Effects of clinorotation on COL1A1- EGFP gene expression.
Bone-formation related gene plays a critical role in bone loss induced by space microgravity, however the exact mechanism is unclear. In this study, we aim to investigate the effect of microgravity on the activity of alpha 1(I) collagen (COL1A1) gene promoter and the expression of osteoblast-related genes. COL1A1 promoter was digested by restriction enzymes resulting in three DNA fragments. The fragments were ligated with the enhanced green fluorescent protein report gene, and subcloned into expression vectors. ROS17/2.8 cells transfected by these vectors were screened by G418, and enhanced green fluorescent protein (EGFP) positive colonies were isolated and cultured under clinostat condition. EGFP and Collagen type I expression level were detected by fluorescence intensity analysis and immunocytochemistry methods respectively. The results showed that the expression of EGFP and collagen type I was increased 24 h, 48 h after the cells were cultured under stimulated microgravity, illustrating that the activity of COL1A1 promoter might be increased. In conclusion, osteoblasts can compensatively increase the expression of type I collagen by enhancing the activity of COL1A1 promoter under short-term simulated microgravity conditions.
Dai
Zhongquan
Z
Laboratory of Space Cellular and Molecular Biology, Institute of Space Medico-Engineering, Beijing 100094, China.
Li
Yinghui
Y
Ding
Bai
B
Zhang
Yuguo
Y
Liu
Weiquan
W
Liu
Pengpeng
P
D000818:Animals; D045744:Cell Line, Tumor; D024042:Collagen Type I / Q000235:genetics*; D000090602:Collagen Type I, alpha 1 Chain; D005453:Fluorescence; D005786:Gene Expression Regulation; D005822:Genetic Vectors / Q000235:genetics; D049452:Green Fluorescent Proteins / Q000235:genetics*; D007150:Immunohistochemistry; D051381:Rats; D017207:Rats, Sprague-Dawley; D014162:Transfection; D018474:Weightlessness Simulation
D024042:Collagen Type I; D000090602:Collagen Type I, alpha 1 Chain; C485184:enhanced green fluorescent protein; D049452:Green Fluorescent Proteins