HIGH GLUCOSE INHIBITS NITRIC-OXIDE PRODUCTION IN CULTURED RAT MESANGIAL CELLS

Hyperglycemia directly contributes to the development of diabetic neph ropathy. A high-serum glucose concentration alters intraglomerular hem odynamics and promotes deposition of extracellular matrix in the kidne y. Nitric oxide (NO) is a short-lived messenger molecule that particip ates in the regulation of renal blood flow, GFR, and mesangial matrix accumulation. Therefore, in this study it was tested whether high gluc ose directly modulates NO synthesis by rat mesangial cells in vitro by measuring the accumulation of nitrite, the stable metabolite of NO, i n the incubation media. Raising the external glucose concentration to 33.3 mM for 24 to 72 h reduced nitrite levels in cell supernatants in a time-dependent manner to a nadir of 14 +/- 3% of the amount in norma l glucose media (5.6 mM) (P < 0.01). The decline in NO synthesis in hi gh glucose media was paralleled by decreased cyclic guanosine monophos phate generation; however, there was no alteration in rat mesangial ce ll expression of inducible NO synthase protein. The suppressive effect of high glucose on NO production by mesangial cells was not modified by inhibition of protein kinase C (H-7), the addition of antioxidants (vitamin E or superoxide dismutase), or a pan-specific anti-transformi ng growth factor-beta antibody. An elevated ambient glucose caused a t ime-dependent reduction in mesangial cell L-arginine content. Addition of L-arginine (10 to 20 mM) to external media partially reversed the inhibitory effect of high glucose on mesangial cell NO production in a dose-dependent manner. The highest dose of L-arginine (20 mM) increas ed mesangial cell L-arginine content to comparable levels in normal an d high glucose media. These results indicate that high glucose causes depletion of L-arginine in mesangial cells and compromises NO synthesi s. Limitation in the metabolic precursor and other, as yet unidentifie d, factors act to reduce NO production by mesangial cells in the prese nce of an elevated ambient glucose level, a change that may play a rol e in the development of diabetic glomerulosclerosis.