Studies On The Heterogeneity Of Basal Intracellular Calcium Concentration And Its Relations To The Reactivity Of Mouse Peritoneal Macrophages

As important inflammatory cells, macrophages play an important role on the development of inflammation. There have been many reports on the reactivity of macrophages in the literature with pooled macrophage samples, but the results were contradictory. Considering the existence of the different epitops in macrophages, the function of macrophages may also be heterogeneous, so the actual changes of the individual cells can be hardly detected if the measurements were made in the pooled cell samples. Therefore, in the present work, the cytosolic free calciurn([Ca2+]i), superoxide anions(02-) and the reactivity of the peritoneal macrophages(PM) in mice were studied at the single cell level. [Ca2+]i was determined by Fura-2. The modified method is as follows:①PM were adhered for 30-90mm; ②the loading medium containing PM Fura-2/AM and 0.04% pluronic F-127 was incubated with PM for 1hr at room temperature; ③The Grynkiewicz constant was determined by "EGTA/AM method"in each mouse instead of a constant value used in every mouse. The values of basal [Ca2+]i determined in 392 PMs of 7 mice showed normal distribution(54±24nM, n=392) with wide range(less than 2OnM to more than lOOnM), among which about 50% were in the range of 40-6OnM. When stimulated with PMA or tMLP, [Ca2+]i was increased, the peak values were positively correlated with the basal [Ca2+]i (PMA stimulated cells: r=0.52, p