大鼠Kruppel样因子14腺病毒载体的构建及其对心肌细胞肥大的影响  

Effects of new constructed rat Kruppel⁃like factor 14 adenovirus vector on cardiomyocyte hypertrophy

作　　者：任志磊[1] 杨崇哲[1] 银孟卓 黄凯华 刘丰[1] REN Zhilei;YANG Chongzhe;YIN Mengzhuo;HUANG Kaihua;LIU Feng(Department of Geriactrics,Guangzhou First People′s Hospital,Guangzhou Medical University,Guangzhou 510180,China)

机构地区：[1]广州医科大学附属广州市第一人民医院老年病科,广州510180

出　　处：《实用医学杂志》2021年第4期441-445,共5页The Journal of Practical Medicine

基　　金：国家自然科学基金项目(编号:81300176);广东省自然科学基金项目(编号:S2011040001711)。

摘　　要：目的检测生理状态和病理性肥大时大鼠乳鼠心肌细胞(NRVM)内Kruppel样因子14(KLF14)的表达水平。构建大鼠KLF14腺病毒载体,感染大鼠NRVM稳定高表达KLF14,探讨其对心肌细胞肥大的影响。方法根据NCBI数据库KLF14的序列合成并构建pHBAd⁃MCMV⁃RFP⁃KLF14重组质粒,PCR和测序鉴定后通过大肠杆菌扩增,在HEK 293细胞中包装成KLF14腺病毒。用KLF14腺病毒或阴性对照红色荧光蛋白(RFP)腺病毒感染NRVM,使用异丙肾上腺素(ISO)刺激NRVM诱导心肌细胞肥大,RT⁃PCR检测BNP和KLF14的表达水平。结果PCR和测序鉴定证实KLF14腺病毒质粒构建成功。RT⁃PCR检测感染KLF14腺病毒可以显著上调KLF14在NRVM中的表达,增加ISO上调的BNP的表达。结论生理情况下,KLF14在心肌细胞中低丰度表达,病理性肥大时表达升高。KLF14过表达促进ISO诱导的心肌细胞肥大。Objective The expression level of KLF14 in the neonatal rat cardiomyocytes(NRVM)of rats was detected under physiological and pathological hypertrophic conditions.To construct rat Kruppel⁃like factor 14(KLF14)adenovirus vector and infect NRVM,and then to investigate its effect on cardiomyocyte hypertrophy.Methods pHBAd⁃MCMV⁃RFP⁃KLF14 recombinant plasmid was synthesized according to NCBI database sequence.The plasmid sequence was verified by PCR and sequencing,and then amplified using bacillus coli.Adenoviral vector was packaged in HEK 293 cell.NRVM were transfected with KLF14 adenovirus or negative control red fluorescent protein(RFP)adenovirus,then stimulated with isoprenaline(ISO)to induce myocytes hypertrophy.Real⁃time PCR(RT⁃PCR)was used to measure the expression of BNP and KLF14 expression.Results The KLF14 adenovirus plasmid was successfully constructed by PCR and sequencing.RT⁃PCR showed that the KLF14 expression in NRVM was significantly increased after KLF14 adenovirus transfection,and overexpression of KLF14 aggravated ISO⁃induced BNP expression.Conclusions KLF14 is expressed at low abundance in cardiomyocytes under physiological conditions,while upregulated in pathological hypertrophy.Over expression of KLF14 promotes ISO⁃induced cardiomyocytes hypertrophy.

关 键 词：KLF14 腺病毒载体 心肌细胞肥大 B型钠尿肽 

分 类 号：R-33[医药卫生]