先天性尿道下裂与SRD5A2及SRY基因突变关系研究

Identification of mutations of SRD5A2 gene and SRY gene in patients with hypospadias

目的 探讨中国人先天性尿道下裂的发生与５α还原酶２（ Ｓ Ｒ Ｄ５ Ａ２） 基因突变及 Ｓ Ｒ Ｙ 基因突变的关系。方法 收集先天性尿道下裂患儿静脉血２３ 例，抽提 Ｄ Ｎ Ａ。经 Ｐ Ｃ Ｒ 分别扩增 Ｓ Ｒ Ｄ５ Ａ２ 基因的第１ 至第５ 外显子及 Ｓ Ｒ Ｙ 基因的 Ｄ Ｎ Ａ 片段。对各扩增片段采用单链构象多态性诊断法（ Ｐ Ｃ Ｒ Ｓ Ｓ Ｃ Ｐ） ，筛选基因突变标本，有电泳异常的标本进行测序分析。结果 经 Ｐ Ｃ Ｒ Ｓ Ｓ Ｃ Ｐ 发现３ 例 Ｓ Ｒ Ｄ５ Ａ２ 基因的第４ 外显子 Ｄ Ｎ Ａ 扩增片段电泳带位置异常，经 Ｄ Ｎ Ａ 序列分析证实２ 例为纯合子型第２２７ 位密码子由 Ｃ Ａ Ａ 替代 Ｃ Ｇ Ａ（ Ａｒｇ２２７ Ｇｌｎ） ；１ 例为双重杂合子型突变，第２２７ 位密码子由 Ｃ Ａ Ａ 替代 Ｃ Ｇ Ａ，同时第１８６ 位密码子由 Ｔ Ｔ Ｇ 替代 Ｔ Ｔ Ｔ（ Ｐｈｅ１８６ Ｌｅｕ） 。 Ｐ Ｃ Ｒ Ｓ Ｓ Ｃ Ｐ 检测 Ｓ Ｒ Ｙ 基因片段未发现异常电泳带。结论 Ｓ Ｒ Ｄ５ Ａ２ 基因突变，可能是先天性尿道下裂的病因之一， Ｓ Ｒ Ｄ５ Ａ２ 基因的第２２７ 位密码子可能为中国人该基因的突变热点，而尿道下裂中 Ｓ Ｒ Ｙ 基因的突变少见。

Objective To identify possible molecular mechanism underlined hypospadias and any relationship of the mutations of SRD5A2 gene and SRY gene to hypospadias.Methods Twenty three blood samples from the patients with hypospadias were obtained from Aug. 1996 to Jan. 1998. DNA was extracted from blood leukocytes. Exons 1 to 5 of the SRD5A2 gene and exon of the SRY gene were amplified by PCR. Mutation detection was performed using PCR SSCP/silver staining and direct DNA sequencing.Results In 3 cases (named 5R2 China 1,5R2 China 2,and 5R2 China 3), DNA sequencing revealed that a homozygous change from nucleotide G to A occurred in 5R2 China 1 and 2, leading to a substitution of glutamine to arginine in the codon 227 (Arg 227 to Gln). In the third patient (5R2 China 3), DNA sequencing revealed two different heterozygous mutations (Arg 227 to Gln, Phe 186 to Leu) in exon 4 of the SRD5A2 gene. No mutation of SRY gene was found in all patients. Conclusion The mutation of SRD5A2 gene affects the differentiation of the external genitalia and may play a role in the etiology of hypospadias. Since no mutation of SRY gene was found, this study might suggest that the mutation of SRY gene is not an important event in hypospadias. Condon 227 is a hotspot site of mutation within the gene. The mutation of codon 186(Phe 186Leu) represents a new form of SRD5A2 mutation.