激素耐药肾病伴泌尿生殖器异常患儿的临床及WT1基因检测

目的:了解WT1基因突变在3例激素耐药肾病伴泌尿生殖器异常患儿的临床特点和致病作用.方法:采用间接免疫荧光及免疫组织化学的方法对2例患儿的肾组织行足细胞分子(nephrin,podocin,α-actinin4,WT1及CD2AP)表达;采用PCR及RT-PCR的方法检测WT1基因突变及 KTS/-KTS比例.结果:3例患儿发病年龄分别为6月、1岁及10岁,确诊年龄分别为7月、9岁及15岁.例1和例3为男性表型伴泌尿外生殖器异常,例2为女性表型;3例患儿染色体核型均为46,XY.临床均为激素耐药肾病,例1和例2肾脏病理为局灶节段性肾小球硬化(FSGS).对2例肾活检患儿肾组织标本的分析显示足细胞分子表达均发生改变;例1 WT1无表达,例2 WT1在足细胞核内的分布与正常对照不同.WT1基因分析示,WT1基因序列中例1未发现突变,例2为IVS 9 5 G＞A杂合突变,例3为WT1外显子91186 G＞A的杂合突变.结论:对于早发激素耐药肾病且病理为FSGS的女性患者或伴有泌尿生殖器异常的男性患者应行染色体核型和WT1基因分析.WT1突变引起足细胞分子表达发生改变,提示这些足细胞分子参与了蛋白尿的形成或发展.

Clinical characteristics and WT1 genetic analysis of patients withsteroid resistant nephrotic syndrome accompaniedwith genitourinary malformations

OBJECTIVE: To understand WT1 mutations in patients with steroid resistant nephrotic syndrome (SRNS) accompanied with genitourinary malformations. METHODS: Three cases of SRNS accompanied with genitourinary malformations were enrolled. The expression of podocyte molecules (nephrin, podocin, alpha-actinin-4, WT1, and CD2AP) in 2 cases was analyzed with the immunofluorescence and immunohistochemistry techniques. The genomic DNA and cDNA of WT1 were analyzed by using PCR and RT-PCR, respectively. GeneScan and GeneScan software were used to quantify the ratio of +KTS/-KTS isoforms. RESULTS: The onset ages of 3 cases were 6 months, 1 year, and 10 years old, respectively. The diagnosis age was 7 months, 9 years, and 15 years old, respectively. The phenotype of Case 1 and Case 3 was male accompanied with genitourinary malformations. Case 2 was phenotypic female. Karyotype analysis of 3 cases revealed 46, XY. Three cases were diagnosed as SRNS. Focal segmental glomerulosclerosis (FSGS) was confirmed in 2 cases. Podocyte molecular expression altered in renal tissues of 2 cases. In addition, WT1 staining was negative in Case 1. WT1 expression in Case 2 showed diffuse nuclear staining with less obvious speckles compared with controls. WT1 IVS 9 +5 G>A mutation was detected in Case 2 and WT1 Exon 9 1186 G>A mutation was detected in Case 3. No WT1 mutation was detected in Case 1. CONCLUSION: Karyotype analysis and WT1 genetic testing should be done in all female patients with early onset steroid resistant FSGS and in male patients with SRNS accompanied with genitourinary malformations. Abnormal podocyte molecular expression suggests that more podocyte molecules might be involved in the pathogenesis of proteinuria in WT1 mutational patients.