Effects of advanced glycation end products on cytosolic Ca2+ signaling of cultured human mesangial cells

Advanced glycation end product (AGE) accumulation in a high glucose (HG) environment is thought to mediate some of the vascular complications of diabetes. Transmembrane signaling of contractile cells is generally inhibited by HG, with implications for systemic and target organ hemodynamics. In the kidney, glomerular mesangial cells grown in HG media are hyporesponsive to the effects of vasoconstrictor agents, possibly explaining the hyperfiltration and increased capillary pressure that eventually lead to diabetic glomerulopathy. To verify whether AGE binding to specific mesangial receptors could mediate these effects of HG, cultured human mesangial cells (HMC) were exposed to in vitro glycated bovine serum albumin (BSA) for 60 min at 37 degrees C before measurement of cytosolic Ca2+ ([Ca2+](i)) by microfluorometric techniques in monolayers or single cells. AGE-BSA (2 mg/ml) reduced Ca2+ release from intracellular stores by 1 mu M angiotensin II from peak [Ca2+](i) levels of 843 +/- 117 to 390 +/- 50 nM in monolayers and from 689 +/- 68 to 291 +/- 36 nM in individual cells (P < 0.05). Nonglycated BSA and BSA exposed to 250 mM glucose-6-phosphate for 30 d in the presence of 250 mM aminoguanidine (AMGD), an inhibitor of nonenzymatic glycation, had no effect on the angiotensin II-induced [Ca2+](i) spike (peak 766 +/- 104 and 647 +/- 87 nM, monolayers/ single cells, respectively, P = NS). AGE also inhibited store-operated Ca2+ influx through plasma membrane channels, assessed by addition of 1 to 10 mM extracellular Ca2+ to cells previously held in Ca2+-free media (control 339 +/- 46/593 +/- 51, +AGE-BSA 236 +/- 25/390 +/- 56, +AMGD 483 +/- 55/ 374 +/- 64 nM [Ca2+](i) monolayers/single cells at 10 mM Ca2+, respectively; +AGE-BSA, P < 9.05 versus control). Contrary to PIG, AGE-BSA did not translocate protein kinase C isoforms alpha, zeta, and delta to the plasma membrane. Culture of HMC in HG supplemented with 1 mM AMGD prevented downregulation of [Ca2+](i) signaling. These data suggest that glycated macromolecules or matrix components may inhibit transmembrane Ca2+ signaling of glomerular cells through binding to a specific AGE receptor, thus mediating some of the known functional effects of HG on the kidney.