| 摘要: |
| ［摘要］ 目的 构建糖原合成酶激酶3β（GSK-3β）特异的RNA干扰（RNAi）腺病毒，观察其抑制高游离脂肪酸（FFA）诱导下人脐静脉内皮细胞（HUVEC）GSK-3β的表达情况。方法 利用体外同源重组技术构建GSK-3β特异的siRNA腺病毒表达载体，在HEK293A细胞中包装并扩增病毒、空斑以实验法测定病毒滴度。GSK-3β特异的siRNA腺病毒感染HUVEC，以Western印记法测定其对GSK-3β蛋白表达的影响。结果 成功构建了GSK-3β特异的siRNA腺病毒，获得高滴度的腺病毒液；构建的重组腺病毒感染HUVEC，可显著抑制正常培养及游离脂肪酸诱导下的细胞的GSK-3β蛋白的表达，Ad-DEST组与Ad-640组比较，Ad-640组明显下降（P<0.05）。结论 构建的GSK3β特异的RNAi腺病毒能有效抑制HUVEC GSK-3β蛋白的表达，有可能保护高脂诱导的HUVEC损伤。 |
| 关键词: RNA干扰 糖原合成酶激酶3β 游离脂肪酸 人脐静脉内皮细胞 |
| DOI：10.3969/j.issn.1674-3806.2010.01.01 |
| 分类号:R 587.1 |
| 基金项目:福建省自然科学基金（2007J 0072） |
|
| Effect of GSK-3β-targeting RNAi recombinant adenovirus on the GSK-3β expression of human umbilical vein endothelial cells induced by high free fatty acid |
|
CHEN Yu-fang, CHEN Gang, LIN Li-xiang
|
|
Department of Endocrinology, Fujian Provincial Hospital, Fujian Medical University, Fuzhou 350003, China
|
| Abstract: |
| ［Abstract］ Objective To construct RNAi recombinant adenovirus expressive vectors specific to GSK-3β and explore if it can inhibit the GSK-3β expression of human umbilical vein endothelial cells induced by high free fatty acid.Methods Homologous recombination and cloning techniques were used to construct RNAi recombinant adenovirus expressive vectors specific to GSK-3β. Then, the adenovirus plasmids was transfected into HEK293A cells to produce adenovirus and amplify the adenovirus stock. Plaque forming assay was used to titer the adenovirus stock. The GSK-3βexpression were detected by Western blot.Results The RNAi adenovirus vectors specific to GSK-3β were successfully produced with high titer. The expression of GSK-3β protein could be down-regulated efficiently by the RNAi adenovirus in HUVEC both in the condition of normol and high free fatty acid(P<0.05).Conclusion RNAi adenovirus is an important tool that inhibit the expression of GSK-3β efficiently. It may partly protect HUVEC from impairment which induced by FFAs. |
| Key words: RNA interference Glycogen synthetase kinase-3 beta (GSK-3β) Free fatty acid(FFA) Human umbilical vein endothelial cells(HUVEC) |
