摘要目的　检测AR基因5调控区可能引起启动子功能及蛋白表达的基因改变。方法　应用PCR-SSCP对AR基因5'调控区进行筛选，所有变异体均进行DNA序列分析并克隆至氯霉素乙酰转移酶报告基因载体（pCAT），然后转染Hela细胞，检测CAT活性求出启动子相对活性。同时进行凝胶滞留试验与足纹分析以确定DNA与蛋白质的相互作用。结果　在145名2型糖尿病患者及123名正常对照的醛糖还原酶基因5'调控区除野生型外，发现两种多态性C(-106)T 和 C(-12)G。在正常对照和2型糖尿病患者中，基因型WT/WT, WT/C(-12)G 和 WT/C(-106)T的发生率无显著性差异.在有视网膜病变和无视网膜病变的2型糖尿病患者中, WT/C(-106)T 的发生率分别为 31.5%和17.5% (P<0.05), WT/C(-12)G的发生率分别为 10.5%和2.5%（P>0.05）,在有并发症的患者中, WT/C(-12)G 和 WT/C(-106)T的总发生率为41.8%,明显高于无视网膜病变患者的发生率20.0%(P<0.025),野生型, C(-12)G和C(-106)T 的相对转录活性分别为15.7%, 31.0% 和 32.2%,DNA-蛋白质相互作用实验表明,这些变异未改变DNA与反式作用因子的结合位点。结论　在中国人群中,AR基因5调控区C(-106)T 和 C(-12)G多态与2型糖尿病视网膜病变密切相关。

abstractsObjective To screen the 5' regulatory region of the aldose reductase (AR) gene for genetic variabilities causing changes in protein expression and affecting the promoter function. Methods The screenings were carried out by polymerase chain reaction-single strand conformation polymorphism (PCR-SSCP). All SSCP variants were submitted for DNA sequencing and inserted into the plasmid chloromycetin acetyl transferase (CAT) enhancer vector. The constructs were used to transfect Hela cells,and CAT assays were performed to assess promoter activity. Gel mobility shift and footprinting assays were also performed to determine the interaction between the DNA and nuclear proteins. Results Two polymorphisms, C(-106)T and C(-12)G, were identified in the regulatory region in 123 Chinese control subjects and 145 patients with type 2 diabetes mellitus. The frequencies of genotypes WT/WT, WT/C(-12)G and WT/C(-106)T were not significantly different between the subjects and patients. In the patients with and without retinopathy, frequencies of WT/C(-106)T were 31.5% and 17.5% (P<0.05) respectively, and the frequencies of WT/C(-12)G were 10.5% and 2.5% (P>0.05) respectively. The total frequency of WT/C(-12)G and WT/C(-106)T in patients with retinopathy was 41.8%, significantly higher than that (20.0%) in patients without retinopathy (P<0.025). The relative transcription activities of the wild-type, the C(-12)G and the C(-106)T were 15.7%, 31.0% and 32.2%, respectively. The results of DNA-protein interaction assays showed that these variations did not change the binding site of DNA with trans-acting factors. Conclusion The polymorphisms C(-12)G and C(-106)T strongly associated with diabetic retinopathy in the Chinese population have been identified in the regulatory region of the aldose reductase gene.