冠状动脉粥样硬化性心脏病新致病基因MEF2A第7外显子突变的检测

Determination of exon 7 mutation of new virulence gene MEF2A in patients with coronary atherosclerotic heart disease

目的:检测中国冠状动脉粥样硬化性心脏病患者MEF2A基因第7外显子是否存在新突变位点。方法:从1995-03/1996-08在武汉同济医院和武汉协和医院对来自湖北、黑龙江、河南、湖南等省的怀疑冠状动脉粥样硬化性心脏病的患者156例进行临床调查。利用聚合酶链反应-单链构象多态性分析结合变性高效液相色谱分析对所有对象的MEF2A基因第7外显子进行筛查,然后选取变性高效液相色谱分析图谱的峰型出现双或多峰者以及聚合酶链反应-单链构象多态性分析图谱条带数目或位置与对照有差异者的片段进行DNA直接测序,测序结果与正常序列对照确定是否存在突变。结果:参与实验的患者156例全部进入结果分析。①MEF2A基因第7外显子区域有4例患者变性高效液相色谱分析图谱呈双峰或多峰,结果疑为异常。②有7例患者的单链构象多态性分析电泳出现异常条带,即泳动比正常多出条带或移动速度与对照有差异,表明这7例可能出现MEF2A基因突变。③所有异常标本的DNA直接测序结果与正常序列对照未发现第7外显子区域基因突变。结论:冠状动脉粥样硬化性心脏病患者在MEF2A基因第7外显子未发现新的突变,变性高效液相色谱分析和聚合酶链反应-单链构象多态性结果与DNA测序结果并非平行关系,单链构象多态性分析同样存在假阳性。

AIM： To explore the mutation of MEF2A gene exon 7 in Chinese patients with coronary atherosclerotic heart disease （CAHD）. METHODS： Totally 156 suspicious patients, who came from Hubei, Heilongjiang, Henan, Hunan provinces clinically in Wuhan Union Hospital and Wuhan Tongji Hospital from March 1995 to August 1996, were investigated. The exon 7 by denaturing high performance liquid chromatography （HPLC） and polymerase chain reaction-single strand conformation polymorphism （PCR-SSCP） was screened and selected probably abnormal samples that had double or multiple peaks in HPLC or had difference with PCR-SSCP to sequence. Sequencing results would be contrasted with normal sequence to ascertain the existence of mutation. RESULTS： Totally 156 patients were involved in the result analysis. ① Four cases were thought to be abnormal probably by DHPLC showing two or more apices. ②SSCP in exon 7 was thought to be abnormal in 7 eases because of appearance of increaseor decrease of strand or different speed of mobility. It was suggested that mutation might existed in this 7 cases. ③But no mutation was found in the patients by DNA sequence analysis. CONCLUSION： No new mutation in exon 7 of MEF2A is found in Chinese patients with CAHD. Results of DHPLC and PCR-SSCP with result of DNA sequence are not parallel. SSCP also has false positive.