Osteopontin (OPN) encodes a secreted glycosylated phosphoprotein containing a GRGDS motif that can mediate cell attachment through the αvβ3 integrin, and has recently been shown to down‐regulate nitric oxide synthase (NOS) expression. We report here that primary cultures of renal proximal tubule epithelial (PTE) cells prepared from human kidneys of different developmental stages and ages show a positive correlation between developmental age and the expression, at the mRNA level, of both OPN and constitutive NOS. However, OPN and NOS responded in different manners, as assessed by mRNA measurements, to hypoxia‐reoxygenation injury. The OPN mRNA level, assessed by Northern blotting, increased slightly during 60 min of hypoxia and more substantially during subsequent reoxygenation of primary PTE cells derived from the kidneys of young but not of aged donors. The abundance of NOS mRNA, measured using a cDNA probe to the constitutive form of the enzyme, was enhanced during hypoxia in kidneys derived from humans of all ages, and then decreased during reoxygenation—possibly as the result of increased OPN expression. PTE cells from aged kidneys are more susceptible to cell death under hypoxic conditions than PTE cells from young kidneys. An investigation of the effect of an oxidant on OPN and NOS mRNA levels revealed that within 30 min of exposure to H2O2, NOS mRNA levels decreased simultaneously with an increase in OPN mRNA levels. Nitric oxide (NO), the product of NOS, is at low levels an important signal transduction molecule participating in the regulation of vascular tone and renal reabsorption; at high levels it is cytotoxic. We suggest that the diminished ability of cells from old kidneys to down‐regulate NO production and to increase OPN expression after hypoxia‐reoxygenation may contribute to their increased susceptibility to oxidant injury. © 1994 Wiley‐Liss, Inc.