Effect of Simulated Microgravity Conditions on Differentiation of Adipose Derived Stem Cells towards Fibroblasts Using Connective Tissue Growth Factor.

Abstract

Background: Mesenchymal stem cells (MSCs) are multipotent cells able to differentiating into a variety of mesenchymal tissues including osteoblasts, adipocytes and several other tissues. Objectives: Differentiation of MSCs into fibroblast cells in vitro is an attractive strategy to achieve fibroblast cell and use them for purposes such as regeneration medicine. The goal of this study was investigate the simulated microgravity effect on differentiation of Adipose Derived Stem Cells (ADSCs) to fibroblasts. Materials and Methods: To fibroblast differentiation 100 ng.mL-1 of connective tissue growth factor (CTGF), and for simulation microgravity, 2D clinostat was used. After isolation the human ADSCs from adipose, cells were passaged, and at passages 3 they were used for characterization and subsequent steps. After 7 days of CTGF and simulated microgravity treatment, proliferation, and differentiation were analyzed collectively by MTT assay, quantitative PCR analyses, and Immunocytochemistry staining. Results: MTT assay revealed that CTGF stimulate the proliferation but simulated microgravity didn't have statistically significant effect on cell proliferation. In RNA level the expression of these genes are investigated: collagen type I (COLI), elastin (ELA), collagen type III (ColIII), Matrix Metalloproteinases I(MMP1), Fibronectin 1 (FN1), CD44, Fibroblast Specific protein (FSP-1), Integrin Subunit Beta 1 (ITGB1), Vimentin (VIM) and Fibrillin (FBN). We found that expression of ELN, FN1, FSP1, COL1A1, ITGB1, MMP1 and COL3A1 in both condition, and VIM and FBN1 just in differentiation medium in normal gravity increased. In protein level the expression of COL III and ELN in simulated microgravity increased. Conclusions: These findings collectively demonstrate that the simulated microgravity condition alters the marker fibroblast gene expression in fibroblast differentiation process.