Buja L M, Fattor R A, Miller J C, Chien K R, Willerson J T
Department of Pathology, University of Texas, Health Science Center, Dallas.
Lab Invest. 1990 Sep;63(3):320-31.
We evaluated the contributions of calcium loading and impaired energy production to metabolic and ultrastructural manifestations of cell injury in a cultured neonatal rat ventriculocyte model. Direct calcium loading was produced by incubation in K(+)-free medium to inhibit the Na+,K(+)-ATPase and promote Na(+)-Ca2+ exchange, and inhibition of energy metabolism was produced by incubation with 30 microM iodoacetic acid (IAA). Measurements were made of total cell calcium, [3H] arachidonic acid (AA) release (an index of membrane phospholipid degradation), ATP, and ultrastructural features of cell damage. Inhibition of the Na(+),K(+) pump resulted in the rapid onset of cellular calcium loading, increased [3H]AA release, and moderate ATP reduction. After return to control medium for 24 hours, myocytes previously exposed to K(+)-free medium for 1 hour showed recovery of ATP level and little additional [3H]AA release. However, after 2 to 3 hours of calcium loading, the ATP level remained moderately depressed, residual [3H]AA release was greater, and a mixed population of relatively normal and severely damaged myocytes was observed by electron microscopy. IAA treatment for 1 hour resulted in moderate ATP reduction without calcium accumulation or [3H]AA release, whereas IAA treatment for 3 hours resulted in marked ATP reduction associated with calcium accumulation and [3H]AA release. Reversal experiments showed substantial recovery of ATP level after 1 hour of IAA exposure, and marked ATP depression and [3H]AA release associated with widespread irreversible injury after 3 hours. Thus, the data indicate that increased calcium accumulation itself can initiate accelerated membrane phospholipid degradation, but that progression to irreversible injury is influenced by other factors, including the magnitude of ATP depression associated with calcium loading.
我们在培养的新生大鼠心室肌细胞模型中评估了钙超载和能量产生受损对细胞损伤的代谢及超微结构表现的影响。通过在无钾培养基中孵育以抑制钠钾ATP酶并促进钠钙交换来实现直接钙超载，通过与30微摩尔碘乙酸（IAA）孵育来抑制能量代谢。测量了细胞总钙、[3H]花生四烯酸（AA）释放（膜磷脂降解的指标）、ATP以及细胞损伤的超微结构特征。钠钾泵的抑制导致细胞钙超载迅速发生、[3H]AA释放增加以及ATP适度降低。在恢复到对照培养基24小时后，先前暴露于无钾培养基1小时的心肌细胞显示ATP水平恢复，且[3H]AA释放几乎没有进一步增加。然而，在钙超载2至3小时后，ATP水平仍适度降低，残余[3H]AA释放更多，并且通过电子显微镜观察到存在相对正常和严重受损的心肌细胞混合群体。IAA处理1小时导致ATP适度降低，而没有钙积累或[3H]AA释放，而IAA处理3小时导致与钙积累和[3H]AA释放相关的显著ATP降低。逆转实验表明，IAA暴露1小时后ATP水平有显著恢复，而暴露3小时后则出现与广泛不可逆损伤相关的显著ATP降低和[3H]AA释放。因此，数据表明钙积累增加本身可引发膜磷脂降解加速，但向不可逆损伤的进展受其他因素影响，包括与钙超载相关的ATP降低程度。
