先天性巨结肠与其同源病的RET基因突变的研究

Mutations of RET proto-oncogene in Hirschsprung's disease and its allied disorders

目的 了解先天性巨结肠与巨结肠同源病RET基因突变及其突变的差异。方法 30例散发性先天性巨结肠 ,14例巨结肠同源病 ,10例正常对照。取外周血 ,提取DNA ,聚合酶链反应(PCR)扩增RET基因第 13外显子 ;单链构象多态 (SSCP)分析外显子突变 ,并通过测序明确突变的位点和类型。结果 30例散发性先天性巨结肠RET基因 13外显子检测 5例存在基因突变 ,共发现三种突变类型 :有 3例在碱基 18888位点 ,胸苷酸残基T被鸟苷酸残基G置换 ,导致亮氨酸的静默突变 ;有 1例在碱基 18919位点 ,腺苷酸残基A被鸟苷酸残基G置换 ,导致赖氨酸突变为谷氨酸 ,为错义突变 ;有 1例在碱基 18974位点 ,插入一个鸟苷酸残基G ,导致框架移位突变。RET基因 13外显子突变率为 16 .7% (5 /30 )。巨结肠同源病和正常对照组未见RET基因 13外显子突变。结论 先天性巨结肠与RET基因突变有关 ,而巨结肠同源病未发现RET基因突变 ,提示这两种疾病具有分子遗传学差异。

Objective To study the difference of RET proto-oncogene mutation between Hirschsprung's disease (HD) and allied disorders of Hirschsprung's disease (ADHD). Methods Blood samples were obtained from 30 sporadic HD patients, 14 ADHD patients and 10 normal children as control. Genomic DNA was extracted from the blood samples. Exon 13 of RET proto-oncogene were amplified by Polymerase chain reaction (PCR). The mutation was analyzed by single strand conformational polymorphism (SSCP) and DNA sequencing. Results 5 cases of exon 13 RET mutation were detected in the 30 sporadic HD patients. 3 different types of mutation were identified: a transnversion T→G at nucleotide 18888 caused a silent mutation (Leu→Leu) in 3 cases; a transition A→G at nucleotide 18919 caused a missense mutation (Lys→Glu) in 1 caes; a insertion G at nucleotide 18974 caused a frameshift mutation. The Exon 13 of RET proto-oncogene mutation rate in sporadic HD patient was 16.7%(5/30). No mutation was found in ADHD or control cases. Conclusions Mutation of RET proto-oncogene is involved in the etiopathogenesis of HD. Since there was no mutation found in ADHD patients, it indicates that the genetic pathways of HD and ADHD are different.