Effects of transforming growth factor-β1 on the expression of Wnt signaling pathway related genes in the differentiation of bone marrow stromal cells

Abstract

Objective To observe the transforming growth factor-β1 (TGF-β1) on the expression of Wnt signaling pathway related genes during the chondrogenesis of bone marrow stromal cells (BMSCs) in vitro.Methods BMSCs cultured in TGF-β1 medium were observed by inverted phase contrast microscope and identified by toluidine blue staining.The expression levels of glycosaminoglycan (GAG) and target genes including Wnt1,Wnt3a,β-catenin,Sox9 and collagen Ⅱ mRNA under different concentrations of TGF-β1 were detected by using enzyme linked immunosorbent assay (ELISA) and real-time fluorescence quantitative polymerase chain reaction (FQ-PCR) respectively.Results At day 21 after TGF-β1 induction,toluidine blue staining showed that more than 95％ cells differentiated into chondrocytes; the amount of GAG in the cell cultured medium in 2 μg/L TGF-β1-induced group was (69.16 ± 1.18) mg/L,significantly higher than other groups (P ＜ 0.05).FQ-PCR results showed that Wnt1,Wnt3a and β-catenin mRNA expression in 2 μg/L TGF-β1-induced group was inhibited [0.0266 ±0.0047,0.0154 ±0.0033,0.0187 ±0.0021,(P ＜0.05)],but in contrast Sox9 and collagen Ⅱ mRNA expression levels were increased [0.0682 ± 0.0024,0.1134 ± 0.0048 (P ＜ 0.05)] as compared with the control group.Conclusion TGF-β1 could promote BMSCs chondrogenic differentiation,and the expression of Wnt/β-catenin signaling pathway related genes Wnt1,Wnt3a and β-catenin mRNA was inhibited and the expression of chondrocyte specific genes Sox9 and collagen Ⅱ mRNA was increased. Key words: Transforming growth factor-beta1; Bone marrow stromal cells; Wnt signaling pathway; Glycosaminoglycan