Amadori products and Advanced Glycated End‐products in glomerular tissues of short‐ and long‐term diabetic animals

High blood glucose levels play major roles in the pathogenesis of diabetic nephropathy through glycation of long‐lived molecules such as those of the extracellular matrix. Upon exposure to high glucose levels, long‐lived proteins get glycated into early glycated molecules (Schiff bases and Amadori products) and then, with time, into advanced glycated end‐products (AGEs). Using specific antibodies against glucitol‐lysine and anti‐AGEs, we revealed by immuno‐electron microscopy and by western blot the two categories of glycated proteins in glomerular basement membrane (GBM) of short‐ (3 months) and long‐term (6 and 15 months) diabetic animals. The use of the anti‐glucitol lysine in conjunction with protein A‐gold led to labelings over different tissular and cellular compartments, but mainly over the GBM. Tissues from control normoglycaemic animals yield very low signals, while tissues from 3 months diabetic animals displayed strong labelings. This was confirmed by western blot using purified GBM material isolated from glomerular tissue of normoglycaemic and hyperglycaemic animals. The use of the anti‐AGE antibody yield similar results. Labelings over tissues from normoglycaemic animals were very low, while those of diabetic animals were significantly increased. Surprisingly, levels of AGEs labelings, already high on tissues of 3 months diabetic animals, did not increase with age; tissues from 15 months diabetic animals displaying labelings similar in intensity to those of 3 months diabetic animals. This must be related to the turnover of the extracellular matrix proteins which, although being long‐lived proteins, do have a half‐life of about 2–3 months. No accumulation of Amadori or AGEs products is expected beyond this length of time. Supported by the Canadian Institutes of Health Research.