Inflammation alters regional mitochondrial Ca2+ in human airway smooth muscle cells

Delmotte P, Yang B, Thompson MA, Pabelick CM, Prakash YS, Sieck GC. Inflammation alters regional mitochondrial Ca2+ in human airway smooth muscle cells. Am J Physiol Cell Physiol 303: C244-C256, 2012. First published June 6, 2012; doi:10.1152/ajpcell.00414.2011.-Regulation of cytosolic Ca2+ concentration ([Ca2+](cyt)) in airway smooth muscle (ASM) is a key aspect of airway contractility and can be modulated by inflammation. Mitochondria have tremendous potential for buffering [Ca2+](cyt), helping prevent Ca2+ overload, and modulating other intracellular events. Here, compartmentalization of mitochondria to different cellular regions may subserve different roles. In the present study, we examined the role of Ca2+ buffering by mitochondria and mitochondrial Ca2+ transport mechanisms in the regulation of [Ca2+](cyt) in enzymatically dissociated human ASM cells upon exposure to the proinflammatory cytokines TNF-alpha and IL-13. Cells were loaded simultaneously with fluo-3 AM and rhod-2 AM, and [Ca2+](cyt) and mitochondrial Ca2+ concentration ([Ca2+](mito)) were measured, respectively, using real-time two-color fluorescence microscopy in both the perinuclear and distal, perimembranous regions of cells. Histamine induced a rapid increase in both [Ca2+](cyt) and [Ca2+](mito), with a significant delay in the mitochondrial response. Inhibition of the mitochondrial Na+/Ca2+ exchanger (1 mu M CGP-37157) increased [Ca2+](mito) responses in perinuclear mitochondria but not distal mitochondria. Inhibition of the mitochondrial uniporter (1 mu M Ru360) decreased [Ca2+](mito) responses in perinuclear and distal mitochondria. CGP-37157 and Ru360 significantly enhanced histamine-induced [Ca2+](cyt). TNF-alpha and IL-13 both increased [Ca2+](cyt), which was associated with decreased [Ca2+](mito) in the case of TNF-alpha but not IL-13. The effects of TNF-alpha on both [Ca2+](cyt) and [Ca2+](mito) were affected by CGP-37157 but not by Ru360. Overall, these data demonstrate that in human ASM cells, mitochondria buffer [Ca2+](cyt) after agonist stimulation and its enhancement by inflammation. The differential regulation of [Ca2+](mito) in different parts of ASM cells may serve to locally regulate Ca2+ fluxes from intracellular sources versus the plasma membrane as well as respond to differential energy demands at these sites. We propose that such differential mitochondrial regulation, and its disruption, may play a role in airway hyperreactivity in diseases such as asthma, where [Ca2+](cyt) is increased.