Role of protein kinase C beta2 activation in TNF-alpha-induced human vascular endothelial cell apoptosis.
Role of protein kinase C beta2 activation in TNF-alpha-induced human vascular endothelial cell apoptosis.
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蛋白激酶 C beta2 激活在 TNF-α 诱导的人血管内皮细胞凋亡中的作用。
DOI：
10.1139/y09-004
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发表时间：
2009-03
影响因子：
2.1
通讯作者：
中科院分区：
文献类型：
--
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循环炎性细胞因子肿瘤坏死因子α (tnf - α)在引发或加剧血管内皮损伤的病理条件下增加，如糖尿病。蛋白激酶C （PKC）已被证明在tnf α诱导的人内皮细胞凋亡中起关键作用。然而，PKC的特定亚型在tnf - α诱导的人内皮细胞凋亡中所起的相对作用尚未得到解决。我们研究了一种选择性pkcβ(2)抑制剂（CGP53353）对人血管内皮细胞（细胞系ECV304）中tnf - α诱导的细胞凋亡的影响，以及对活性氧和一氧化氮产生的影响，并将其与rotlerin（一种已被证明可以降低PKCdelta蛋白水平的试剂）的作用进行了比较。培养的人血管内皮细胞（ECV304）用4种治疗方案中的一种治疗24小时：单独使用40 ng/mL tnf - α (tnf - α)、tnf - α与10微mol/L rottlerin （T+rottlerin）、tnf - α与1微mol/L CGP53353 （T+CGP）或不治疗（对照）。MTT法检测细胞活力，流式细胞术检测细胞凋亡。tnf - α诱导的内皮细胞凋亡与细胞内过氧化氢（约为对照组的20倍）和超氧化物（约为对照组的16倍）产生的急剧增加有关，分别通过二氯荧光素和二氢乙啶荧光染色测定。这种增加伴随着超氧化物歧化酶和谷胱甘肽过氧化物酶活性的降低，随后，脂质过氧化产物丙二醛的增加。CGP53353，而不是rotlerin，消除或减弱了所有这些变化。我们得出结论，PKCbeta(2)在tnf α诱导的人血管内皮细胞凋亡中起主要作用。
The circulatory inflammatory cytokine tumor necrosis factor alpha (TNF-alpha) is increased in pathologic conditions that initiate or exacerbate vascular endothelial injury, such as diabetes. Protein kinase C (PKC) has been shown to play a critical role in TNF-alpha-induced human endothelial cell apoptosis. However, the relative roles played by specific isoforms of PKC in TNF-alpha-induced human endothelial cell apoptosis have not been addressed. We investigated the effects of a selective PKCbeta(2) inhibitor (CGP53353) on TNF-alpha-induced apoptosis in human vascular endothelial cells (cell line ECV304) and on the production of reactive oxygen species and nitric oxide, and compared its effects with rottlerin, a reagent that has been shown to reduce PKCdelta protein levels. Cultured human vascular endothelial cells (ECV304) were treated for 24 h with one of 4 regimes: 40 ng/mL TNF-alpha alone (TNF-alpha), TNF-alpha with 10 micromol/L rottlerin (T+rottlerin), TNF-alpha with 1 micromol/L CGP53353 (T+CGP), or untreated (control). Cell viability was measured by MTT assay, and cell apoptosis was assessed by flow cytometry. TNF-alpha-induced endothelial cell apoptosis was associated with dramatic increases in production of intracellular hydrogen peroxide (approximately 20 times greater than control) and superoxide (approximately 16 times greater than control), as measured by dichlorofluorescein and dihydroethidium fluorescent staining, respectively. This increase was accompanied by reduced activity of superoxide dismutase and glutathione peroxidase and, subsequently, an increase in the lipid peroxidation product malondialdehyde. CGP53353, but not rottlerin, abolished or attenuated all these changes. We conclude that PKCbeta(2) plays a major role in TNF-alpha-induced human vascular endothelial cell apoptosis.
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5.7
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