中国组织工程研究 ›› 2017, Vol. 21 ›› Issue (1): 115-121.doi: 10.3969/j.issn.2095-4344.2017.01.021

• 干细胞培养与分化 stem cell culture and differentiation • 上一篇    下一篇

重组腺病毒介导脂联素对人脐静脉内皮细胞损伤的作用和机制

王雪梅,魏  琴,段明军,张  春,杨毅宁   

  1. 新疆医学动物模型研究重点实验室,新疆医科大学第一附属医院临床医学研究院,新疆维吾尔自治区乌鲁木齐市  830011
  • 出版日期:2017-01-08 发布日期:2017-03-15
  • 通讯作者: 杨毅宁,博士,主任医师,教授,博士生导师,新疆医学动物模型研究重点实验室,新疆维吾尔自治区乌鲁木齐市 830011
  • 作者简介:王雪梅,女,1983年生,新疆维吾尔自治区库尔勒市人,回族,2015年新疆医科大学劳动卫生与环境卫生专业毕业,博士,副研究员,主要从事心血管疾病动物模型制作和发病机制研究。
  • 基金资助:

    新疆维吾尔自治区自然科学基金青年基金项目(2015211C095);新疆重大疾病医学重点实验室-省部共建国家重点实验室培育基地开放课题(SKLIB-XJMDR-2014-16)

Effects of recombinant adenovirus-mediated adiponectin on human umbilical vein endothelial cell injury and the underlying mechanism

Wang Xue-mei, Wei Qin, Duan Ming-jun, Zhang Chun, Yang Yi-ning   

  1. Xinjiang Key Laboratory of Medical Animal Model Research, Clinical Medical Research Institute of First Affiliated Hospital of Xinjiang Medical University, Urumqi 830011, Xinjiang Uygur Autonomous Region, China
  • Online:2017-01-08 Published:2017-03-15
  • Contact: Yang Yi-ning, M.D., Professor, Chief physician, Doctoral supervisor, Xinjiang Key Laboratory of Medical Animal Model Research, Clinical Medical Research Institute of First Affiliated Hospital of Xinjiang Medical University, Urumqi 830011, Xinjiang Uygur Autonomous Region, China
  • About author:Wang Xue-mei, M.D., Associate researcher, Xinjiang Key Laboratory of Medical Animal Model Research, Clinical Medical Research Institute of First Affiliated Hospital of Xinjiang Medical University, Urumqi 830011, Xinjiang Uygur Autonomous Region, China
  • Supported by:

    the Natural Science Foundation for the Youth of Xinjiang Uygur Autonomous Region, No. 2015211C095; the State Key Lab Incubation Base of Xinjiang Major Diseases Research, No. SKLIB-XJMDR-2014-16

摘要:

文章快速阅读:

文题释义:
基因转染:
是“将具生物功能的核酸转移或运送到细胞内并使核酸在细胞内维持其生物功能”。其中,核酸包括DNA(质粒和线性双链DNA),反义寡核苷酸及RNAi(RNA interference)。基因转染技术已广泛应用于基因组功能研究(基因表达调控,基因功能,信号转导和药物筛选研究)和基因治疗研究。
感染复数(multiplicity of infection,MOI):传统的感染复数概念起源于噬菌体感染细菌的研究。噬菌体的数量单位为pfu(plaque forming unit,空斑形成单位)。一般认为感染复数是一个比值,没有单位,其实其隐含的单位是pfu number/cell。后来感染复数被普遍用于病毒感染细胞的研究中,含义是感染时病毒与细胞数量的比值。

 

