Synergistic Effects of Hyperosmolarity and Transforming Growth Factor Beta (TGF-β) on Chondrogenic Differentiation and Reducing Neotissue Ossification

Abstract

Background: Mesenchymal stem cells (MSCs), are multipotent stem cells that their differentiation to chondrocytes is considered in recent years. Platelet-rich plasma (PRP), because of rich of several growth factors, and also hyperosmolarity, because of stimulating differentiation culture medium to original medium of chondrocytes, can affect this differentiation. Considering hypertrophy and ossification as resulted adverse factors, this study aimed to achieve an optimal medium for chondrogenic differentiation of human MSCs. Methods: MSCs were extracted from adipose tissue, and differentiated to chondrocyte line. The effect of base medium containing transforming growth factor beta (TGF-β), PRP, hyperosmolarity, base + hyperosmolarity and PRP + hyperosmolarity on chondrogenic differentiation was assessed using special staining methods such as Alcian blue, hematoxylin and eosin (H&E), and collagen type II and X immunocytochemistry, measurement of biochemical indexes including alkaline phosphatase (ALP) activity and deposit of calcium, and study of expression of Col2, Smad2, Sox9, and Aggrecan genes using real-time polymerase chain reaction (RT-PCR). Findings: Human adipose tissue derived MSCs, with differentiation ability to adipocyte and osteocyte lines, were presented characterizations of chondrocytes in all differentiation groups. In base + hyperosmolarity group, ALP activity and deposit of calcium were less than other groups. Conclusion: Base + hyperosmolarity differentiation group can be a better culture medium for chondrogenic differentiation.

Keywords

• Mesenchymal stem cells
• Cell differentiation
• Platelet-Rich Plasma
• Chondrocytes