Study On The Pathogenesis Of Postmenopausal Osteoporosis Rats From The SDF-1/CXCR4 Pathway Based On "spleen-kidney Correlation"

Purpose:This paper is guided by the TCM theory "spleen and kidney correlation",to explore if the bone and skeletal muscle in the SDF-1/CXCR4 signaling pathway related factors will be changed in protein expression when the rats with postmenopausal osteoporosis are prevented and treated by TCM treatments of nourishing the kidney-yin,warming the kidney-yang,tonifying spleen and tonifying kidney and spleen,and to analysis and extend the mechanism of the modern medicine of TCM prevention and treatment of osteoporosis.By observing the expression of SDF-1/CXCR4 signal pathway in the bone and muscle of rats under different treatment methods,we can provide experimental basis for selecting the best TCM treatment for osteoporosis.Material and method:A total of 91 female SPF Wistar rats were randomly divided into normal group,model group,tonifying kidney-yin group,tonifying kidney-yang group,invigorating spleen group,tonifying kidney and invigorating spleen group and Fosamax group.Ovariectomies were performed in all the rats except the normal group to establish the rat model of postmenopausal osteoporosis.Intragastric administration was started on the second day after surgery,once a day for 10 weeks.The normal group and model group were given0.9% sodium chloride solution,and the tonifying kidney-yin group,tonifying kidney-yang group,invigorating spleen group,tonifying kidney and invigorating spleen group and Fosamax group were given Zuogui pill,Yougui pill,Buzhong Yiqi Decoction,Bushen Jianpi Prescription and Fosamax liquid by intragastric administration,respectively.Anesthesia and specimen collection were performed after the last gavage.Abdominal aorta blood was extracted from open abdomen to make serum.The femur of left and right posterior limbs and the white muscle of left posterior gastrocnemius were extracted.The BMD of left posterior limb femur was detected by dual-energy X-ray absorptiometry,and the protein expressions of ALP and TRACP in serum,SDF-1,CXCR4,PI3 K and HIF-1? in bone and skeletal muscle of each group were detected by enzyme linked immunosorbent assay.Results:Part 1: Model establishment and evaluation1.Bone mineral density measurement results: Compared with the normal group,the density of left hindlimb femur of the model group was significantly decreased(P<0.01);Compared with model group,BMD of rats in all treatment groups was significantly increased(P<0.01).2.Determination of bone metabolism indexes: Compared with the normal group,serum ALP and TRACP levels in model group were significantly increased(P<0.01);Compared with model group,serum ALP and TRACP levels in treatment groups were significantly decreased(P<0.01),and the decrease was most obvious in Fosamax group,followed by tonifying kidney and invigorating spleen group.Part 2:Effects of different treatments of traditional Chinese medicine on SDF-1/CXCR4 signaling pathway in bone tissue and skeletal muscle of rats with postmenopausal osteoporosis.Experiment 1: Proteins expressions of SDF-1,CXCR4,PI3 K and HIF-1 ? in bone tissue under different treatments of TCM.1.The content of SDF-1 in bone tissue of model group was significantly higher than that of normal group(P<0.01).Compared with model group,the contents of SDF-1 in bone tissue of rats in tonifying kidney-yang group,Fosamax group and invigorating spleen group were significantly decreased(P<0.01),and the same was true in tonifying kidney-yin group(P<0.05).There was also a great degree of decrease in the tonifying kidney and invigorating spleen group,but it was not statistically significant.2.Compared with the normal group,the content of CXCR4 in bone tissue of rats in model group was significantly increased(P<0.01);Compared with model group,CXCR4 content in bone tissue of rats in all treatment groups was significantly decreased in different degrees(P<0.01),and the decrease was most obvious in Fosamax group,followed by tonifying kidney-yang group.3.Compared with the normal group,the expression level of PI3 K protein in bone tissue of rats in model group was significantly down-regulated(P<0.01);Compared with model group,the protein expression levels of PI3 K in Fosamax group,invigorating spleen group,tonifying kidney-yang group