Effects Of GDF11 On The Proliferation And Differentiation Of Osteoclast-like Cells And UMR106 Osteoblast-like Cells

Research background and purpose:Osteoporosis(OP)is a common bone metabolism-related disease.The proliferation and differentiation of osteoblasts(OBs)are inhibited,resulting in weakened bone formation,and increase with age or hormone changes in the body lead to increased bone resorption capacity of osteoclasts(OCs),which may be the main cause of osteoporosis.Changes in the expression of some genes and proteins directly affect the proliferation and differentiation of osteoclasts and osteoblasts,and further affect bone formation and bone resorption.RANKL is a crucial factor in the process of osteoclast proliferation and differentiation.RANKL acts on RANK receptor on the pre-osteoclast cell membrane transmits signals to tumor necrosis factor receptor-associated factors(TRAF6).TRAF6 is an important linker in the process of osteoclast formation,it can cause a chain reaction of downstream signals thereby promoting osteoclast's differentiation,proliferation and bone resorption.The strength of tartrate-resistant acid phosphatase(TRAP)activity reflects the differentiation of osteoclasts,which is a sign of osteoclast differentiation and maturity,TRAP-positive multinucleated cells can be considered mature osteoclasts.Runx2 is a specific transcription factor for osteoblast differentiation,it associated with osteoblast gene expressions,at the same time,it can promote the secretion of extracellular matrix by osteoblasts.As the downstream transcription factor of Runx2,Osterix directly regulate the bone formation of osteoblasts as a factor of osteoblast bone formation.Researching the biological activities of osteoblasts and osteoclasts to exploring effective and less side effects drugs for osteoporosis must be the main goals of scientific bone metabolism research.As a member of the superfamily of growth transforming factor-?(TGF-?),growth differentiation factor 11(GDF11)involved in the development of multiple systems such as the skeletal system,the nervous system,and the kidney and other organs.At the same time,the bones system also has its gene and protein expression.However,its effect on bone metabolism is not clear at present.The main purpose of this study is to explore the effect and mechanism of GDF11 on osteoblasts and osteoclasts.Materials and Methods:The first part: The osteoclast-like cells were deal with blank control group,0.1,1,2,5,10,20,40 ng/ml GDF11 group,then CCK-8 method was used to detect the proliferation of cells.Osteoclast-like cell differentiation detected by Tartrate-resistant acid phosphatase kit.Bone resorption capacity of cells determined by bone resorption test.qPCR assay the expression of RANK,TRAF6 and p38 mRNA in the cells.Part 2: Cultured UMR106 cells and divided them into blank control group and experimental group including GDF11 group,GDF11 + siRNA group and siRNA group.CCK-8 method was used to detect cell proliferation.The expressions of Runx2 and Osterix mRNA were determined by qPCR.Result:The first part1.Under the 400 x microscope,RAW264.7 cells can be seen round nucleus,the cytoplasm is transparent,and the morphological characteristics and growth trends are similar to osteoclasts.2.Compared with the control,GDF11 at concentrations of 0.1,1 and 2 ng/ml promoted cell proliferation,and 40 ng/ml GDF11 inhibited cell proliferation.3.Compared with the control group,the number of TRAP positive Multi-core osteoclasts in the GDF11 group increased obviously.4.Compared with the control group,the number of bone resorption pores in each group was increased compared with the control group.5.Compared with the control group,the expression of RANK and TRAF6 mRNA in the cells of GDF11,GDF11 + RANKL and RANKL groups were significantly increased;the expression of p38 mRNA in GDF11 and RANKL cells was significantly increased.The second part1.Compared with the control group,the expression of Runx2 mRNA in siRNAtransfection group decreased obviously.2.Compared with the control group,the cell proliferation of the GDF11 group was promoted;compared with the siRNA + GDF11 and siRNA group,GDF11 cell proliferation was significantly increased.3.Compared with the control group,the expressions of Runx2 and Osterix mRNA in the GDF11 group were obviously increased;while the expressions of Runx2 and Osterix mRNA in the GDF11 + siRNA group and the siRNA group were significantly reduced;compared with the GDF11 group,the Runx2 and Osterix mRNA expression of cells in the GDF11 + siRNA and siRNA group were reduced obviously.Conclusion:1.GDF11 promotes RAW264.7 osteoclast-like cell proliferation.2.GDF11 may promote osteoclast-like cell proliferation and differentiation by activating the RANK,TRAF6 and P38 signaling pathway.3.GDF11 enhances the expression of Osterix by promoting the Runx2 signaling pathway,thereby promoting the proliferation of UMR106 cells.