Investigating Mitochondrial Metabolism in Contracting HL-I Cardiomyocytes Following Hypoxia and Pharmacological HIF Activation Identifies HIF-Dependent and Independent Mechanisms of Regulation
Investigating Mitochondrial Metabolism in Contracting HL-I Cardiomyocytes Following Hypoxia and Pharmacological HIF Activation Identifies HIF-Dependent and Independent Mechanisms of Regulation
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DOI：
10.1177/1074248414524480
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发表时间：
2014-11-01
影响因子：
2.6
通讯作者：
Heather, Lisa C.
中科院分区：
文献类型：
--
作者：
Ambrose, Lucy J. A.;Abd-Jamil, Amira H.;Heather, Lisa C.
低氧是心脏病的结果，并下调线粒体新陈代谢，但其在心脏中发生的分子机制尚不完全清楚。因此，我们的目标是使用收缩HL-I心肌细胞模型来研究缺氧对线粒体代谢的影响。细胞暴露于低氧(2%O-2)6、12、24和48小时以表征代谢反应。随后用低氧诱导因子(HIF)激活化合物二甲氧甘氨酸(DMOG)处理细胞，以确定低氧诱导的线粒体变化是否依赖于HIF，并评估该培养的心肌细胞系是否适合进行心血管药理学研究。缺氧细胞在24小时后糖酵解增加，葡萄糖转运蛋白1和乳酸水平分别增加5倍和5倍。缺氧24小时后，线粒体网络更加碎裂，但柠檬酸合成酶活性没有变化，表明线粒体含量没有变化。与对照组相比，细胞耗氧量降低了30%，同时电子传递链(ETC)复合体1和IV以及乌头酸酶活性分别降低了81%、96%和72%。DMOG对HIF的药理激活使细胞耗氧量降低43%，与乌头酸酶和复合体1活性分别降低26%和30%相一致，表明这种适应是由HIF介导的。相反，低氧介导的复合体IV活性的降低不能被DMOG处理所复制，这表明该复合体的调节不依赖于HIF。综上所述，缺氧24小时增加了无氧糖酵解，降低了线粒体呼吸，这与收缩HL-I细胞的ETC和三羧酸循环酶活性的变化有关。在这种心肌细胞系中，药理上的HIF激活使得能够识别HIF依赖和独立的线粒体代谢变化。
Hypoxia is a consequence of cardiac disease and downregulates mitochondrial metabolism, yet the molecular mechanisms through which this occurs in the heart are incompletely characterized. Therefore, we aimed to use a contracting HL-I cardiomyocyte model to investigate the effects of hypoxia on mitochondrial metabolism. Cells were exposed to hypoxia (2% O-2) for 6, 12, 24, and 48 hours to characterize the metabolic response. Cells were subsequently treated with the hypoxia inducible factor (HIF)-activating compound, dimethyloxalylglycine (DMOG), to determine whether hypoxia-induced mitochondrial changes were HIF dependent or independent, and to assess the suitability of this cultured cardiac cell line for cardiovascular pharmacological studies. Hypoxic cells had increased glycolysis after 24 hours, with glucose transporter 1 and lactate levels increased 5-fold and I 5-fold, respectively. After 24 hours of hypoxia, mitochondrial networks were more fragmented but there was no change in citrate synthase activity, indicating that mitochondrial content was unchanged. Cellular oxygen consumption was 30% lower, accompanied by decreases in the enzymatic activities of electron transport chain (ETC) complexes 1 and IV, and aconitase by 81%, 96%, and 72%, relative to controls. Pharmacological HIF activation with DMOG decreased cellular oxygen consumption by 43%, coincident with decreases in the activities of aconitase and complex 1 by 26% and 30%, indicating that these adaptations were HIF mediated. In contrast, the hypoxia-mediated decrease in complex IV activity was not replicated by DMOG treatment, suggesting HIF-independent regulation of this complex. In conclusion, 24 hours of hypoxia increased anaerobic glycolysis and decreased mitochondrial respiration, which was associated with changes in ETC and tricarboxylic acid cycle enzyme activities in contracting HL-I cells. Pharmacological HIF activation in this cardiac cell line allowed both HIF-dependent and independent mitochondrial metabolic changes to be identified.