摘要
背景:
血管内皮细胞紧贴于血管壁,在血管稳态平衡的调节中发挥着至关重要的作用。
目的:分析腺病毒介导的脂联素过表达对氧化型低密度脂蛋白(ox-LDL)诱导的人脐静脉内皮细胞动脉粥样化损伤、核因子κB p65核蛋白及其信号通路下游促炎因子的负性调控作用。
方法:实验分4组:正常对照组,采用人脐静脉内皮细胞常规培养,未用ox-LDL诱导;ox-LDL诱导组,用30 mg/L的ox-LDL诱导人脐静脉内皮细胞48 h;二硫代氨基甲酸吡咯烷(PDTC)干预+ox-LDL诱导组,先用100 µmol/L的PDTC处理细胞1 h,再用ox-LDL 30 mg/L诱导48 h;Ad-APN-eGFP干预+ox-LDL诱导组,以MOI=100 pfu/cell转染人脐静脉内皮细胞2 h后,用ox-LDL 30 mg/L诱导48 h。各组干预后检测各项指标。
结果与结论:①ox-LDL诱导人脐静脉内皮细胞造成内皮细胞损伤模型,激活核因子κB信号通路,核因子κB p65核蛋白的表达量增加;②rAd-APN-eGFP转染能够抑制核因子κB通路的激活,使核因子κB p65核蛋白表达量降低,降低核因子κB调节的下游促炎分子VCAM-1、ICAM-1、MCP-1的基因和蛋白表达,增加抗炎因子内皮型一氧化氮合酶的基因和蛋白表达,差异有显著性表达;③结果说明,rAd-APN-eGFP载体转染人脐静脉内皮细胞,通过有效抑制ox-LDL诱导人脐静脉内皮细胞的核因子κB激活,减轻ox-LDL诱导的细胞促炎损伤和细胞凋亡,与使用PDTC干预具有相近的效果;④结果证实rAd-APN-eGFP和PDTC一样可通过抑制核因子κB通路的激活来减轻动脉粥样硬化这种炎症反应。

 

中国组织工程研究杂志出版内容重点:干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程
ORCID:
0000-0002-6540-4546(王雪梅)

关键词: 组织构建, 内皮细胞, 脂联素, 动脉粥样硬化, 人脐静脉内皮细胞, 氧化型低密度脂蛋白, NF-κB信号通路, 新疆维吾尔自治区自然科学基金

Abstract:

BACKGROUND: Vascular endothelial cells closely adhere to the blood vessel wall so as to play a crucial role in the regulation of vascular homeostasis.
OBJECTIVE: To analyze the inhibitory effects of recombinant adenovirus-mediated adiponectin overexpression on oxidized low density lipoprotein (ox-LDL)-induced atherosclerotic injury in human umbilical vein endothelial cells, nuclear factor κB (NF-κB) p65 nuclear protein and pro-inflammatory factors in the downstream of signaling pathway.
METHODS: There were four groups in this experiment: human umbilical vein endothelial cells were cultured in the conventional medium without ox-LDL induction as control group; human umbilical vein endothelial cells induced by 30 mg/L ox-LDL for 48 hours as ox-LDL induction group; cells firstly treated with 100 µmol/L pyrrolidine dithiocarbamate (PDTC) for 1 hour, and then induced by 30 mg/L ox-LDL for 48 hours as PDTC with ox-LDL induction group; cells transfected by adenovirus bearing a vector encoding for adiponectin and enhanced green fluorescent protein (Ad-APN-eGFP) (multiplicity of infection=100 pfu/cell) for 2 hours, and then induced by 30 mg/L ox-LDL for 48 hours as Ad-APN-eGFP with ox-LDL induction group. Afterwards, all relative indicators were detected in each group.
RESULTS AND CONCLUSION: Endothelial cell injury models were obtained by ox-LDL induction, and the NF-κB signaling pathway was activated with the increased expression of NF-κB p65 nuclear protein. Ad-APN-eGFP could inhibit the NF-κB signaling pathway, which significantly decreased the expression of NF-κB p65 nuclear protein, as well as the gene and protein expressions of VCAM-1, ICAM-1 and MCP-1 in the downstream of signaling pathway, but significantly increased the gene and protein expressions of endothelial nitric oxide synthase. These results show that Ad-APN-eGFP-tranfected human umbilical vein endothelial cells can relieve ox-LDL-induced pro-inflammatory injury and reduced apoptosis via effectively inhibiting the NF-κB activation, which achieves similar outcomes with the PDTC intervention. In other words, Ad-APN-eGFP and PDTC both can reduce inflammatory reactions such as atherosclerotic injury by inhibiting the NF-κB signaling pathway.

 

中国组织工程研究杂志出版内容重点:干细胞;骨髓干细胞;造血干细胞;脂肪干细胞;肿瘤干细胞;胚胎干细胞;脐带脐血干细胞;干细胞诱导;干细胞分化;组织工程

Key words: Atherosclerosis, Endothelial Cells, Lipoproteins, LDL, Tissue Engineering

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