Evidence for Mechanistic Alterations of Ca(2+) Homeostasis in Type 2 Diabetes Mellitus

Altered cytosolic Ca(2+) is implicated in the aetiology of many diseases including diabetes but there are few studies on the mechanism(s) of the altered Ca(2+) regulation. Using human lymphocytes, we studied cytosolic calcium (Ca(i)) and various Ca(2+) transport mechanisms in subjects with Type 2 diabetes mellitus and control subjects. Ca(2+)-specific fluorescent probes (Fura-2 and Fluo-3) were used to monitor the Ca(2+) signals. Thapsigargin, a potent and specific inhibitor of the sarco(endo)plasmic reticulum Ca(2+)-ATPase (SERCA), was used to study Ca(2+)- store dependent Ca(2+) fluxes. Significant (P < 0.05) elevation of basal Ca(i) levels was observed in lymphocytes from diabetic subjects. Ca(i) levels were positively correlated with fasting, plasma glucose and HbAlc. There was also a significant (P < 0.05) reduction in plasma membrane calcium (PMCA) ATPase activity in diabetic subjects compared to controls. Cells from Type 2 diabetics exhibited an increased Ca(2+) influx (as measured both by Fluo-3 fliorescence and C45a assays) as a consequence of of thapsigargin-mediated Ca(2+) store depletion. Upon addition of Mn(2+) (a surrogate of Ca(2+)), the fura-2 fluorescence decayed in an exponential fashion and the rate and extent of this decline was steeper and greater in cells from type 2 diabetic patients. There was also a significant (P < 0.05) difference in the Na(+)/Ca(2+) exchange activity in Type 2 diabetic patients, both under resting conditions and after challenging the cells with thapsigargin, when the internal store Ca(2+) sequestration was circumvented. Pharmacological activation of protein kinase C (PKC) in cells from patients resulted in only partial inhibition of Ca(2+) entry. We conclude that cellular Ca(2+) accumulation in cells from Type 2 diabetes results from (a) reduction in PMCA ATPase activity, (b) modulation of Na(+)/Ca(2+) exchange and (3) increased Ca(2+) influx across the plasma membrane.