Microarray Analysis of Gene Expression During Differentiation of Human Mesenchymal Stem Cells Treated with Vitamin E in vitro into Osteoblasts
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연구목적: 비타민 E는 자유 유리기에 의한 골흡수 세포인자의 증가에 대한 보호교과가 있는 것으로 알려져 있다. 이에본 연구에서는 비티만 E가 인간 중간엽 줄기세포의 골모세포분화에 있어 유전자발현에 미치는 영향을 알아보고자 하였다. 연구방법: 인간골수줄기세포를 골분화배지에서 배양하면서 비타민 E를 첨가하였다. 발색면역법을 이용하여 골모세포의분화정도를 측정하였고 마이크로어레이 기법으로 유전자발현의 증감을 확인한 후 real-time RT-PCR로 분석하였다. 결 과: 비타민 E는 골모세포분화과정에 있어 세포의 증식을 증가시켰다. 중간엽줄기세포에서 골분화과정에 관여하는 것으로 알려진 68개 유전자의 분석결과 Alkaline phosphatase, transforming growth factor-beta 1, fibroblast growth factor receptor 1, matrix metalloproteinase 2, muscle segment homeobox 2, bone morphogenetic protein 1, biglycan, vascular endothelial growth factor B, dentin sialophosphoprotein, cartilage oligomeric matrix protein, runt-related transcription factor 2, fibroblast growth factor receptor 3, SMAD2가 2배 이상 유전자발현이 증가되었고, osteopetrosis-associated transmembrane protein 1,microphthalmia-associated transcription factor, epidermal growth factor receptor은 감소하였다. Real-time RT-PCR 분석결과runt-related transcription factor 2와 transforming growth factor-beta 1의 발현이 증가하였다. 결 론: 비타민 E는 인간 중간엽 줄기세포의 골 분화를 증가시키는 작용이 있음을 확인할 수 있었다.
Abstract
Objective: Supplementation with vitamin E is able to protect bone against free radical-induced elevation of bone-resorbing cytokines. We examined gene expression by microarray analysis during the differentiation of human mesenchymal stem cells treated with vitamin E into osteoblasts in vitro. Methods: Human bone marrow stem cells were cultured in osteogenic differentiation medium and vitamin E was added. A colorimetric immunoassay for the quantification of cell proliferation was used to measure osteoblast differentiation. Gene expression was analyzed using a microarray technique. We also used a real time reverse transcription-polymerase chain reaction (RT-PCR). Results: It was found that vitamin E enhanced cell proliferation when compared to cells cultured in media without vitamin E. We focused on 68 genes which are related to osteogenesis and osteoclastogenesis. Alkaline phosphatase, transforming growth factor-beta 1, fibroblast growth factor receptor 1, matrix metalloproteinase 2, muscle segment homeobox 2, bone morphogenetic protein 1, biglycan, vascular endothelial growth factor B, dentin sialophosphoprotein, cartilage oligomeric matrix protein, runt-related transcription factor 2, fibroblast growth factor receptor 3, and SMAD2 were upregulated >2-fold compared to the control. Conversely, osteopetrosis-associated transmembrane protein 1, microphthalmia-associated transcription factor, and epidermal growth factor receptor were downregulated > 2-fold compared to the control. Vitamin E produced a 1.5-fold increase in the expression of runt-related transcription factor 2 and transforming growth factor-beta 1 as determined by real time RT-PCR. Conclusion: Vitamin E had a positive effect on the gene expressions regarding osteogenic differentiation of mesenchymal stem cells. [Korean Journal of Bone Metabolism, 18(1): 23-32, 2011]
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