Regulation of ICAM-1/CD54 expression on human endothelial cells by hydrogen peroxide involves inducible NO synthase.

Expression of the inducible isoform of nitric oxide synthase (iNOS) is stimulated by cytokines in human epithelial cells. This work indicates that incubation of human umbilical cord endothelial cells with combinations of interleukin-1beta, tumor necrosis factor alpha, and interferon-gamma stimulated the synthesis of iNOS mRNA, as detected by reverse transcriptase-polymerase chain reaction. It is important to note that 50, 100, and 200 microM hydrogen peroxide was able to stimulate iNOS directly. Furthermore, 100 microM H2O2 enhanced synthesis of the oxidation products, nitrite (NO2-) and nitrate (NO3-) at 12 and 36 h. iNOS protein, detected by Western blot analysis, as well as L-citrulline levels, were also increased. When endothelial cell monolayers were incubated for 1 h with 100 microM H2O2 and subsequently with cytokines, iNOS mRNA was further augmented. Under the same conditions, we regularly observed an inhibition (25%) of intercellular adhesion molecule-1 (ICAM-1/CD54) expression. The latter was reversed when the NOS inhibitor N(G)-monomethyl-L-arginine was added, as shown by flow cytometry. These data suggest a specific effect of endogenous hydroperoxides on the biosynthesis and processing of the human endothelial iNOS isoform. We propose that H2O2 induces a temporary NO-dependent modulation of adhesion molecule expression to limit the tissue destruction that accompanies the vascular recruitment of leukocytes.

Naftidrofuryl-driven regulation of endothelial ICAM-1 involves nitric oxide.

Naftidrofuryl is a selective inhibitor of the 5-HT2 receptor expressed on human endothelial cells. This drug has been used over the years to cope with cerebral or peripheral ischemic accidents; however, no clear mechanism of action of this molecule has been highlighted to explain its vascular effects. In the present work, we demonstrate that the involvement of nitric oxide can account for the effects of naftidrofuryl. Indeed, naftidrofuryl potently inhibited the TNF-alpha-triggered increase of intercellular adhesion molecule-1 (ICAM-1) expression as well as stress fiber formation in human umbilical vein endothelial cells (HUVEC). Moreover, naftidrofuryl induced the expression of type II nitric oxide synthase (NOS II) messenger and protein, leading to a noticeable increase in nitric oxide synthesis. Furthermore, using the specific NOS II inhibitor 1400W, we verified that the observed effects of naftidrofuryl were NOS II-dependent. The biology of nitric oxide accounts for the reduction of the vasospasm associated with stroke and the strong inhibition of platelet aggregation. In conclusion, our work provides evidence for the inhibition of leukocyte recruitment by downregulation of CD54/ICAM-1, an additional key factor to be dealt with during thrombotic accidents. Importantly, it also highlights a novel NOS II-dependent mechanism of action for naftidrofuryl.