Bioenergetic characterization of mouse podocytes
Bioenergetic characterization of mouse podocytes
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DOI：
10.1152/ajpcell.00563.2009
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发表时间：
2010-08-01
影响因子：
5.5
通讯作者：
Kopp, Jeffrey B.
中科院分区：
文献类型：
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作者：
Abe, Yoshifusa;Sakairi, Toru;Kopp, Jeffrey B.
Abe Y，Sakairi T，Kajiyama H，Shrivastav S，Beeson C，Kopp JB.小鼠足细胞的生物能量表征。美国生理学杂志细胞生理学299：C464-C476，2010年。首次发表于2010年5月5日; doi：10.1152/ajpcell.00563.2009。线粒体功能障碍导致足细胞损伤，但正常足细胞生物能量学尚未得到表征。我们使用转化的小鼠足细胞系和Seahorse Bioscience XF 24胞外通量分析仪测量了耗氧率（OCR）和胞外酸化率（ECAR）。基础OCR和ECAR分别为55.2 +/- 9.9 pmol/min和3.1 +/- 1.9毫pH单位/min。复合物V抑制剂寡霉素将OCR降低至与基线速率的45%相似，表明与ATP合成偶联的细胞耗氧量接近55%。鱼藤酮，一种复合物I抑制剂，降低OCR到接近基线速率的25%，表明线粒体呼吸占总细胞呼吸的75%。因此，大约75%的线粒体呼吸与ATP合成偶联，大约25%的线粒体呼吸与质子泄漏偶联。羰基氰化物对三氟甲氧基苯腙（FCCP），从ATP产生的电子传递解偶联，增加OCR和ECAR类似的360%和840%的控制水平。FCCP加鱼藤酮使ATP含量降低60%，糖酵解抑制剂2-脱氧葡萄糖使ATP降低35%，2-脱氧葡萄糖与FCCP或鱼藤酮组合使ATP降低> 85%。乳酸脱氢酶抑制剂草胺酸和2-脱氧葡萄糖没有降低ECAR，2-脱氧葡萄糖对OCR没有影响，尽管2-脱氧葡萄糖使ATP含量降低了25%。FCCP诱导的线粒体解偶联与某些底物（包括乳酸、葡萄糖、丙酮酸和棕榈酸）的OCR增加相关。在原代小鼠足细胞中重复这些实验产生了类似的数据。我们的结论是线粒体在维持足细胞能量平衡中起主要作用，而糖酵解作用较小。
Abe Y, Sakairi T, Kajiyama H, Shrivastav S, Beeson C, Kopp JB. Bioenergetic characterization of mouse podocytes. Am J Physiol Cell Physiol 299: C464-C476, 2010. First published May 5, 2010; doi: 10.1152/ajpcell.00563.2009.-Mitochondrial dysfunction contributes to podocyte injury, but normal podocyte bioenergetics have not been characterized. We measured oxygen consumption rates (OCR) and extracellular acidification rates (ECAR), using a transformed mouse podocyte cell line and the Seahorse Bioscience XF24 Extracellular Flux Analyzer. Basal OCR and ECAR were 55.2 +/- 9.9 pmol/min and 3.1 +/- 1.9 milli-pH units/min, respectively. The complex V inhibitor oligomycin reduced OCR to similar to 45% of baseline rates, indicating that similar to 55% of cellular oxygen consumption was coupled to ATP synthesis. Rotenone, a complex I inhibitor, reduced OCR to similar to 25% of the baseline rates, suggesting that mitochondrial respiration accounted for similar to 75% of the total cellular respiration. Thus similar to 75% of mitochondrial respiration was coupled to ATP synthesis and similar to 25% was accounted for by proton leak. Carbonyl cyanide p-trifluoromethoxyphenylhydrazone (FCCP), which uncouples electron transport from ATP generation, increased OCR and ECAR to similar to 360% and 840% of control levels. FCCP plus rotenone reduced ATP content by 60%, the glycolysis inhibitor 2-deoxyglucose reduced ATP by 35%, and 2-deoxyglucose in combination with FCCP or rotenone reduced ATP by >85%. The lactate dehydrogenase inhibitor oxamate and 2-deoxyglucose did not reduce ECAR, and 2-deoxyglucose had no effect on OCR, although 2-deoxyglucose reduced ATP content by 25%. Mitochondrial uncoupling induced by FCCP was associated with increased OCR with certain substrates, including lactate, glucose, pyruvate, and palmitate. Replication of these experiments in primary mouse podocytes yielded similar data. We conclude that mitochondria play the primary role in maintaining podocyte energy homeostasis, while glycolysis makes a lesser contribution.