| 摘要: | 葛瑞夫玆氏症（Graves’ disease）是自體免疫甲狀腺疾病，在臨床上病人會有甲狀腺機能亢進（hyperthyroidism）、瀰漫性甲狀腺腫（diffuse goiter）、甲狀腺凸眼症（ophthalmopathy）及脛前黏液水腫（pretibial myxedema）。流行病學研究顯示，環境和基因因素在葛瑞夫玆氏症發生過程佔重要地位。因為每個人體內基因的多型性（polymorphisms），導致不同個體對葛瑞夫玆氏症的感受性也不同。此外，不同個體對於物質所誘發的免疫反應強弱不同，導致個體罹患葛瑞夫玆氏症的危險性也不同。
目前，所被探討的候選基因（candidate gene）－毒殺性T淋巴細胞關聯抗原-4（cytotoxic T lymphocyte-associated antigen-4，CTLA-4）－，參與T細胞活化調節的B7/CTLA-4/CD28訊息路徑。其所表現的蛋白質在整個T細胞所引起的免疫反應中，扮演一個負調節（down regulation）者的角色以及維持體內的免疫平衡。它會阻斷CD28和B7所產生的協同刺激訊息（co-stimulation signal），使整個免疫反應進入休眠的階段。我們所要探討的是位在CTLA-4基因上的兩個單一核苷酸之多型性變異（single nucleotide polymorphisms，SNPs），分別位在表現序列 1 +49（exon 1 +49）SNP以及3’ 非編碼區（3’ noncoding region）的CT60 SNP。在本篇論文中，利用族群病體－對照研究（population-based case-control study），來探討在CTLA-4基因上exon 1 +49 SNP以及CT60 SNP與台灣族群葛瑞夫玆氏症之間的基因關聯性，以及兩個SNPs之間的連鎖不平衡關係。
我們共收集了107個台灣的葛瑞夫玆氏病人以及101個健康對照組的DNA檢體。利用聚合酶鏈鎖反應－限制內切酶片段長度多型性（polymerase chain reaction－restriction fragment length polymorphism assay，PCR-RFLP）分析exon 1 +49 SNP及CT60 SNP的A/G核苷酸變異。另外，利用化學冷光（chemiluminescent）免疫分析儀器－IMMULITE 2000，來分析病人血清中的甲狀腺素（thyroxine，T4）及三碘甲腺素（3,5,3,’-triiodothyronine，T3），以及甲狀腺刺激激素（thyroid stimulating hormone，TSH）的濃度。
我們發現病人及對照組在exon 1 +49 SNP以及CT60 SNP中的基因型（genotype）以及對偶基因（allele）的分佈有顯著差異（p < 0.05）。但是，病人及對照組在表現型（phenotype）的分佈並無顯著差異。病人在exon 1 +49 SNP及CT60 SNP基因型同時為G/G所帶的比例明顯比對照組多（p = 0.022）。然而，exon 1 +49 SNP及CT60 SNP基因型和病人臨床表徵（發病年齡，血清中T3、T4、TSH）之間並無相關聯性。
此篇論文首次發現：CTLA-4基因上的exon 1 +49 SNP 及CT60 SNP與台灣族群葛瑞夫茲氏症的感受性相關，但是其真正的分子病理機轉仍有待研究。
The objective of this study was to examine the polymorphisms in the cytotoxic T lymphocyte-associated antigen-4 (CTLA-4) gene and its relationship with Graves’ disease in the Taiwanese population.
Graves' disease (GD) is an autoimmune disease caused by a combination of environmental and genetic factors. One of the candidate genes is CTLA-4, which acts as fundamental regulator of T cell homeostasis and can limit the immune response by blocking CD28-B7 interaction. Two A/G polymorphic sites of the CTLA-4 gene have been described, one at exon 1 +49 and the other at the 3’ noncoding region－CT60. In the present study, a population-based case-control study was employed to investigate the genetic association, or linkage disequilibrium, between CTLA-4 alleles and GD patients in the Taiwanese population.
We studied DNA samples from 107 GD patients and 101 control subjects. The exon 1 +49 and CT60 A/G polymorphisms were typed using a polymerase chain reaction－restriction fragment length polymorphism assay (PCR-RFLP). The serum concentrations of 3,5,3,’-triiodothyronine (T3), thyroxine (T4) and thyroid stimulating hormone (TSH) were analyzed using chemiluminescent immunoassay by IMMULITE 2000.
There were significant differences in genotype frequencies and allele frequencies (p < 0.05) in two polymrophsim sites between patients with GD patients and controls. And there was also a significant difference in phenotype frequence in CT60 SNP but no difference in phenotype frequencie in exon 1 +49 between patients with GD patients and controls. Furthermore, we found that both G/G genotypes in exon 1 + 49 SNP and CT60 SNP occurred with much higher frequence in patients than in control subjects (p = 0.022). Comparing with patinets, there was much higher frequence occurring both A/G genotypes in exon 1 +49 SNP and CT60 SNP in control subjects.( p = 0.015) Significant linkage disequilibrium was found between exon 1 +49 SNP and CT60 SNP in both GD patients and controls (D’ = 1.00). Because of the tight linkage disequilibrium, a combination of these polymorphisms enhances the role of CTLA-4 in GD. To investigate the possible pathological phenotypes corresponding the genotype changes, we focused on the T3, T4, TSH and age of onset. We found there were no association between the mean values of T3, T4, TSH and age of on set and CTLA-4 genotypes in patients.
We provide, for the first time, supporting evidence that CTLA-4 polymorphisms are associated with susceptibility to GD in the Taiwanese population. However, the exact molecular mechanism remains unclear. |
