We examined the effects of in vitro nonenzymatic glycosylation (NEG) of the extracellular matrix (ECM) on various aspects of the adhesive interaction between the ECM and human kidney mesangial (HKM) cells, and also on the ability of HKM cells to spread and proliferate. Isolated type IV collagen (tIV) or intact complexes of glomerular basement membrane and mesangial matrix (GBM/MM) were used as substrates following control incubation or modest glycation. We observed that HKM cells adhered less effectively to glycated tIV at early time intervals and were delayed in attaining maximal levels of adhesion compared to cells interacting with control tIV. The nature of the adhesive interaction was also different since antibodies which blocked the function of β1-containing integrins more effectively inhibited adhesion between HKM cells and glycated tIV than cells and control tIV. HKM cells interacting with glycated tIV also demonstrated less cell surface microspike and ruffle formation at five minutes after plating, less extensive spreading throughout the examined time intervals (≥ 90 min after plating), and slightly increased cell numbers 5 to 10 days after plating when compared to cells interacting with control tIV. However, increased cell numbers were not observed when HKM cells were grown on glycated GBM/MM. Similar changes in response to glycated substrate were observed when HKM cells were grown in either 5 or 25 mM glucose. In conclusion, relatively modest glycation of the ECM alone was sufficient to result in specific changes in HKM cell behavior in vitro. It is possible that the matrix glycation which occurs in the kidneys of diabetic patients could alter mesangial cell behavior in situ in ways which may contribute to the development of diabetic nephropathy.