Kobayashi S, Miescher E, Clemens M G
Division of Pediatric Surgery, The Johns Hopkins University School of Medicine, Baltimore, Maryland 21287, USA.
Transplantation. 1999 Feb 15;67(3):451-7. doi: 10.1097/00007890-199902150-00019.
Calcium accumulation of cells and mitochondria during reperfusion or reoxygenation has been implicated as a potential factor in cell injury as the result of mitochondrial damage. The objective of this study was to disclose whether or not low extracellular calcium ion concentration ([Ca2+]ex) in the medium at the time of reoxygenation might prevent calcium accumulation and attenuate hepatocytes injury after severe hypoxia.
Isolated rat hepatocytes were incubated under a hyperoxic or hypoxic atmosphere for 60 min. During the ensuing 60-min hyperoxic reoxygenation, medium [Ca2+]ex was varied from 0.6 microM to 2.0 mM by altering total calcium and addition of chelators.
Incubation in low [Ca2+]ex reduced total cellular calcium and mitochondrial calcium in both the hyperoxic and hypoxic group. Under hyperoxic/hyperoxic incubation (control), hepatocytes were able to maintain potassium balance when [Ca2+]ex was >3.0 microM (pCa=5.5) and cellular viability (% lactate dehydrogenase release) at all levels of extracellular calcium. Under hypoxic/hyperoxic incubation (reoxygenation), however, loss of the ability to restore potassium balance as well as apparent increase in lactate dehydrogenase release were observed at severely low [Ca2+]ex (<30 microM; pCa=4.5). This low [Ca2+]ex-induced exacerbation of hepatocytes viability could not be generated under mild reoxygenation such as normoxia.
In normal isolated hepatocytes, very low [Ca2+]ex levels produce only very subtle changes in membrane permeability of isolated hepatocytes. After hypoxia, however, hypocalcemia acts synergistically with hyperoxic reoxygenation to produce more severe damage. These results suggested that [Ca2+]ex should be maintained on the physiological level to attenuate hepatocytes injury after severe hypoxia.
再灌注或复氧期间细胞和线粒体的钙积累被认为是线粒体损伤导致细胞损伤的一个潜在因素。本研究的目的是揭示复氧时培养基中低细胞外钙离子浓度（[Ca2+]ex）是否能预防钙积累并减轻严重缺氧后的肝细胞损伤。
将分离的大鼠肝细胞在高氧或低氧气氛下孵育60分钟。在随后60分钟的高氧复氧期间，通过改变总钙量和添加螯合剂，使培养基[Ca2+]ex从0.6微摩尔/升变化至2.0毫摩尔/升。
在高氧和低氧组中，低[Ca2+]ex孵育均降低了细胞总钙和线粒体钙。在高氧/高氧孵育（对照）下，当[Ca2+]ex>3.0微摩尔/升（pCa = 5.5）时，肝细胞能够维持钾平衡，且在所有细胞外钙水平下细胞活力（乳酸脱氢酶释放百分比）均正常。然而，在低氧/高氧孵育（复氧）下，在极低的[Ca2+]ex（<30微摩尔/升；pCa = 4.5）时，观察到恢复钾平衡的能力丧失以及乳酸脱氢酶释放明显增加。这种低[Ca2+]ex诱导的肝细胞活力恶化在轻度复氧如常氧条件下不会发生。
在正常分离的肝细胞中，极低的[Ca2+]ex水平仅引起分离肝细胞膜通透性的非常细微的变化。然而，缺氧后，低钙血症与高氧复氧协同作用产生更严重的损伤。这些结果表明，应将[Ca2+]ex维持在生理水平以减轻严重缺氧后的肝细胞损伤。
