Supplementary Material for: Pathways Responsible for Apoptosis Resulting from Amadori-Induced Oxidative and Nitrosative Stress in Human Mesothelial Cells

<i>Background:</i> Apoptosis and inflammatory/oxidative stress have been associated with hyperglycemia in human peritoneal mesothelial cells (HPMCs) and other cell types. We and others have highlighted the role of early products of non-enzymatic protein glycation in inducing proinflammatory conditions and increasing apoptotic rates in HPMCs. Loss of HPMCs seems to be a hallmark of complications associated with peritoneal membrane dysfunction. The aim of this work is to elucidate the mechanisms by which Amadori adducts may act upon HPMC apoptosis. <i>Methods:</i> HPMCs isolated from different patients were exposed to different Amadori adducts, i.e. highly glycated hemoglobin (10 n<i>M</i>) and glycated bovine serum albumin (250 µg/ml), to study cell death and several proapoptotic markers by different experimental approaches. <i>Results:</i> Amadori adducts, but not their respective controls, impaired cell proliferation and cell viability by means of apoptosis in a time-dependent manner. They regulated the intrinsic mitochondrial cell death signaling pathway and modulated activation of caspases, Bax, iNOS, p53, NF-ĸB, and mitogen-activated protein kinases (p38 and JNK) through different reactive oxygen and nitrosative species. <i>Conclusions:</i> Our data strongly support the idea that long-term hyperglycemia could act as an inducer of apoptosis in HPMCs through Amadori adducts, involving different oxidative and nitrosative reactive species.