Study On Expression Patterns Of KLF2and KLF15during The Lineage Commitment Process Of Human Bone Marrow Mesenchymal Stem Cells

Background Mature adipocytes, myocytes and osteoblasts are derived from a common mesenchymal precursor, the mesenchymal stem cell (MSC). The processes of adipogenesis, myogenesis and osteogenesis are closely related to each other. Interestingly, regulators of differentiation along one lineage are often found to function in the differentiation along another, some of which are seen as regulatory switches for these differentiation pathways. Work in unipotent cell lines has shown that members of Kruppel-like factor(KLF) family, KLF2and KLF15act as fine tuners of adipogenesis. It is also found that the two factors are both highly expressed in muscle tissue, while their roles during myogenesis have not been clarified thus far. Moreover, there is a large gap in our knowledge about the expression and function of the KLF family during osteogenesis. Analysis of KLF2and KLF15expression profiles during adipogenic, myogenic, and osteogenic differentiation from pluripotent MSCs will lay the basis for further investigations into the signal pathway involved in muscle development and bone formation and their relations with fat storage.Objectives The purpose of this research is to find out whether there is any alteration of transcription factor KLF2and KLF15, during the process of induced adipogenic, myogenic, and osteogenic differentiation from human bone marrow mesenchymal stem cells (hBMSCs) in vitro, and to discuss the possible roles of the two factors in these processes.Methods Cells were isolated by density gradient centrifugation from human bone marrow, and treated with established lineage-specific agents in vitro. Meanwhile, the expression of KLF2, KLF15and several differentiation marker genes in cells were detected by quantitative RT-PCR and immunofluorescent staining.Results The hBMSCs gained the ability to undergo lineage commitment after exposure to adipogenic, myogenic, and osteogenic stimuli, and stained positive for Oil red O, Alizarin red S and myosin immunofluorescence, respectively.Both KLF2and KLF15showed time-course expression during adipogenesis and myogenesis. KLF2expression decreased while KLF15increased during these processes. KLF15expression was also upregulated during osteogenesis. KLF2expression did not show significant change during osteogenesis.Glut4expression level was upregulated during adipogenesis as well as myogenesis. PPARγ, MyoD and Runx2expression increased after adipogenic, myogenic, and osteogenic induction respectively.Conclusion KLF2and KLF15expression profiles during adipogenic differentiation of hBMSCs were in accordance with that of3T3-L1preadipocytes as previously reported. The significant change of KLF2and KLF15expression during adipogenesis and myogenesis suggest that they may not only function in adipogenesis, but also in myogenesis. The induction treatment seems to have the opposite effect on KLF2and KLF15expression, which suggest the two factors possibly act in a competitive way by binding to the same target sites. The upregulation of KLF15after osteogenic induction suggest KLF15might be related to the initiation and maintenance of osteogenesis.