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) en vironment is thought to mediate some of the vascular complications of diabe tes. Transmembrane signaling of contractile cells is generally inhibited by HG, with implications for systemic and target organ hemodynamics. In the k idney, glomerular mesangial cells grown in HG media are hyporesponsive to t he effects of vasoconstrictor agents, possibly explaining the hyperfiltrati on and increased capillary pressure that eventually lead to diabetic glomer ulopathy. To verify whether AGE binding to specific mesangial receptors cou ld mediate these effects of HG, cultured human mesangial cells (HMC) were e xposed to in vitro glycated bovine serum albumin (BSA) for 60 min at 37 deg rees C before measurement of cytosolic Ca2+ ([Ca2+](i)) by microfluorometri c techniques in monolayers or single cells. AGE-BSA (2 mg/ml) reduced Ca2release 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 +/- 6 8 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 am inoguanidine (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 a ddition of 1 to 10 mM extracellular Ca2+ to cells previously held in Ca2+-f ree 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 C a2+, respectively; +AGE-BSA, P < 9.05 versus control). Contrary to PIG, AGE -BSA did not translocate protein kinase C isoforms alpha, zeta, and delta t o the plasma membrane. Culture of HMC in HG supplemented with 1 mM AMGD pre vented downregulation of [Ca2+](i) signaling. These data suggest that glyca ted macromolecules or matrix components may inhibit transmembrane Ca2+ sign aling of glomerular cells through binding to a specific AGE receptor, thus mediating some of the known functional effects of HG on the kidney.