in cells in each experimental group showed that compared with the normal group,the zuogui pill group,the inducing liquid group and the fosamax group increased significantly(p < 0.01);The detection results on the 12 th day are relatively better than that on the 9th and 15 th day.2.the detection results of GPR48 concentration in cells in each experimental group showed that compared with the normal group,the zuogui pill group increased significantly(p < 0.01);Compared with the induction liquid group and the fosamax group,the zuogui pill group has significant difference(P<0.01).3.the detection results of BMP-2 concentration in the cells of each experimental group showed that compared with the normal group,the BMP-2 concentration in the inducing solution group,zuogui pill group and fosamax group was significantly increased(P<0.01).Among them,the inducing liquid group has the highest content,and the Zuogui pill group is higher than the normal group(P<0.01).4.the SHH concentration in BMSCs of rats cultured with drug-containing serum in each group showed that compared with the normal group,SHH in the induction fosamax group,zuogui pill group and liquid group was significantly increased(P<0.01).Among them,the inducing liquid group has the highest content,and the Zuogui pill group is higher than the normal group(P<0.01).5.the detection results of Smo concentration in BMSCs of rats cultured with drug-containing serum in each group showed that compared with the normal group,the Smo levels in the induction fosamax group,zuogui pill group and liquid group were significantly increased(P<0.01).Among them,the inducing liquid group has the highest content,and the Zuogui pill group is higher than the normal group(P<0.01).6.the detection results of Gli1 concentration in BMSCs of rats cultured with drug-containing serum in each group showed that compared with the normal group,Gli1 concentration in the induction fosamax group,zuogui pill group and liquid group was significantly increased(P<0.01).Among them,the inducing liquid group has the highest content,and the Zuogui pill group is higher than the normal group,even higher than the fosamax group(P<0.01).Conclusion:1.ALP,GPR48,BMP2,Shh,SMO and Gli1 can be expressed in BMSCs of normal rats, which indicates that GPR48 signaling pathway and hedgehog signaling pathway are related signaling factors in cells of rats.2.Zuogui pill can up-regulate the protein concentration of ALP,GPR48,BMP2,Shh,SMO and Gli1 in the BMSCs of rats,which indicates that Zuogui pill may achieve the purpose of prevention and treatment of osteoporosis by activating Shh,SMO,Gli1 and other target genes in Hedgehog signal pathway and regulating the expression level of signal factors.3.The drug-containing serum of Zuogui pill can promote the differentiation of rat BMSCs into osteoblasts,which has been proved again,fully verifying the scientificity of the kidney marrow relationship of "kidney storing essence,main bone,generating marrow". |
PDF Full Text Request