Acquisition of Stress Tolerance in Vascular Smooth Muscle Cells by The Induction of Mn-Sod

氧化应激诱导血管组织中的平滑肌细胞死亡，特别是在动脉粥样硬化斑块的纤维帽中，并通过斑块不稳定导致血管结构的破坏。我们研究了锰超氧化物歧化酶（Mn-SOD），这是在各种组织中诱导，并作为一个救援蛋白在响应外部压力，是否在血管平滑肌细胞（VSMC）中表达，并减弱氧化应激引起的细胞损伤。从大鼠主动脉分离培养大鼠主动脉平滑肌细胞（RASMC），用TNF-α（0-10 ng/ml）或热休克（42℃，1h）诱导细胞Mn-SOD。采用酶联免疫吸附法检测大鼠动脉平滑肌细胞中Mn-SOD的含量。在TNF-α作用后6 h，Mn-SOD的浓度从55.2± 11.8ng/mg蛋白增加到121.7± 10.6ng/mg蛋白（p<0.05）。Mn-SOD在热休克后12 h也从67.3±5.7 ng/mg蛋白增加到92.0±7.9 ng/mg蛋白（p<0.05 ...更多信息 热休克）。缺氧（6 h）-复氧（2 h）作为氧化应激，在TNF-D或热休克后6 h，细胞内Mn-SOD明显诱导，乳酸脱氢酶（LDH）释放作为细胞损伤的指标。TNF-α使缺氧-复氧后LDH释放显著减少至无TNF-α的对照培养物的32.1%（p<0.01）。热休克也使LDH释放减少到没有热休克的对照培养物的51.3%（p<0.05）。当Mn-SOD的反义寡脱氧核苷酸（500 nM）抑制Mn-SOD的诱导作用时，TNF-α对缺氧-复氧后LDH释放的抑制作用没有观察到。采用脂质体转染法将大鼠Mn-SOD基因导入RASMC，观察缺氧-复氧后细胞损伤情况。LDH释放降低至没有脂质转染的对照培养物的56.4%（p<0.01）。这些结果表明，由TNF-α或热休克诱导的内部Mn-SOD或通过使用含有大鼠Mn-SOD基因的载体转移的外部Mn-SOD增强血管平滑肌细胞对氧化应激的耐受性。少

Oxidative stress induces death of smooth muscle cells in vascular tissue, especially in the fibrous cap of atherosclerotic plaque, and leads to disruption of vessel structure via plaque unstabilization. We examined whether manganese superoxide dismutase (Mn-SOD), which is induced in various tissue and acts as one of rescue proteins in response to external stress, is expressed in vascular smooth muscle cells (VSMC) and attenuates cellular injury caused by oxidative stress. Rat aortic smooth muscle cells (RASMC) were cultured from rat aorta and cells were exposed to TNF-a (0-10 ng/ml) or heat shock (42℃, 1 h) to induce Mn-SOD. The concentration of Mn-SOD in RASMC was examined by enzyme linked immunosorbent assay. The concentration of Mn-SOD increased from 55.2±11.8 ng/mg protein to 121.7±10.6 ng/mg protein at 6 h after the exposure of TNF-a (p<0.05 vs. without TNF-a). Mn-SOD also increased from 67.3±5.7 ng/mg protein to 92.0±7.9 ng/mg protein at 12 h after heat shock (p<0.05 vs. without … More heat shock). RASMC were exposed to anoxia (6 h) - reoxygenation (2 h) as oxidative stress at 6 h after exposure to TNF-D or to heat shock, when Mn-SOD was markedly induced in cells, and lactate dehydrogenase (LDH) release was examined as an index of cellular injury. TNF-a attenuated LDH release after anoxia-reoxygenation significantly to 32.1% of control cultures without TNF-a (p<0.01). Heat shock also decreased LDH release to 51.3% of control cultures without heat shock (p<0.05). When Mn-SOD induction was inhibited by antisense oligodeoxyribonucleotide to Mn-SOD (500 nM), attenuation of LDH release after anoxia-reoxygenation by TNF-a was not observed. Rat Mn-SOD expression was transferred into RASMC by lipofection and the cellular injury after anoxia-reoxygenation was examined. The LDH release decreased to 56.4% of control cultures without the lipofection (p<0.01). These results suggest that internal Mn-SOD, which is induced by TNF-a or heat shock, or external Mn-SOD, which is transferred by using a vector containing the gene of rat Mn-SOD, enhances tolerance of vascular smooth muscle cells to oxidative stress. Less