筛选大鼠血管钙化消退差异表达基因及初步分析

目的：探讨大鼠在血管钙化消退过程中基因表达的变化。方法：选取6周龄SPF级雄性SD大鼠24只,随机分成3组（n=8）,分别为对照组、钙化组和消退组。钙化组和消退组制作血管钙化模型（vitamin D3 plus nieotine,VDN）,对照组生理盐水和花生油灌胃。钙化组和对照组于实验第8周处死,消退组继续饲养至16周处死,测定各组大鼠动脉组织钙含量并作病理检查。采用抑制性消减杂交方法将消退组和钙化组大鼠血管cDNA作差减杂交,分离消退组较钙化组高表达或低表达基因的cDNA片段,建立消退组和钙化组的差异表达文库,扩增、鉴定文库并测序阳性克隆,BLAST比对测序序列。随机挑选4个基因进行RT-PCR验证和DNA条带半定量分析。结果：①血管组织钙含量测定显示钙化组（（15．34±2．51）mg／g）较对照组（（5．20±0．75）mg／g）血管组织钙含量明显升高（P〈0．01）;消退组（（12．73±1．89）mg／g）较钙化组降低（P〈0．05）;②构建了血管钙化的差减文库,对所获阳性克隆进行测序和BLAST比对分析,获得28个表达上调基因和22个表达下调基因。RT-PCR验证示Prdx3,Ank2,Ror2,Abcel等基因在消退组和钙化组间差异表达,消退组较钙化组表达增高,平均约为后者的1．7倍。结论：VDN模型诱导的大鼠血管钙化可以发生主动消退。钙化消退过程中焦磷酸合成相关基因、谷氨酸信号相关基因、还原及凋亡调节基因表达上调,同时见较多骨化相关基因及氧化活性等基因表达下调。钙化抑制基因的表达增多而钙化促进基因表达的下降可能是钙化发生主动消退的内源性机制。

Identification of differentially expressed genes in rats and preliminary analysis in regression of vascular calcification

Objective: To investigate the differentially expressed genes in rat in the process of regression of vascular calcification by using the suppression subtractive hybridization(SSH).Methods: 24 SD male rats which aged 6 weeks and specific pathogen free grade were selec-ted and randomly divided into 3 groups(n=8):control group,calcification group and regression group respectively.Vascular calcification mo-del(vitamin D3 plus nicotine,VDN) were made from rats in calcification group and regression group,and rats ...