低氧诱导因子-1α在小鼠次级骨化中心形成中的表达

Expression of hypoxia inducible factor-1α in the center of murine developing secondary ossification

目的研究低氧诱导因子-1α(HIF-1α)在小鼠肢体次级骨化中心形成阶段的时空表达规律,并探讨其在长骨次级骨化中心发生和发育过程中的作用。方法解剖镜下整体观察股骨次级骨化中心的发生、发展过程;以地高辛标记的HIF-1αcRNA为探针,采用组织切片原位杂交技术,观察HIF-1αmRNA在次级骨化中心形成阶段的时空表达规律;通过免疫组织化学法观察HIF-1α蛋白在次级骨化中心形成中的表达及分布。结果出生6d,在股骨髁部见到不透光的骨化影,提示次级骨化中心开始出现;其后不透光影在股骨髁内不断膨大。原位杂交显示,出生6d时股骨髁中心出现小片状荧光增强区,出生8d时荧光增强区呈放射样扩大,表示HIF-1αmRNA有大量表达。免疫组织化学法显示,出生6d时股骨髁中心少量HIF-1α蛋白表达,出生8d时次级骨化中心的细胞核内出现大量HIF-1α蛋白表达,之后很快消失。结论HIF-1α在次级骨化中心发育中呈规律性的时空表达,提示其可能参与小鼠长骨次级骨化中心形成的骨发育过程。

Objective To investigate the spatial temporal expression of hypoxia inducible factor-1α （HIF 1α） during the developing murine secondary ossification and appraise the role of HIF-1α in the development of the secondary ossification. Methods The developmental stage of the murine secondary ossification was observed by stereomicroscope, and the expression of HIF-1α at various periods in developing secondary ossification was detected by in situ hybridization （ISH） using Dig-labeled cRNA probe of HIF-1α. The distributional expression of HIF1α protein was also detected by using immunohistochemistry. Results The secondary ossification center appeared 6 days（Pn6） postnatal, and at the same stage HIF-1α mRNA expression was detected by ISH, showing small fluorescent area in the center and radiating from fluorescent area at Pn8, indicating greatly increased HIF-1α mRNA expression. Scanty HIF-1α protein-positive cell was detected at the center of secondary ossification at Pn6, whereas it increased dramatically at PnS. Conclusion The current results demonstrate the expression pattern and the role of HIF-1α in the development of the secondary ossification.