Abstract
The present study aimed to evaluate the effect of tissue inhibitor of metalloproteinase-1 (TIMP-1) on the proliferation and osteogenic differentiation potential of human bone marrow-derived MSCs (hBMSCs). hBMSCs with stable TIMP-1 overexpression or TIMP-1 knockdown were generated. Osteogenic differentiation was assessed by Alizarin Red S staining, alkaline phosphatase (ALP) activity and expression of specific markers. Compared with the vehicle controls, TIMP-1 knockdown significantly promoted the growth of hBMSCs. TIMP-1 knockdown up-regulated β-catenin and cyclin D1 proteins. During osteogenic differentiation, TIMP-1 knockdown elevated the deposition of calcium nodules, ALP activity and the mRNA levels of the osteogenic markers sex determining region Y-box 9 (Sox9), CCAAT-enhancer-binding protein and peroxisome proliferator-activated receptor γ. During osteogenic differentiation, TIMP-1 knockdown significantly enhanced the up-regulation of osteocalcin proteins. Meanwhile, TIMP-1 overexpression attenuated the Wnt/activator Wnt3a-induced up-regulation cyclin D1 and Runt-related transcription factor 2 (RUNX-2) (during osteogenic differentiation) proteins, while TIMP-1 knockdown restored the inhibitor Dickkopf 1-induced inhibition effect on the expression of β-catenin, cyclin D1 and RUNX-2. TIMP-1 plays a negative regulatory role in the proliferation and osteogenesis of hBMSCs, at least partially, through Wnt/β-catenin signaling.
Highlights
Bone marrow-derived mesenchymal stem cells (BMSCs) are commonly adopted in various cell-based therapy for tissue repair and regeneration [1]
The results demonstrated that the proliferation of LV-Tissue inhibitor of metalloproteinase-1 (TIMP-1) human bone marrow-derived MSC (hBMSC) slightly decreased from day 3 (D3) to D5, while the proliferation of shRNA-tissue inhibitors of matrix metalloproteinases (TIMPs)-1 hBMSCs significantly increased from D3 to D5 as compared with the vehicle control
We investigated the effect of TIMP-1 on the proliferation and osteogenic differentiation potential of hBMSCs by using lentiviral-mediated overexpression and shRNA interference technique
Summary
Bone marrow-derived mesenchymal stem cells (BMSCs) are commonly adopted in various cell-based therapy for tissue repair and regeneration [1]. Previous study has shown that MSCs can move to the fracture sites and differentiate into osteocytes for bone healing [4]. In this respect, understanding the mechanism of osteogenic differentiation of MSCs is helpful for developing the strategy of hMSCs therapy for bone defects or fractures. Matrix metalloproteinases (MMPs) are a family of proteolytic enzymes involved in remodeling of ECM, and their enzyme activity is regulated by binding with tissue inhibitors of matrix metalloproteinases (TIMPs) [6]. Tissue inhibitor of metalloproteinase-1 (TIMP-1) is a multifunctional protein and ubiquitously expresses in a variety of human cells and tissues and participates in a variety of biological processes [7]
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