目的

分析21-羟化酶缺乏症（21-hydroxylase deficiency，21-OHD）患儿和家系基因突变特征及突变基因型与临床表型的关系。

方法

采用病例试验诊断研究设计。收集2011年12月至2013年5月期间就诊于上海交通大学医学院附属上海儿童医学中心的16例21-OHD 患儿及父母外周静脉血，抽提基因组DNA。根据细胞色素P450c21（Cytochrome P450c21，CYP21A2）基因与其假基因间序列的差异，用高特异性引物扩增该基因10个外显子及内含子序列，用PCR扩增产物直接测序法对21-OHD患儿及其家系进行基因分型。

结果

用PCR产物直接测序法检测与分析16例21-OHD患儿的突变基因型，共检出6种CYP21A2基因突变，其中IVS2-13A/C＞G[50%（16/32）]、p.I172N[25%（8/32）]为最常见的2种类型，c.1451_1452delinsC[3%（1/32）]及p. R149P[3%（1/32）]均为罕见突变，除2例患儿未获得父母标本外，其余患儿基因突变均来自父母。失盐型患儿中最常见的突变类型为IVS2-13A/C＞G[75%（15/20个等位基因）]，而单纯型患者中最常见的突变为p.I172N[67%（8/12个等位基因）]，患儿基因型与临床表型间具有很高一致性。

结论

PCR产物直接测序法能较特异地检出21-OHD基因突变位点，并可为疾病确诊及临床表型预测提供可靠依据。

Objective

To analyze the relationship between the genetic mutations and clinical parameters in 21-hydroxylase deficiency (21-OHD) patients and their pedigrees.

Methods

Sixteen 21-OHD patients from Shanghai Children' s Medical Center and their parents were enrolled from December 2011 to May 2013. Genomic DNA was extracted from the peripheral blood sample. All the 10 exons of CYP21A2 gene were amplified by PCR using specific primers and directly sequenced to detect disease-causing mutations.

Results

Analysis of the 16 patients revealed 6 kinds of mutations in CYP21A2 gene. The most frequent mutations of CYP21A2 gene were IVS2-13A/C>G [50%(16/32)] and p.I172N [25%(8/32)], while the c.1451_1452delinsC[3%(1/32)] and p. R149P [3%(1/32)] were rare mutations. All these gene variations detected in the patients were also identified in the parent samples, except for the two cases with incomplete pedigree. The most frequent molecular defect of the salt-wasting form and simple virilizing form was IVS2-13A/C>G [75%(15/20)] and p.I172N [67%(8/12)] respectively. Correlation between genotypes and phenotypes was compatible with the reported data.

Conclusion

The method in this study based on the directly sequencing can identify mutations on CYP21A2 gene with high specificity, which provided a reliable method for molecular diagnosis of 21-OHD.