Evidence for Mechanistic Alterations ofCa2+Homeostasis in Type 2 Diabetes Mellitus

Abstract

Altered cytosolicCa2+is implicated in the aetiology of many diseases including diabetes but there are few studies on the mechanism(s) of the alteredCa2+regulation. Using human lymphocytes, we studied cytosolic calcium (Cai) and variousCa2+transport mechanisms in subjects with Type 2 diabetes mellitus and control subjects.Ca2+-specific fluorescent probes (Fura-2 and Fluo-3) were used to monitor theCa2+signals. Thapsigargin, a potent and specific inhibitor of the sarco(endo)plasmic reticulumCa2+-ATPase (SERCA), was used to studyCa2+- store dependentCa2+fluxes. Significant (P< 0.05) elevation of basalCailevels was observed in lymphocytes from diabetic subjects.Cailevels 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 increasedCa2+influx (as measured both by Fluo-3 fliorescence andC45aassays) as a consequence of of thapsigargin-mediatedCa2+store depletion. Upon addition ofMn2+(a surrogate ofCa2+), 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 theNa+/Ca2+exchange activity in Type 2 diabetic patients, both under resting conditions and after challenging the cells with thapsigargin, when the internal storeCa2+sequestration was circumvented. Pharmacological activation of protein kinase C (PKC) in cells from patients resulted in only partial inhibition ofCa2+entry. We conclude that cellularCa2+accumulation in cells from Type 2 diabetes results from (a) reduction in PMCA ATPase activity, (b) modulation ofNa+/Ca2+exchange and (3) increasedCa2+influx across the plasma membrane.