6q24新生儿暂时性糖尿病的临床特点及分子遗传学研究

Clinical features and molecular genetic analysis of 6q24-related transient neonatal diabetes mellitus

目的探讨6q24新生儿暂时性糖尿病(6q24-related transient neonatal diabetes mellitus,6q24-TNDM)的临床特征和分子遗传学特点。方法回顾性分析2017年上海市儿童医院新生儿科收治的2例6q24-TNDM患儿的临床资料。采用焦磷酸测序方法,对6q24区域内甲基化差异修饰区域(differentially methylated region,DMR)中16个胞嘧啶-磷酸-鸟嘌呤二核苷酸(cytidine-phosphate-guanosine,CpG)位点的甲基化水平进行定量分析。结果2例患儿均为男性,例1为5日龄,例2为11日龄,均为剖宫产娩出。例1胎龄37周+4,出生体重2340 g;例2胎龄38周+2,出生体重2600 g。2例均为小于胎龄儿,生后均因高血糖入院,体格检查见皮肤弹性略差,皮下脂肪偏少,入院时血糖分别为12.95和8.00 mmol/L,血胰岛素低于正常值(分别为<1.39和3.94 pmol/L),C-肽低于正常值(分别为0.05和0.14 nmol/L),尿糖阳性,尿酮体阴性,血清抗谷氨酸脱羧酶抗体、抗胰岛素抗体、胰岛细胞抗体阴性,胰腺大小正常。皮下注射胰岛素2周余,2例患儿好转出院,分别在生后69和42 d停用胰岛素,糖尿病缓解。2例患儿产前诊断提示染色体核型正常,单核苷酸多态性芯片提示6号染色体单亲二体,入院后二代测序均未检测到明确的致病性点突变。使用焦磷酸测序发现患儿6q24区域DMR中16个CpG位点的甲基化水平均低于10%(健康对照儿童相应位点的甲基化水平约为40%),呈现明显的低甲基化。结论对于出生时小于胎龄儿、高血糖时血胰岛素和C-肽水平低的TNDM患儿,采用焦磷酸测序技术对甲基化差异修饰区域中CpG位点的甲基化水平进行定量分析,可以对6q24-TNDM进行快捷直接诊断,有助于指导治疗和评估预后。

Objective To investigate the clinical and molecular genetic characteristics of 6q24-related transient neonatal diabetes mellitus(6q24-TNDM).Methods The clinical data of two neonates with 6q24-TNDM admitted to Shanghai Children's Hospital,Shanghai Jiao Tong University in 2017,were retrospectively collected.The methylation levels of 16 cytidine-phosphate-guanosine(CpG)sites from the methylated differentially modified region(DMR)in 6q24 were quantitatively analyzed by pyrosequencing.Results Case 1,aged 5 d,was born at 37+4 gestational weeks due to fetal growth restriction,and case 2 was 11-days old and born at 38+2 gestational weeks.Both infants were male and small for age.They were born through a cesarean section.The birth weight of case 1 and case 2 were 2340 g and 2600 g,respectively.They were admitted due to hyperglycemia with blood glucose of 12.95 and 8.00 mmol/L on admission,respectively.Physical examination showed slightly poor skin elasticity and thin subcutaneous fat.Laboratory examination revealed lower serum insulin(<1.39 and 3.94 pmol/L)and peptide C(0.05 and 0.14 nmol/L)levels,positive results of urine glucose,negative tests for urine ketone,serum anti-glutamic acid decarboxylase antibody,anti-insulin antibody,and islet cell antibody in both cases.Normal size of the pancreas was observed by ultrasonography.The infants were improved and were discharged after subcutaneous insulin infusion for more than two weeks.The treatment was discontinued at 69 d and 42 d postnatally for case 1 and case 2.Prenatal diagnosis of the two infants showed normal karyotypes and uniparental disomy of chromosome 6 indicated by single nucleotide polymorphism chip.No pathogenic mutations were detected by next-generation sequencing after admission.The methylation levels of 16 CpG sites in DMR of 6q24 in the two cases,which were quantitatively analyzed by pyrosequencing,were lower than 10%(normal value in healthy matched controls:40%),indicating an obvious hypomethylation.Conclusions For children with TNDM who are small for gestational age at birth,presenting hyperglycemia with decreased serum insulin and C-peptide levels,pyrosequencing can be used to quantitatively analyze the methylation levels of CpG sites in 6q24 DMR,which can quickly and directly assist in the diagnosis of 6q24-TNDM,thereby contributing to the treatment and prognosis assessment.