Preliminary data suggest different intracellular calcium handling of Th1 and Th2 lymphocytes that may contribute to distinct cytokine production patterns. In this study we explored the contribution of the main mechanisms in charge of the elevation and decrease of cytoplasmic free calcium levels, i.e., the endoplasmic calcium release, the calcium release activated calcium (CRAC) channel, the mitochondrial calcium uniporter (MCU), the sarco/endoplasmic reticulum calcium ATPase (SERCA), and the plasma membrane calcium ATPase (PMCA) during the first 10 min of activation in human Th1 and Th2 lymphocytes applying a kinetic flow cytometry approach. We isolated peripheral blood mononuclear cells from 10 healthy individuals. Cells were stained with CD4, CXCR3 and CCR4 cell surface markers to identify Th1 and Th2 cells, respectively and loaded with Fluo-3/AM calcium sensitive dye. Cells were activated with phytohemagglutinine and alterations of cytoplasmic free calcium levels were monitored for 10 min after specific inhibition of the above mechanisms. Our results revealed delicate differences in calcium flux kinetics of Th1 and Th2 lymphocytes. The lower activity of MCU, and therefore of CRAC channels, along with the higher activity of the SERCA pump account for the notion that Th2 cells go through a lower level of lymphocyte activation compared with Th1 cells upon identical activating stimuli. The observed differences in calcium flux of Th1 and Th2 cells may contribute to different calcium handling kinetics and, hence, to distinct cytokine production patterns by these subsets.
