Methotrexate induces cell swelling and necrosis in renal tubular cells

Background The present study was carried out to investigate if methotrexate (MTX) has a direct lethal effect in renal tubular cells, and if so, to further clarify the mechanisms of cell death. Materials and Methods Renal tubular cells (LLC-PK1 cells) were incubated with MTX (0.01 µM, 0.1 µM, and 1 µM), either alone or in combination with 0.1 µM amiloride (Na+/H+ antiporter inhibitor) or 1 µM carbachol (M-cholinergic agonist). Cell viability was then determined by means of trypan blue (TB) exclusion tests and MTT assays. Results After 4 hr incubation with 0.1 µM MTX the number of viable cells was decreased by 18% in comparison with control cells, and the proportion of dead cells was increased by 38%. Cell death induced by MTX was time-dependent and did not show apoptotic features. On the contrary, cell swelling was discovered. This cell death was prevented by co-incubating the cells with amiloride or carbachol. Conclusions MTX induces cell swelling and cell death in renal tubular LLC-PK1 cells. The tubular cell death induced by MTX is time-dependent. Cell death can be prevented by co-incubating with amiloride, thus indicating that the Na+/H+ antiporter and possibly other volume regulatory factors in renal tubular cells are involved in MTX-induced renal failure. © 2005 Wiley-Liss, Inc.