醌型二氢生物喋呤还原酶基因表达变化对高糖环境下NRK-52E细胞葡萄糖调节蛋白75表达的影响

Influence of QDPR gene expression on GRP75 in NRK-52E cells under high glucose ambience

目的探讨醌型二氢生物喋呤还原酶(QDPR)基因对高糖环境下肾小管上皮细胞系NRK-52E葡萄糖调节蛋白75(GRP75)的影响及其在DN中的可能作用机制。方法构建过表达及敲低QDPR基因的NRK-52E模型,分别予5.4 mmol/L和30 mmol/L葡萄糖培养基培养,Western blot检测GRP75在OLETF大鼠肾皮质及细胞模型各组的表达水平,流式细胞术检测细胞周期。结果与LETO鼠比较,OLETF大鼠肾皮质GRP75含量降低[(1.53±0.27)vs(0.79±0.26),P<0.01];与NRK-52E组比较,NRK-52E高糖组的GRP75含量降低[(0.62±0.03)vs(0.46±0.06)]、G0/G1期细胞增多[(39.80±1.31)%vs(50.35±0.33)%]、S期细胞减少[(48.55±1.85)%vs(37.17±0.10)%](P<0.05);过表达QDPR使高糖环境下GRP75含量降低[(0.85±0.10)vs(0.45±0.16),P<0.05]、G0/G1期细胞增多[(43.73±0.48)%vs(61.87±0.17)%,P<0.01],S期细胞减少[(42.42±0.66)%vs(25.29±0.12)%,P<0.01];敲低QDPR不影响GRP75表达量与细胞周期。结论 GRP75在DN模型中含量减少,QDPR基因可能通过GRP75及细胞周期影响DN发生发展。

Objective To explore the effect of quinoid dihydropteridine reductase （QDPR） gene expression on glucose-regulated proteins 75 （GRP75） in renal tubular NRK-52E cells under high glucose ambience and its possible underlying mechanism in the development of diabetic nephropathy（DN）. Methods The QDPR overexpression and knockdown models were established in NRK-52E cell lines. All the cells were cultured in 5.4 mmol/L and 30 mmol/L glucose culture medium respectively. GRP75 expression levels in renal cortex of OLETF rats and NRK-52E cells were tested by western blot. Cell eycle profiles were determined by flow cytometric analysis. Results GRP75 protein decreased in renal cortex of OLETF rats [（1. 53±0. 27） vs （0. 79±0. 26） ,P〈O. 01]. GRP75 protein decreased [（0. 62±0. 03） vs （0. 46±0.06）],G0/G1 phase cells increased[（39. 805= 1.31）% vs （50. 35±0. 33）%]and S phase cells decreased [（48. 55±1.85）M vs （37. 17±0.10）%]（P〈0.05） in NRK-52E high glucose group. QDPR overexpression decreased the level of GRP75 protein [（0. 85±0. 10） vs （0. 45±0. 16）,P〈0.05], and S phase cells I-（42. 42±0. 66）M vs （25. 29±0. 12）%,P〈0. 01], increased G0/G1 phase cells[（43. 73±0. 48）% vs （61.87±0. 17）% ,P〈0.01] in high glucose environment. GRP75 expression levels and cell cycle were not effected when QDPR were knockdown. Conclusion GRP75 decreases in DN model. QDPR gene may influence the occurrence and process of DN by regulating the expression of GRP75 and cell cycle.