In vitro differentiation of bone marrow mesenchymal stem cells induced by activin A and all-trans retinoic acid into insulin-producing cells / 中国组织工程研究

ABSTRACT

BACKGROUND: The increasing prevalence of diabetes mellitus has been become one of the diseases which threaten the heath of human being in the 21st century. Islet transplantation is considered to be the most effective approach to cure type Ⅰ diabetes mellitus. However, lack of donor tissue limits the application of this therapy. However, recent progress of stem cell research shows that stem cell therapy may be a potential means to solve this problem.OBJECTIVE: To take activin A and all-trans retinoic acid (AR) in inducing the differentiation of bone marrow mesenchymal stem cells (MSCs) and explore its possibility DESIGN: A randomized controlled experiment.SETTING: Institute of Biotechnology, Academy of Military Medical SciencesMATERIALS This experiment was conducted at the Institute of Biotechnology, Academy of Military Medical Sciences from November 2004to June 2005. Six male Sprague-Dawley rats, with body mass of 150-160g, were provided by the Experimental Animal Center of Academy of Military Medical Sciences.METHODS: Femoral bone marrow of the rats was extracted under aseptic condition. Bone marrow mesenchymal stem cells (MSCs) were isolated with density gradient centrifugation. Passaged MSCs were randomly divided into 4 groups high concentration of glucose (HG), AR, beta-mercaptoethanol (ME) and negative control groups. MSCs were induced to differentiate into IPCs with conditional medium containing high concentration glucose, activin A, RA and ME etc. After induction, phenotypes of differentiated cells were examined by immunocytochemistry and reverse transcription-polymerase chain reaction (RT-PCR).MAIN OUTCOME MEASURES: Expression of insulin and glucagon of differentiated cells were examined by immunocytochemistry. Insulin-1 mR-NA expression of differentiated cells was detected by RT-PCR.RESULTS: After bone marrow mesenchymal stem cells were induced,there were scattered insulin-and glucagon-positive cells in the HG group,many insulin-and glucagon-positive cells in the AR and ME groups, and these cells formed insulin-like structure. The expression of insulin-1mRNA could be observed in the HG, AR and ME groups. Insulin-and glucagonpositive cells and the expression of insulin-1mRNA were not observed in the negative control group.CONCLUSION: We adopt an induction scheme based on AR and other matured factors, and successfully make bone marrow mesench.ymal stem cells induce and differentiate into insulin positive reaction cells and form insulin-like structure, but its induction efficiency needs further improvement.