Ladilov Y V, Balser C, Piper H M
Physiologisches Institut, Justus-Liebig-Universität, Giessen, Germany.
Circ Res. 1998 Mar 9;82(4):451-7. doi: 10.1161/01.res.82.4.451.
The aim of this study was to investigate whether treatment with the protein kinase C (PKC) agonist 1,2-dioctanoyl-sn-glycerol (1,2DOG) can protect isolated adult Wistar rat cardiomyocytes against simulated ischemia and reoxygenation. Cytosolic Ca2+ (assessed by fura 2 fluorescence), pHi (assessed by BCECF fluorescence), and cell length were measured during 80 minutes of simulated ischemia (anoxia, pHo 6.4) and 20 minutes of reoxygenation (pHo 7.4) and compared between control cells and cells treated with 20 micromol/L 1,2DOG before anoxia (10-minute treatment and 10-minute washout), before and during anoxia (two-step treatment), or only during anoxia. Treatment before anoxia attenuated rigor contracture but did not influence anoxic Ca2+ overload. In contrast, two-step treatment before and during anoxia accelerated rigor contracture but reduced the rate of anoxic Ca2+ accumulation. During reoxygenation, control cells developed irreversible hypercontracture (reduction of cell length to 43+/-2% of the initial cell length, n=62), which was accompanied by spontaneous oscillations of cytosolic Ca2+ (19.6+/-1.6 per minute). Two-step treatment with 1,2DOG before and during anoxia significantly reduced hypercontracture (reduction of cell length to 60+/-2%, P<.01 versus control, n=41) and suppressed spontaneous Ca2+ oscillations (2.8+/-0.9 per minute, P<.01 versus control). These effects could not be reproduced by treatment with 1,2DOG before anoxia or during anoxia or by a two-step treatment with the PKC-inactive 1,3-dioctanoyl-sn-glycerol and were fully abolished with 1 micromol/L bisindolylmaleimide (PKC inhibitor). We conclude that a two-step activation of PKC before and during anoxia is required for effective protection of cardiomyocytes against anoxic Ca2+ overload and reoxygenation-induced hypercontracture.
本研究的目的是调查用蛋白激酶C（PKC）激动剂1,2 - 二辛酰 - sn - 甘油（1,2DOG）进行处理是否能够保护成年Wistar大鼠离体心肌细胞免受模拟缺血和复氧损伤。在80分钟的模拟缺血（缺氧，细胞外pH 6.4）和20分钟的复氧（细胞外pH 7.4）过程中，测量胞质Ca2+（通过fura 2荧光评估）、细胞内pH（通过BCECF荧光评估）和细胞长度，并将对照组细胞与在缺氧前（10分钟处理和10分钟洗脱）、缺氧前及缺氧期间（两步处理）或仅在缺氧期间用20 μmol/L 1,2DOG处理的细胞进行比较。缺氧前处理减轻了强直收缩，但不影响缺氧性Ca2+超载。相反，缺氧前及缺氧期间的两步处理加速了强直收缩，但降低了缺氧性Ca2+积累的速率。在复氧期间，对照细胞出现不可逆的过度收缩（细胞长度减少至初始细胞长度的43±2%，n = 62），同时伴有胞质Ca2+的自发振荡（每分钟19.6±1.6次）。缺氧前及缺氧期间用1,2DOG进行两步处理可显著减轻过度收缩（细胞长度减少至60±2%，与对照组相比P<0.01，n = 41）并抑制Ca2+自发振荡（每分钟2.8±0.9次，与对照组相比P<0.01）。缺氧前或缺氧期间用1,2DOG处理或用PKC无活性的1,3 - 二辛酰 - sn - 甘油进行两步处理均无法重现这些效应，并且用1 μmol/L双吲哚基马来酰亚胺（PKC抑制剂）可完全消除这些效应。我们得出结论，缺氧前及缺氧期间对PKC进行两步激活是有效保护心肌细胞免受缺氧性Ca2+超载和复氧诱导的过度收缩所必需的。
