Diabetic nephropathy is a major cause of the increased morbidity and mortality in insulin-dependent diabetes mellitus. The most significant renal lesion of diabetic nephropathy is expansion of the glomerular mesangium. Thickening of the glomerular basement membrance is also apparent. Mesangial expansion is largely due to the accumulation of extracellular matrix (ECM) proteins such as fibronectin, laminin, and type IV collagen. To determine whether high glucose is responsible for the observed increase in mesangial cell ECM protein accumulation, mesangial cells were grown in tissue culture medium containing 10 mmol/l (millimolar) glucose (normal) or 30 mmol/l glucose (high). The degree of ECM protein accumulation was determined by immunocytochemistry and a solid-phase enzyme-linked immunosorbent assay (ELISA) developed in the laboratory. Mesangial cells cultured for 1 week contained fibronectin as the most abundant ECM protein, followed by laminin and type IV collagen. Type IV collagen was seen only after the cells had piled up into 'hillocks' (approximately 4 weeks of continuous growth without passaging). After 4 weeks in 30 mmol/l glucose, mesangial cells contained increased amounts of all three matrix proteins. Fibronectin and laminin were increased by approximately 60%, while type IV collagen was increased 50%. Cells subcultured in medium containing 30 mmol/l glucose for 8 months displayed a twofold increase in fibronectin and laminin. Thus, high glucose per se can cause changes in mesangial cell ECM. This cell culture model should be useful in elucidating the mechanisms involved.