Hypoxia and oxidative stress are considered to be underlying factors in the deterioration of renal function and pathogenesis in acute kidney injury (AKI) and chronic kidney disease, including diabetic nephropathy (DN). However, the long-term role of hypoxia in DN is unknown. Here, we investigated the distribution, severity, and time course of hypoxia during DN development in our well-established severely diabetic transgenic (Tg) DN mouse model that mimics human DN up to 80 wk of age, using pimonidazole adduct immunohistochemistry. The relationship between pimonidazole adduct distribution and hypoxia-inducible factor (HIF) expression was also examined. We found 1) persistent pimonidazole immunostaining mainly in the outer zone of the outer medulla, extending into the inner zone, 2) significant expansion of area and intensity up to 40 wk of age, and 3) characteristic subcellular localization mainly at apical sites in vesicular form by laser scanning microscopy of thin slices. The distribution of pimonidazole adducts was different from that of HIF reported previously, indicating that hypoxia does not directly contribute to persistent abnormal HIF expression. These results suggest that pimonidazole adducts produced under low [Formula: see text] conditions are sustained by a mechanism distinct from direct ischemia. We propose that in the long course of DN development, persistent hyperfiltration and hyperexcretion of glucose, albumin, and water increase metabolism and energy expenditure in the tubules, and such chronic stimulation leads to relative ischemia and local hypoxia, which may contribute in part to the loss of nephrons.NEW & NOTEWORTHY This study provides new insights into hypoxia during the long course of diabetic nephropathy development. Hypoxia was persistently localized only in limited areas and its distribution differed significantly from that of hypoxia-inducible factors. These findings suggests that in the long course of diabetic nephropathy development, increased energy requirements and limited blood supply may lead to relative ischemia and induction of local and persistent hypoxia, which may contribute in part to the loss of nephrons.
Keywords: diabetic nephropathy; hypoxia; hypoxia-inducible factor; pimonidazole adduct; type 2 diabetes.