| Background:Osteoporosis is characterized by loss of bone mass and destruction of bone structure,which leads to a decrease in bone strength and an increase in the risk of fracture,and has a serious negative impact on the general health and quality of life of postmenopausal women and the elderly.It is estimated that osteoporosis affects 200 million women worldwide,and its prevalence increases with age: the prevalence rate is about 10% among women over 60 years old,about 20% of women over70 years old,about 40% of women over 80 years old,and more than 60%of women over 90 years old.Osteoporosis is classified into primary and secondary osteoporosis.The primary osteoporosis is still of unclear etiology and complex pathogenesis,therefore,its treatment remains unsatisfactory,such as late intervention,poor specificity,unstable long-term effect,high cost and so on.Further understanding of the pathophysiological changes and micro-mechanism in the pathogenesis of osteoporosis will help to find a more efficient,economical,targeted and less complication treatment.In recent years,macrophages have attracted much attention as important participants in bone remodeling and repair.Previous studies generally believed that the effect of macrophages on osteoporosis is mainly to regulate the inflammatory response,aging leads to bone loss and causes macrophage dysfunction,aggravates the inflammatory response,releases inflammatory factors to aggravate bone resorption.However,there are relatively few studies on macrophages regulating bone formation.In recent years,with the in-depth study of extracellular vesicles,exosome-mediated intercellular communication has opened up a new perspective for the pathogenesis of complex diseases.Exosome transported proteins and micro RNAs have been confirmed to be involved in the epigenetic regulation of many diseases.It has been found that M1 macrophage exocrine can transport mi R-222 to induce apoptosis of bone marrow mesenchymal stem cells;M2 macrophage derived exosome mi RNA-5106 can induce bone marrow mesenchymal stem cells to differentiate into osteoblasts by targeting salt-induced kinase 2 and 3;exocrine from bone marrow macrophages can transport micro RNAs into bone marrow mesenchymal stem cells,resulting in increased bone loss in mice.In general,macrophage derived exosomes play an important role in the regulation of primary osteoporosis,but its regulatory mechanism has not been fully elucidated.The purpose of this study is to explore the mechanism of bone marrow macrophage exocrine regulating the differentiation of bone marrow mesenchymal stem cells by transporting mi R-670-5p,and to further explain the way that bone marrow macrophages participate in regulating the pathogenesis of osteoporosis,so as to provide new ideas for the prevention and treatment of primary osteoporosis.Objective: to explore the role and mechanism of bone marrow macrophage derived exosomal mi R-670-5p in regulating the differentiation of bone marrow mesenchymal stem cells(BMSCs),and to further explain the pathogenesis of primary osteoporosis and provide a new target for its diagnosis and treatment.Method:(1)Primary bone marrow macrophages from aged and young mice were extracted.Exosomes from bone marrow macrophages were isolated from culture medium and identified by electronic microscope,particle size analysis and Western blot.Bone marrow macrophage exosomes were then co-cultured with BMSC,after which the BMSCs were induced to differentiation.ALP,ARS,ORO staining and q RT-PCR were used to compare the effects of bone marrow macrophage exocrine bodies of different age groups on osteogenic differentiation and adipogenic differentiation of BMSCs.The differentially expressed micro RNAs in bone marrow macrophage exocrine bodies of aged mice and young mice were found by micro RNA array and verified by q RT-PCR in mice and human patients.(2)The effect of mi R-670-5p,which had significantly increased expression in bone marrow macrophages of aged mice on BMSC differentiation was investigated by transfection experiment,differentiation induced experiment,staining,q RT-PCR and immunofluorescence experiments.The in vivo effect of mi R-670-5p mimic was investigated by intramedullary injection and transfection,snd the changes of bone structure were detected by micro-CT,HE staining and OCN immunohistochemical staining.The target gene of mi R-670-5p was predicted by Targetscan,mi RWalk,DIANAmicro T,and Tarbase,verified by q RT-PCR,Western blot and double luciferase reporter experiment,and its effect on Wnt/ β-catenin signaling pathway was further explored.(3)The mouse model of primary osteoporosis was established by ovariectomy.The in vivo effect of mi R-670-5p inhibitor was investigated by intramedullary injection and transfection.The changes of bone structure were detected by micro-CT,HE staining and OCN immunohistochemical staining.Result:(1)compared with young mice,the bone marrow macrophage derived exosomes of aged mice inhibited the osteogenic differentiation of BMSC and promoted its adipogenic differentiation.The expression of mi R-670-5p significantly increased in bone marrow macrophage exosomes from aged mouse,and showed the same trend in exosomes of patients with primary osteoporosis;(2)mi R-670-5p could inhibit the osteogenic differentiation of BMSC and promote its adipogenic differentiation,and can promote bone loss in vivo.Mi R-670-5p can target ESRRA gene,downregulate its m RNA and protein levels,and downregulate Wnt/β-catenin signaling pathway;(3)inhibition of mi R-670-5p can partially save the impaired osteogenic differentiation of BMSC in ovary missed mice and inhibit its adipogenic differentiation.Inhibition of mi R-670-5p can reduce bone loss in ovary missed mice in vivo.Conclusion: The expression of bone marrow macrophage exosomal mi R-670-5p is associated with primary osteoporosis.Bone marrow macrophage derived exosomes transport more mi R-670-5p into bone marrow mesenchymal stem cells.Mi R-670-5p down-regulates the activity of Wnt/ β-catenin signal pathway through targeted inhibition of ESRRA gene expression,inhibits osteogenic differentiation of bone marrow mesenchymal stem cells and promotes adipogenic differentiation of bone marrow mesenchymal stem cells,thus promoting the occurrence and development of primary osteoporosis.Inhibition of mi R-670-5p can partially save the osteogenic differentiation ability of bone marrow mesenchymal stem cells in osteoporotic mice,inhibit their adipogenic differentiation,and effectively improve bone loss and bone marrow steatosis in mice,which has a certain potential for the prevention and treatment of primary osteoporosis.Figures: 25;tables: 14;references: 145. |
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