kt-5720 and herbimycin

The effects of pretreatment with protein kinase C and protein kinase A inhibitors on the intraventricular insulin-induced attenuation of the antinociceptive effect of [D-Ala2, N-MePhe4, Gly-ol5]enkephalin (DAMGO) were studied in mice. Intracerebroventricular (i.c.v.) pretreatment with insulin dose- and time-dependently attenuated the antinociceptive effect of i.c.v. DAMGO (5.6 ng) in mice. Intracerebroventricular pretreatment with a highly selective tyrosine kinase inhibitor, herbimycin A, at doses of 200 and 600 ng for 70 min, dose-dependently reversed the attenuation of the antinociceptive effect of DAMGO (5.6 ng, i.c.v.) caused by insulin. Furthermore, i.c.v. pretreatment with serine/threonin kinase inhibitor, 1-(5-isoquinolinesulfonyl)-2-methylpiperazine hydrochloride (H7), at doses of 3-30 nmol for 60 min, dose-dependently reversed the attenuation of the antinociceptive effect of DAMGO (5.6 ng, i.c.v.) caused by insulin. Intracerebroventricular pretreatment with selective protein kinase C inhibitor, calphostin C, at doses of 1 and 3 pmol for 60 min, but not with a highly protein kinase A inhibitor, (8R, 9S, 11S)-(-)-9-hydroxy-9-n-hexyloxy-carbonyl-8-methyl-2, 3, 9, 20-tetrahydro-8, 11-epoxy-1H, 8H, 11H-2, 7b, 11a-triaqzadibenzo[a, g]cycloocta[c, d, e]-trinden-1-one (KT5720), at dose of 10 pmol for 60 min, reversed the attenuation of the antinociceptive effect of DAMGO (5.6 ng, i.c.v.) caused by insulin. These results suggest that the reduction of DAMGO-induced antinociception by insulin in mice may be, in part, due to the activation of protein kinase C followed by the activation of tyrosine kinase.

Topics: 1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; Analgesics, Opioid; Animals; Benzoquinones; Carbazoles; Cyclic AMP-Dependent Protein Kinases; Dose-Response Relationship, Drug; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalins; Enzyme Inhibitors; Hypoglycemic Agents; Indoles; Injections, Intraventricular; Insulin; Lactams, Macrocyclic; Male; Mice; Mice, Inbred ICR; Naphthalenes; Pain Measurement; Protein Kinase C; Pyrroles; Quinones; Rifabutin; Time Factors

Several protein kinase inhibitors (PKIs) were investigated for their effects on IL-1 beta, TNF alpha and PMA-induced IL-8 production from human umbilical vein endothelial cells (HUVEC). IL-1 beta (ED50 0.07 ng/ml), TNF alpha (ED50 100 ng/ml) and PMA (ED50 20 ng/ml) induced IL-8 production that could be detected as early as 2 h following stimulation. Staurosporine, a potent but non-specific inhibitor of protein kinases, inhibited PMA-induced (IC50 2 nM) but not IL-1 beta or TNF alpha (IC50 > 200 nM) induced IL-8 production. Neither the cAMP-dependent PKI, KT5720, nor the tyrosine PKIs, genistein, tyrphostin (1-100 microM) or lavendustin A (0.0001-1 microM), inhibited IL-8 production elicited by IL-1 beta. However, the macrolide protein kinase inhibitor geldanamycin (IC50 = 30 nM), but not the closely related analog herbimycin A (5-500 nM), inhibited IL-8 production by 60%. Northern blot analysis of IL-8 mRNA revealed that staurosporine suppressed mRNA increase following stimulation by PMA but not by IL-1. It is proposed that a novel protein kinase susceptible to geldanamycin inhibition may be involved in IL-1-mediated signal transduction.

Topics: Alkaloids; Benzoquinones; Carbazoles; Cells, Cultured; Endothelium, Vascular; Humans; Indoles; Interleukin-1; Interleukin-8; Lactams, Macrocyclic; Protein Kinase Inhibitors; Pyrroles; Quinones; Recombinant Proteins; Rifabutin; Staurosporine; Tetradecanoylphorbol Acetate; Tumor Necrosis Factor-alpha; Umbilical Veins