and tonifying kidney-yin group were significantly up-regulated(P<0.01),while the increase of PI3 K in tonifying kidney and invigorating spleen group was not statistically significant(P>0.05).4.Compared with the normal group,the expression of HIF-1? protein in bone tissue of rats in model group was significantly decreased(P<0.01);Compared with model group,the level of HIF-1? in bone tissue of all treatment groups was significantly increased(P<0.01),and the level of HIF-1 ? in Fosamax group was the highest,followed by invigorating spleen group.Experiment 2: Proteins expression of SDF-1,CXCR4,PI3 K and HIF-1 ? in skeletal muscle under different treatments of TCM.1.The expression of SDF-1 protein in skeletal muscle of model group was significantly higher than that of normal group(P<0.01).Compared with model group,the expression of SDF-1protein in skeletal muscle of rats in Fosamax group and invigorating spleen group was significantly decreased(P<0.01),and the expressions of SDF-1 protein in other treatment groups were also decreased,but there was no statistical significance.2.Compared with the normal group,the content of CXCR4 protein in skeletal muscle of the model group was significantly increased(P<0.01);Compared with model group,the treatment groups rats skeletal muscle CXCR4 content all have dropped significantly,when the decrease degree of tonifying kidney and invigorating spleen group(P<0.05)was lower than that of other treatment groups(P<0.01).The decrease of CXCR4 content in skeletal muscle of rats in Fosamax group was the most obvious,which was significantly higher than that in tonifying kidney-yin group and tonifying kidney and invigorating spleen group,and the difference was statistically significant(P<0.01).3.Compared with the normal group,the expression of PI3 K protein in skeletal muscle of rats in model group was significantly decreased(P<0.01);Compared with the model group,the protein expression levels of PI3 K in skeletal muscle of all treatment groups were increased to different degrees,and the expression level of PI3 K in Fosamax group was the most significant(P<0.01),even higher than that in the normal group,followed by the invigorating spleen group,tonifying kidney-yang group(P<0.01)and tonifying kidney-yin group(P<0.05).4.Compared with normal group,HIF-1 ? protein expression in skeletal muscle of model group was significantly decreased(P<0.01);Compared with model group,HIF-1? protein expression in skeletal muscle was significantly up-regulated in all treatment groups(P<0.01).Conclusion:1.The TCM treatments of nourishing the kidney-yin,warming the kidney-yang,tonifying spleen and tonifying kidney and spleen can improve the postmenopausal osteoporosis rats femur bone mineral density,significantly lower serum ALP,TRACP levels,showing that the methods of nourishing the kidney-yin,warming the kidney-yang,tonifying spleen and tonifying kidney and spleen can effectively reduce postmenopausal osteoporosis rats bone conversion rate,which is good for prophylaxis and prevention of postmenopausal osteoporosis.2.The methods of nourishing the kidney-yin,warming the kidney-yang,tonifying spleen and tonifying kidney and spleen can regulate related factors in the SDF-1/CXCR4 signaling pathway,inhibit bone resorption,promote bone formation,and play a prevention and treatment role in postmenopausal osteoporosis by down-regulating the expression of SDF-1and CXCR4 in bone tissue,up-regulating the expression of PI3 K and HIF-1? protein.3.The methods of nourishing the kidney-yin,warming the kidney-yang,tonifying spleen and tonifying kidney and spleen can adjust related protein expression on the SDF-1/CXCR4 signaling pathways in skeletal muscle,by reducing the skeletal muscle SDF-1 and CXCR4,increasing the PI3 K and HIF-1 alpha,which is in line with the regulation of bone.This indicates that the coordination of bone and muscle is enhanced,so as to promote bone formation and achieve the purpose of prevention and treatment of postmenopausal osteoporosis.4.Kidney deficiency is the main pathogenesis of postmenopausal osteoporosis,and spleen deficiency is also the key pathogenesis leading to postmenopausal osteoporosis.The methods of nourishing the kidney-yin,warming the kidney-yang,tonifying spleen and tonifying kidney and spleen can effectively treat postmenopausal osteoporosis.