Regulation Of BMP-2 Signal Pathway By Collagen Peptide And Chelated Calcium In Promoting The Activity Of Osteoblasts

Bone morphogenetic protein(BMP-2)plays a vital role in promoting bone formation and inducing osteogenic differentiation.The BMP-2/Smads/Runx2/Osterix signaling pathway plays a central role in the formation of bone marrow mesenchymal cells(BMSCs)into osteoblasts(OB)and the synthesis and secretion of bone extracellular matrix.The objective of this study was to investigate the effects of collagen peptide(CP),Calcium Chaleted with Collagen Peptide(CCCP),Maillard Reaction Product(CP-X),Amino acid chelate calcium(AA-Ca)and Estrogen(E2)on the prevention and treatment of Osteoporosis(OP),to reveal their basis molecular mechanism of BMP-2 signaling pathway on OP control and establishing and internal relationship between the effect of bone improvement and the relative mRNA expression of key genes in the BMP-2 signaling pathway in femoral tissue,laying a foundation for the development and efficient screening research of osteoporosis drugs.OP models were established by removing bilateral ovaries from rats and randomly divided into Model group,CP-L,M,H groups and set up sham group.The oral doses of CP in each dose group were 6.8,20.5,34 g/(kg·d).The Sham and Model groups were given an equal volume of distilled water for 8 w.The wet weight of the uterus and vagina,the serum indexes related with bone metabolism and the bone mineral content were measured,the microstructure of the bone was observed,and the relative expression of key genes of BMP-2 signaling pathway in the femur of each group was determined.The results showed that compared with the Sham group,the uterine morphology of the Model group was shrinking,and the uterine index and vaginal index were significantly reduced(P<0.01).Compared with the Model group,the uterine morphology of all CP dose groups was restored and the uterus index and vaginal index were increased.Serological results showed that compared with the Sham group,the RANKL and IL-6 levels in the serum of the Model group were significantly increased,and the OPG content and OPG/RANKL ratio were significantly decreased(P<0.01).Compared with the Model group,each dose group of CP can reduce the serum RANKL and IL-6content and increase the OPG content and OPG/RANKL ratio,and it has a dose-dependent relationship.The results of tibial bone calcium and bone phosphorus determination showed that the bone calcium and bone phosphorus contents of rats were significantly reduced after ovarian removal(P<0.01).Compared with the Model group,the bone calcium and bone phosphorus contents of the CP groups were significantly increased(P<0.01).Femoral section results showed that the number of trabeculae of rats was significantly reduced,the trabeculae were diluted and broken,and the bone marrow cavity was large and hollow after ovarian extraction.After CP intervention,the trabecular structure of rats in each dose group was restored and the bone microstructure of the CP-M group had the best recovery effect.Further analysis of Micro-CT results shows that compared with the Sham group,the bone volume fraction(BV/TV),the number of trabecular bones(Tb.N),the thickness of trabecular bones(Tb.Th)and the degree of beam connection(Conn.D)are extremely reduced,while the degree of trabecular bone separation(Tb.Sp)is extremely increased(P<0.01).Compared with the Model group,the BV/TV,Tb.N,Tb.Th and Conn.D,in the CP-M group were increased,and the Tb.Sp was decreased(P>0.05).The qRT-PCR results showed that after the ovary removed,the relative expression levels of BMP-2,Smad1,Runx2,and Osterix mRNA in femurs were significantly reduced(P<0.01);compared with the Model group,the relative expression levels of BMP-2,Smad1,Runx2,and Osterix mRNA were significantly increased in CP groups.With the exception of Smad1,the expression levels of other genes were highest in the middle-dose group.The results of previous studies on rats indicate that CCCP,CP-X,AA-Ca,have the preventive and therapeutic effects on osteoporosis.The results of bone mineral content,bone tissue microstructure,bone organic matter content,bone density and other indicators indicated that: E2>Sham>CCCP>Other test groups >Model.The qRT-PCR results of rattus indicated that the expressions of BMP-2,Smad1,Runx2,and Osterix mRNA in the femoral tissues of all the tested substances were significantly higher than those in the Model group(P<0.01).The expression levels of all key genes for medium and high dose groups were: E2>CCCP>Other test groups,which were highly consistent with bone condition.In summary,CP can not only improve cancellous bone through the RANKL/RANK/OPG system axis,but also positively regulate bone metabolism based on the BMP-2 signaling pathway.CCCP,CP-X,AA-Ca and estrogen can also improve bone quality and enhance bone formation based on the positive regulation of BMP-2 signaling pathway.The expression levels of BMP-2,Smad1,Runx2,and Osterix genes in this pathway are positively correlated with bone status,which is helpful for the development of osteoporosis drugs and efficient screening.