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New Approach for Differentiation of Bone Marrow Mesenchymal Stem Cells Toward Chondrocyte Cells With Overexpression of MicroRNA-140

Mahboudi, Hossein*; Soleimani, Masoud; Hanaee-Ahvaz, Hana; Ghanbarian, Hossein*; Bandehpour, Mojgan*; Enderami, Seyed Ehsan; Kazemi, Bahram§

doi: 10.1097/MAT.0000000000000688
Biomedical Engineering

Mesenchymal stem cells are widely stimulated by transforming growth factor beta-3 (TGFβ3) for chondrocyte differentiation. The objective of our study was to establish a new method for differentiation of human mesenchymal stem cells toward chondrocyte by overexpression of MicroRNA-140 (miR-140), and also this method was compared with method of induction with TGFβ3 in high-cell density culture systems. Mesenchymal stem cells were harvested from bone marrow of human. We prepared vectors and then was used for recombinant Lenti virus production in HEK-293 cell. Transducted cells were cultured in monolayer culture system and were harvested after days 7, 14, and 21. Real time polymerase chain reaction (RT-PCR) was performed to evaluate the cartilage-specific genes in the mRNA levels. Also, in order to confirm our results, we have done immunocytochemistry technique. Bone marrow mesenchymal stem cells (BMSCs) were transducted with recombinant Lenti virus, and miR-140 was expressed. Immunocytochemical method confirmed the differentiation of BMSC toward chondrocyte with handling cartilage matrix genes. Also real-time PCR showed that after expression of miR-140 in transducted BMSCs significantly increased gene expression of collagen type II and aggrecan and downregulated expression of collagen type I when compared with the mRNA levels measured in nontransducted BMSCs. These results were compatible compared with TGFβ3 induction method as control positive. In this study, we described a new approach and technique that may be applied for differentiation of BMSCs to chondrocyte instead of stimulation with TGFβ3. Our data implies that miR-140 is a potent chondrogenic differentiation inducer for BMSCs, and we have shown increasing chondrogenic differentiation by using miR-140 overexpression.

From the *Department of Biotechnology, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran

Hematology Department, Faculty of Medical Sciences, Tarbiat Modares University, Tehran, Iran

Department of Stem Cell Biology, Stem Cell Technology Research Center, Tehran, Iran

§Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran.

Submitted for consideration April 2017; accepted for publication in revised form August 2017.

Disclosure: The authors have no conflicts of interest to report.

Supported by the “School of Advanced Technologies in Medicine of Shahid Beheshti University of Medical Sciences” (through Grant No. 9693).

Supplemental digital content is available for this article. Direct URL citations appear in the printed text, and links to the digital files are provided in the HTML and PDF versions of this article on the journal’s Web site (www.asaiojournal.com).

Correspondence: Bahram Kazemi, Department of Biotechnology, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran. Email: bahram_14@yahoo.com.

Copyright © 2018 by the American Society for Artificial Internal Organs