Effects Of Chemokine-like Receptor 1 Gene Deletion On Bone Formation Induced By Dihydrotestosterone In Male Mice

Osteoporosis is a common bone metabolic disease,the prevalence of the disease is rising with the aging of society and it seriously affects human health.Because of the increasing incidence of osteoporosis in elderly men,the mechanism of androgen action on bone has become the focus of attention.Osteoblasts,osteoclasts,bone cells and bone marrow mesenchymal stem cell surface of both have androgen receptor,androgen is directly through the androgen receptor on bone cells to regulat osteoblast.Bone marrow mesenchymal stem cells can be transformed into different types of cells,the fate of the transformation is regulated by extracellular signal molecules.CMKLR1,a factor of Chemerin and its receptor,can mediate immune,inflammatory,metabolic,reproductive and cancer diseases.Chemerin and its receptor CMKLR1 coordination can promote bone marrow mesenchymal stem cells into fat cells,and the role of signaling pathways mediated by CMKLR1 in bone formation has not been reported so far,this article mainly explains the effects of dihydrotestosterone and CMKLR1 signaling pathway on bone formation.In order to elucidate the DHT/CMKLR1 signaling pathway in mouse bone marrow mesenchymal stem cells affect the differentiation ability of mice and the metabolism of bone tissue,this paper mainly from the in vivo and in vitro experiments explored.In vivo experiments,we injected α-NETA and DHT into the body of 25-day-old female C57 mice were divided into control group,DHT group and DHT + α-NETA group,5 mice in each group.After 21 days treated,bone mineral density,trabecular number and bone volume/total volume of femurs and tibias were analyzed from bone micro-CT;using fluorescence quantitative methods to detect ALP,Runx2 and Col1a2 expression to investigate the effect of DHT on bone metabolism after CMKLR1 was antagonized.In vitro experiments were performed to isolate and culture BMSCs from CMKLR1 knockout mice and wild-type malemice,and to observe the osteogenic differentiation ability of BMSCs under the induction of DHT.By adding α-NETA and transient transfection of CMKLR1 siRNA in BMSCs of wildtype mice to antagonistic the CMKLR1 with Chemerin combination and reduce the expression of CMKLR1,through alkaline phosphatase staining to detect ALP activity,through alizarin red staining to detect formation of calcium nodules,through the fluorescence quantitative to detect ALP,Runx2 and Osterix expression,through western blot detect the Runx protein change.The results showed that in vivo DHT increased the bone mineral density,trabecular number and bone volume/total volume bone,bone formation related factor ALP,Runx2 and Col1a2 were also up-regulated;injection of α-NETA,the indicators have declined.In vitro experiment,the results ealso showed alkaline phosphatase activity and formation of calcium nodules were reduced after deletion of CMKLR1,the osteogenic related gene expression and Runx2 protein are also down-regulated.Therefore,DHT can promote bone marrow mesenchymal stem cells to differentiate into osteoblasts,and promote osteoblast mineralization,the deletion of CMKLR1 gene can inhibit the bone marrow mesenchymal stem cells into bone differentiation after DHT induced,Possibly by regulating the osteogenic differentiation of maker related gene Runx2,ALP implementation.Chemerin/CMKLR1 signaling pathway is inhibited and affeced the effect of androgen on bone metabolism after CMKLR1 gene deleted.