Hsu Han-Hsiu, Uemura Toshimasa, Yamaguchi Isamu, Ikoma Toshiyuki, Tanaka Junzo
Department of Life Science and Biotechnology, Biotechnology Research Institute for Drug Discovery, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba Central-5, Ibaraki 305-8562, Japan.
Department of Life Science and Biotechnology, Biotechnology Research Institute for Drug Discovery, National Institute of Advanced Industrial Science and Technology, 1-1-1 Higashi, Tsukuba Central-5, Ibaraki 305-8562, Japan.
J Biosci Bioeng. 2016 Aug;122(2):219-25. doi: 10.1016/j.jbiosc.2016.01.001. Epub 2016 Jan 30.
Fish collagen has recently been reported to be a novel biomaterial for cell and tissue culture as an alternative to conventional mammalian collagens such as bovine and porcine collagens. Fish collagen could overcome the risk of zoonosis, such as from bovine spongiform encephalopathy. Among fish collagens, tilapia collagen, the denaturing temperature of which is near 37°C, is appropriate for cell and tissue culture. In this study, we investigated chondrogenic differentiation of human mesenchymal stem cells (hMSCs) cultured on tilapia scale collagen fibrils compared with porcine collagen and non-coated dishes. The collagen fibrils were observed using a scanning electronic microscope. Safranin O staining, glycosaminoglycans (GAG) expression, and real-time PCR were examined to evaluate chondrogenesis of hMSCs on each type of collagen fibril. The results showed that hMSCs cultured on tilapia scale collagen showed stronger Safranin O staining and higher GAG expression at day 6. Results of real-time PCR indicated that hMSCs cultured on tilapia collagen showed earlier SOX9 expression on day 4 and higher AGGRECAN and COLLAGEN II expression on day 6 compared with on porcine collagen and non-coated dishes. Furthermore, low mRNA levels of bone gamma-carboxyglutamate, a specific marker of osteogenesis, showed that tilapia collagen fibrils specifically enhanced chondrogenic differentiation of hMSCs in chondrogenic medium, as well as porcine collagen. Accordingly, tilapia scale collagen may provide an appropriate collagen source for hMSC chondrogenesis in vitro.
最近有报道称，鱼胶原蛋白是一种用于细胞和组织培养的新型生物材料，可替代牛和猪等传统哺乳动物胶原蛋白。鱼胶原蛋白可以克服人畜共患病的风险，比如牛海绵状脑病。在鱼胶原蛋白中，罗非鱼胶原蛋白的变性温度接近37°C，适用于细胞和组织培养。在本研究中，我们研究了在罗非鱼鱼鳞胶原蛋白原纤维上培养的人间充质干细胞（hMSC）的软骨形成分化，并与猪胶原蛋白和未包被的培养皿进行了比较。使用扫描电子显微镜观察胶原蛋白原纤维。通过番红O染色、糖胺聚糖（GAG）表达和实时PCR检测来评估hMSC在每种类型胶原蛋白原纤维上的软骨形成。结果显示，在罗非鱼鱼鳞胶原蛋白上培养的hMSC在第6天显示出更强的番红O染色和更高的GAG表达。实时PCR结果表明，与在猪胶原蛋白和未包被的培养皿上培养的hMSC相比，在罗非鱼胶原蛋白上培养的hMSC在第4天SOX9表达更早，在第6天聚集蛋白聚糖和胶原蛋白II表达更高。此外，骨γ-羧基谷氨酸（一种成骨特异性标志物）的低mRNA水平表明，罗非鱼胶原蛋白原纤维与猪胶原蛋白一样，在软骨形成培养基中特异性增强了hMSC的软骨形成分化。因此，罗非鱼鱼鳞胶原蛋白可能为体外hMSC软骨形成提供合适的胶原蛋白来源。
