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转染人血红素加氧酶-1基因对大鼠血管平滑肌细胞抗氧化损伤的作用 |
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张敏1,
安威2, 杜海军2, 陈莉2, 张宝慧1
(1北京大学第三医院运动医学研究所, 北京 100083;
2首都医科大学细胞生物系, 北京 100054) |
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摘 要: |
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本实验构建含人血红素加氧酶-1(hHO-1)基因的逆转录病毒载体XM-6/hHO-1,
将其导入离体培养的大鼠血管平滑肌细胞(vascular smooth muscle cells, VSMC), 观察外源性hHO-1基因在VSMC内的表达及其抗活性氧损伤作用。结果表明:(1)
hHO-1基因可在靶细胞中明显表达, 转染VSMC的HO-1蛋白表达和HO酶活性分别比非转染细胞高1.8倍和2.0倍; (2)转染hHO-1的VSMC可对抗大剂量H2O2对细胞的损伤作用,
表现为细胞存活率增加和乳酸脱氢酶(LDH)漏出减少，上述保护作用可被HO特异性抑制剂锌原卟啉IX (Zinc-protoporphyrin IX，
ZnPP-IX)所阻断。研究结果提示, 外源性HO-1的过量表达可增加VSMC对抗氧化损伤的能力。
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关键词:
血红素加氧酶; 逆转录病毒载体; 活性氧; 血管平滑肌
学科分类号: Q78;
R363 |
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Increased resistance against
oxidant-induced injury in the rat vascular smooth muscle cells transfected
with human
heme oxygenase-1 gene
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ZHANG Min1, AN Wei2, DU Hai-Jun2, CHEN Li2, ZHANG
Bao-Hui1
(1Institute of Sports Medicine, The Third
Hospital, Peking University, Beijing 100083; 2Department of Cell Biology,
Capital University of Medical Sciences, Beijing 100054) |
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Abstract:
The heme oxygenase-1 (HO-1), a rate-limiting enzyme in heme metabolism,
has been recently defined as a novel stress-stimulated protein, since the
intracellular expression of HO-1 in response to various stimuli as
oxidation, ischemia and endotoxin injury has been proved to be able to
protect the cells from damage. In this study, a retroviral vector
containing human HO-1 gene was constructed and transfected to rat vascular
smooth muscle cells (VSMCs). Using Southern and Northern blot analyses,
the integration and mRNA expression of HO-1 gene in the transfected cells
were confirmed. The profound protein expression of HO-1 as well as HO
enzyme activity in the transfected cells increased by 1.8-fold and
2.0-fold respectively as compared with the non-transfected cells. It was
found that the HO-1 transfected-VSMCs presented dominant resistance to
toxicity produced by exposure to H2O2, as a significant protective effect
of HO-1 marked by cell survival and LDH leakage was observed when 200, 400
and 600 μmol/L of H2O2 were used. The protection of HO-1 rapidly declined
after the transfected-VSMCs were pretreated 24 h with an HO-1 specific
inhibitor (ZnPP-IX). The results of this investigation suggest that the
functional expression of HO-1 gene within VSMCs raises an alternative
ability to protect the vascular cells against active oxygen injury.
Key words:
heme oxygenase; retroviral vector; active
oxygen; vascular smooth muscle cells |
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