Department of Biochemistry and Molecular Biology, Mayo Clinic, Rochester, MN, USA.
PLoS One. 2012;7(11):e49083. doi: 10.1371/journal.pone.0049083. Epub 2012 Nov 21.
Fuchs endothelial corneal dystrophy (FECD) is a common, familial disease of the corneal endothelium and is the leading indication for corneal transplantation. Variation in the transcription factor 4 (TCF4) gene has been identified as a major contributor to the disease. We tested for an association between an intronic TGC trinucleotide repeat in TCF4 and FECD by determining repeat length in 66 affected participants with severe FECD and 63 participants with normal corneas in a 3-stage discovery/replication/validation study. PCR primers flanking the TGC repeat were used to amplify leukocyte-derived genomic DNA. Repeat length was determined by direct sequencing, short tandem repeat (STR) assay and Southern blotting. Genomic Southern blots were used to evaluate samples for which only a single allele was identified by STR analysis. Compiling data for 3 arms of the study, a TGC repeat length >50 was present in 79% of FECD cases and in 3% of normal controls cases (p<0.001). Among cases, 52 of 66 (79%) subjects had >50 TGC repeats, 13 (20%) had <40 repeats and 1 (2%) had an intermediate repeat length. In comparison, only 2 of 63 (3%) unaffected control subjects had >50 repeats, 60 (95%) had <40 repeats and 1 (2%) had an intermediate repeat length. The repeat length was greater than 1000 in 4 FECD cases. The sensitivity and specificity of >50 TGC repeats identifying FECD in this patient cohort was 79% and 96%, respectively Expanded TGC repeat was more specific for FECD cases than the previously identified, highly associated, single nucleotide polymorphism, rs613872 (specificity = 79%). The TGC trinucleotide repeat expansion in TCF4 is strongly associated with FECD, and a repeat length >50 is highly specific for the disease This association suggests that trinucleotide expansion may play a pathogenic role in the majority of FECD cases and is a predictor of disease risk.
福克森内皮角膜营养不良（FECD）是一种常见的、家族性的角膜内皮疾病，也是角膜移植的主要指征。转录因子 4（TCF4）基因的变异已被确定为该病的主要原因之一。我们通过在 3 个阶段的发现/复制/验证研究中，在 66 名患有严重 FECD 的患者和 63 名正常角膜的参与者中，确定了 TCF4 内含子 TGC 三核苷酸重复的长度，以检测 TCF4 基因内的 TGC 三核苷酸重复与 FECD 之间的关联。使用侧翼 TGC 重复的 PCR 引物扩增白细胞衍生的基因组 DNA。通过直接测序、短串联重复（STR）分析和 Southern 印迹确定重复长度。基因组 Southern 印迹用于评估仅通过 STR 分析鉴定为单个等位基因的样品。对研究的 3 个部分的数据进行汇总，FECD 病例中有 79%（66 例中的 52 例）存在 TGC 重复长度>50，而正常对照组中只有 3%（63 例中的 2 例）（p<0.001）。在病例中，66 例中有 52 例（79%）患者的 TGC 重复长度>50，13 例（20%）患者的 TGC 重复长度<40，1 例（2%）患者的 TGC 重复长度为中间长度。相比之下，63 例无影响对照受试者中只有 2 例（3%）的 TGC 重复长度>50，60 例（95%）的 TGC 重复长度<40，1 例（2%）的 TGC 重复长度为中间长度。在 4 例 FECD 病例中，重复长度大于 1000。在该患者队列中，TGC 重复长度>50 用于识别 FECD 的灵敏度和特异性分别为 79%和 96%。与先前确定的高度相关的单核苷酸多态性 rs613872 相比，扩展的 TGC 重复对 FECD 病例更具特异性（特异性=79%）。TCF4 中的 TGC 三核苷酸重复扩展与 FECD 强烈相关，TGC 重复长度>50 对该疾病具有高度特异性。这种关联表明三核苷酸扩展可能在大多数 FECD 病例中发挥致病作用，并且是疾病风险的预测因子。
