Potent inhibition of inducible nitric oxide synthase by geldanamycin, a tyrosine kinase inhibitor, in endothelial, smooth muscle cells, and in rat aorta

Abstract

We have examined whether specific protein tyrosine kinase (PTK) inhibitors (genistein, tyrphostin, or geldanamycin) prevent nitric oxide (NO') production in rat smooth muscle cells (SMC), in murine brain endothelial cells (MBE), and in isolated rat aortas treated with endotoxin (LPS) and/or cytokines. Tyrphostin failed to inhibit either the release of nitrite in both endothelial and smooth muscle cells or vascular hyporeactivity in rat aorta, caused by immunostimulants. Genistein decreased nitrite production in MBE only at high concentration but had no effect on nitrite production in SMC and on the hypocontractility in aortic rings. In contrast, low concentrations of geldanamycin abolished the release of nitrite in MBE and in SMC treated with endotoxin and/or cytokines. Geldanamycin inhibited also the hypocontractility to phenylephrine in aortic rings treated with LPS or interleukin-1. This inhibitor failed to inhibit the release of nitrite and the vascular hyporeactivity once nitric oxide synthase (NOS) was induced by immunostimulants whereas methyl-L-arginine, an inhibitor of NOS, had significant effects. These data suggest that LPS-and cytokines-induced NO" production initiate a common signaling pathway involving a PTK that is inhibited by geldanamycin but not or slightly by tyrphostin or genistein at a point that precedes the induction of NOS.

Transmembrane signalling mechanisms regulating expression of cationic amino acid transporters and inducible nitric oxide synthase in rat vascular smooth muscle cells

“…In agreement with published data [5,7,[9][10][11], our findings with the PTK inhibitors confirm that one or more PTKs are critical for induction of NO synthesis in smooth muscle cells. Unlike effects with RO318220, both genistein and herbimycin A caused significant concentration-dependent inhibition of accumulated nitrite at concentrations well below those found to be cytotoxic.…”

“…However, other studies have found no indication for PKC involvement in iNOS induction in RASMCs [6], mouse astrocytes [7] or human chrondocytes [8]. Increasing evidence now suggests that activation of PTK rather than PKC may be obligatory for induction of iNOS, not only in rat smooth muscle cells [5,6,9] but also in murine macrophages [5,10], astrocytes [7] and mesangial cells [11].…”

Heat shock protein 90 mediates macrophage activation by Taxol and bacterial lipopolysaccharide

“…Structural studies of Taxol's interaction with Hsps will be of interest to help explain how two antitumor drugs, geldanamycin and Taxol, can each bind specifically to Hsp 90 family members and exert predominantly inhibitory and predominantly stimulatory cellular actions, respectively. For example, geldanamycin blocks the action of endothelial nitric oxide synthase (49), whereas Taxol enhances the expression of inducible nitric oxide synthase (50). It will also be important to determine whether LPS interacts with Hsp 90 in the same manner as Taxol.…”

Involvement of JAK2 and MAPK on type II nitric oxide synthase expression in skin-derived dendritic cells

“…The PD-098059 was used to inhibit the MEK activation (8), and the p38 MAPK and JAK2 were inhibited with SB-203580 (36) and tyrphostin B42 (22), respectively. The concentrations of genistein (30 µM), PD-098059 (30 µM), SB-203580 (10 and 20 µM), and tyrphostin B42 (20 µM) were chosen based on the previously published studies (8,14,22,36), and the assay of cellular MTT reduction in the presence of the indicated concentrations of the compounds revealed the lack of a significant toxic effect (data not shown).…”

“…The intracellular signals that regulate the expression of iNOS have been studied in different cell types, and, although it has not been fully characterized yet, iNOS expression appears to be regulated in a cell-specific manner. Protein kinase C (PKC), protein tyrosine kinases (PTKs), and cAMP-dependent protein kinase have been found to be involved in the regulation of iNOS expression (14,17,19,23,(28)(29)(30). Activation of some of these kinases may stimulate the mitogenactivated protein kinases (MAPK), a family of structurally related kinases that are involved in cellular events, such as growth, differentiation, and stress responses (25).…”