The Effects Of Simulated Microgravity On The Culture Of Stabilized Fibrin-chondrocyte Constructs

Articular cartilage damage is common in sports injury and osteoarthritis that affect performances and living severely, however, repair and restore the damage always puzzled the traumatology and orthopedics. With the development of tissue engineering, and construct tissue-engineered cartilage in vitro come true, this provides a promising approach for cartilage repair and restoration. It is well know that mechanical environment is important to cartilage growth and function maintenance in vivo, thus seeking optimum culture condition in vitro to enhance tissue-engineered cartilage capability, in some sense, closely resemble the native cartilage, is the hotspot.Objective: The current study using simulated microgravity by Rotating cell culture system (RCCS), explored the effects of simulated microgravity on the culture of stabilized fibrin-chondrocyte constructs. It can give an experimental reference to well repair and restore cartilage damage. Methods: 1. The chondrocytes from articular cartilage of 3-week-old New Zealand rabbit were isolated and monolayer cultured, then collecting within 4 passages chondrocytes as seed cells.2. The modified scaffold was made of fibrin glue with appropriate concentration antifibrinolysin (aprotinin and tranexamic acid). 3. The seed cells were seeded at 1×10~6/ml in the scaffold, which formed stabilized fibrin-chondrocyte constructs. 4. The constructs were divided into two groups, rotating culture (R) and static control (S) group. Two groups were cultured for three weeks, which the R group subjected to simulated microgravity by rotating cell culture system and the S group were cultured in 12-well plate.5.Gross and histological appearance, biochemical content (DNA and GAG), type II collagen immunohistochemical text of both group samples were assessed during three weeks culture. Results: 1. Using modified scaffold, the constructs can basically maintain the original size and shape after three weeks of culture. Histological and immunohistochemical findings revealed chondrocytes propagation, proteoglycan accumulation and type II collagen expression. 2. Compared to the static culture, the simulated microgravity produced more contents of DNA and GAG in rotating group, and these marked differences appeared at week 3(DNA:R:4.02±0.49ug/ml,S:3.28±0.27ug/ml;GAG:R:OD=42.07±3.40,S:OD=37.26 ±1.74,P<0.05), these suggest that constructs can have better quality by simulated microgravity culture. Conclusion: 1. The study showed that degradative speed of modified fibrin gel scaffold were kept up with cartilaginous tissue formed in vitro for a long duration, and it fitted for rotating culture, which maybe the suitable scaffold for cartilage tissue engineering. 2. The results of this study demonstrated that the simulated microgravity by RCCS can improve the quality of tissue-engineered cartilage formed in vitro. This condition simulated to the parent tissue condition in vivo, thus cells can form big aggregates. With the optimization of the culture conditions, RCCS may be employed for the production of tissue-engineered cartilage and become a promising means for human cartilage reconstruction.