Scheitlin Christopher G, Julian Justin A, Shanmughapriya Santhanam, Madesh Muniswamy, Tsoukias Nikolaos M, Alevriadou B Rita
Departments of Biomedical Engineering and Internal Medicine, Division of Cardiovascular Medicine, and Davis Heart and Lung Research Institute, The Ohio State University, Columbus, Ohio;
Department of Medical Genetics and Molecular Biochemistry and Center for Translational Medicine, Temple University, Philadelphia, Pennsylvania; and.
Am J Physiol Cell Physiol. 2016 Mar 15;310(6):C479-90. doi: 10.1152/ajpcell.00171.2015. Epub 2016 Jan 6.
Shear stress is known to stimulate an intracellular free calcium concentration ([Ca(2+)]i) response in vascular endothelial cells (ECs). [Ca(2+)]i is a key second messenger for signaling that leads to vasodilation and EC survival. Although it is accepted that the shear-induced [Ca(2+)]i response is, in part, due to Ca(2+) release from the endoplasmic reticulum (ER), the role of mitochondria (second largest Ca(2+) store) is unknown. We hypothesized that the mitochondria play a role in regulating [Ca(2+)]i in sheared ECs. Cultured ECs, loaded with a Ca(2+)-sensitive fluorophore, were exposed to physiological levels of shear stress. Shear stress elicited [Ca(2+)]i transients in a percentage of cells with a fraction of them displaying oscillations. Peak magnitudes, percentage of oscillating ECs, and oscillation frequencies depended on the shear level. [Ca(2+)]i transients/oscillations were present when experiments were conducted in Ca(2+)-free solution (plus lanthanum) but absent when ECs were treated with a phospholipase C inhibitor, suggesting that the ER inositol 1,4,5-trisphosphate receptor is responsible for the [Ca(2+)]i response. Either a mitochondrial uncoupler or an electron transport chain inhibitor, but not a mitochondrial ATP synthase inhibitor, prevented the occurrence of transients and especially inhibited the oscillations. Knockdown of the mitochondrial Ca(2+) uniporter also inhibited the shear-induced [Ca(2+)]i transients/oscillations compared with controls. Hence, EC mitochondria, through Ca(2+) uptake/release, regulate the temporal profile of shear-induced ER Ca(2+) release. [Ca(2+)]i oscillation frequencies detected were within the range for activation of mechanoresponsive kinases and transcription factors, suggesting that dysfunctional EC mitochondria may contribute to cardiovascular disease by deregulating the shear-induced [Ca(2+)]i response.
已知剪切应力可刺激血管内皮细胞（ECs）内的细胞内游离钙浓度（[Ca(2+)]i）反应。[Ca(2+)]i是导致血管舒张和EC存活的信号传导的关键第二信使。尽管人们普遍认为剪切诱导的[Ca(2+)]i反应部分归因于内质网（ER）释放Ca(2+)，但线粒体（第二大Ca(2+)储存库）的作用尚不清楚。我们假设线粒体在调节剪切力作用下的ECs中的[Ca(2+)]i方面发挥作用。用Ca(2+)敏感荧光团加载的培养ECs暴露于生理水平的剪切应力下。剪切应力在一定比例的细胞中引发[Ca(2+)]i瞬变，其中一部分细胞表现出振荡。峰值大小、振荡ECs的百分比和振荡频率取决于剪切水平。在无Ca(2+)溶液（加镧）中进行实验时存在[Ca(2+)]i瞬变/振荡，但在用磷脂酶C抑制剂处理ECs时则不存在，这表明ER肌醇1,4,5-三磷酸受体负责[Ca(2+)]i反应。线粒体解偶联剂或电子传递链抑制剂，但不是线粒体ATP合酶抑制剂，可阻止瞬变的发生，尤其抑制振荡。与对照组相比，线粒体Ca(2+)单向转运体的敲低也抑制了剪切诱导的[Ca(2+)]i瞬变/振荡。因此，EC线粒体通过Ca(2+)摄取/释放，调节剪切诱导的ER Ca(2+)释放的时间特征。检测到的[Ca(2+)]i振荡频率在机械反应性激酶和转录因子激活的范围内，这表明功能失调的EC线粒体可能通过解除对剪切诱导的[Ca(2+)]i反应的调节而导致心血管疾病。
