回转对离体大鼠成骨细胞中骨粘连蛋白及骨桥素mRNA的影响

Effects of rotation on osteonectin and osteopontin mRNA level of cultured osteoblasts in rats

为研究模拟失重对成骨细胞细胞外基质mRNA的影响,实验采用离体大鼠成骨细胞水平轴回转模拟失重效应,用RT-PCR技术分别检测成骨细胞中骨桥素(osteopontin,OPN)及骨粘连蛋白(osteonectin,ON)mRNA的水平,并观察细胞培养液中碱性磷酸酶(alkalinephosphatase,ALP)的活性和骨钙素(osteocalcin,BGP)含量的变化。结果观察到,分别回转24、48、72h后,OPN、ON的mRNA含量及细胞培养液中BGP含量均显著下降,细胞培养液中ALP活性也呈下降趋势。上述结果表明,模拟失重后成骨细胞OPN及ON的表达下调,进而使BGP及ALP的分泌量减少,从而导致骨钙化能力降低,提示模拟失重导致的细胞外基质蛋白基因表达下降可能是模拟失重引起骨丢失的原因之一。

Conditions of disuse such as bed rest, space flight, and immobilization result in decreased mechanical loading of bone, whichis associated with reduced bone mineral density and increased fracture risk. Mechanisms involved in this process are not wellunderstood except the suppression of osteoblast function. To investigate the effect of simulated weightlessness on mRNA level ofextracellular matrix proteins, osteoblasts were rotated in horizontal plane as a model of simulated microgravity. Primordial osteoblastsof rats were grown for 2 d and then rotated for 24, 48 and 72 h, respectively. After isolating total RNA in cells, reverse transcriptionpolymerase chain reaction (PT-PCR) was made to examine the mRNA level of osteopontin (OPN) and osteonectin (ON). Meanwhile,the levels of alkaline phosphatase (ALP) and osteocalcin (BGP) in the cultured medium were measured to evaluate the calcific functionof cell. The expression of OPN and ON mRNA fell significantly after rotating for 24, 48 and 72 h, respectively. The contents of BGPdescended significantly, meanwhile, the activity of ALP also showed a degressive tendency. Horizontal rotation decreased the expressionof ON and OPN as well as diminished the secretion of BGP and ALP, which affected the calcific function of osteoblast. The resultsobtained suggest that depression of extracellular matrix proteins expression plays a key role in bone loss during weightlessness.