Regulation of endothelial function by asymmetric dimethylarginine released by vascular smooth muscle cells during oxidative stress

We reported recently that preglomerular vascular smooth muscle cells stably transfected with p22phox (S‐p22phox SMC) released H2O2 and asymmetric dimethylarginine (ADMA) yet these cells lacked an ADMA target since they did not express nitric oxide synthase (NOS). Therefore, we tested the hypothesis that H2O2 and/or ADMA released from VSMCs during oxidative stress regulate endothelial cell (EC) function. Compared to conditioned media from wild type (Wt) SMCs, media from S‐p22phox SMC contained more H2O2 (177 ± 3.9% vs. 100 ± 7.6%; p< 0.001) and ADMA ( 357 ± 4 nmol · L–1 vs. 220 ± 7 nmol · L–1; p < 0.005). NOS Activity (conversion of 3H‐L‐arginine to 3H‐citrulline ) of human glomerular ECs cultured in conditioned media from S‐p22phox SMCs was reduced (2.8 ± 0.2% vs. 3.6 ± 0.4%, p<0.05) compared to Wt cell media. Proliferation of ECs was reduced by 36% by co‐culture with S‐p22phox vs. Wt SMCs at day 5 (176 ± 10 % vs. 274 ± 10 %; P<0.05) and by 46 ±4 % at day 7. Media from both types of SMCs had similar effects as co‐culture. Incubation of ECs for 72 hours with ADMA caused graded reductions in proliferation whereas H2O2 increased proliferation from 1 to 4 μM and deceased proliferation at >10 μM. These results disclosed novel signaling pathways in the blood vessel wall whereby ROS generated by NADPH oxidase in VSMCs may impact the function of ECs by generation of H2O2 and/or ADMA.