Abstract
We have shown that neurokinin A-induced contraction of human sigmoid circular muscle (HSCM) is reduced in patients with ulcerative colitis and that interleukin (IL)-1β may play a role in this change. We now examine changes in the signal transduction pathway mediating neurokinin A-induced contraction of HSCM and explore the role of IL-1β and of H2O2 in these changes. In Fura 2-AM-loaded ulcerative colitis HSCM cells, neurokinin A- and caffeine-induced peak Ca2+ increase and cell shortening were significantly reduced. In normal cells, neurokinin A-induced contraction was decreased by protein kinase C inhibitor chelerythrine and by calmodulin inhibitor CGS9343B [1,3-dihydro-1-[1-[(4-methyl-4H,6H-pyrrolo[1,2-a][4,1]-benzoxazepin-4-yl)methyl]-4-piperidinyl]-2H-benzimidazol-2-one (1:1) maleate]. In ulcerative colitis muscle cells, contraction was inhibited only by chelerythrine but not by CGS9343B. IL-1β treatment of normal HSCM strips and cells reproduced the changes observed in ulcerative colitis. IL-1β-induced reduction in caffeine-induced peak Ca2+ increase and contraction was reversed by catalase, suggesting a role of H2O2. IL-1β-induced H2O2 production was inhibited by mitogen-activated protein kinase (MAPK) kinase inhibitor PD98059 (2′-amino-3′-methoxyflavone) and by cytosolic phospholipase A2 (cPLA2) inhibitor AACOCF3 (arachidonyltrifluoromethyl ketone), but neither by p38 MAPK inhibitor SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole] nor by nuclear factor-κB (NF-κB) inhibitory peptide NF-κB SN50 (H-Ala-Ala-Val-Ala-Leu-Leu-Pro-Ala-Val-Leu-Leu-Ala-Leu-Leu-Ala-Pro-Val-Gln-Arg-Lys-Arg-Gln-Lys-Leu-Met-Pro-OH). IL-1β significantly increased the phosphorylation of extracellular signal-regulated kinase 1 (ERK1)/ERK2 MAPKs and cPLA2 and IL-1β-induced cPLA2 phosphorylation was blocked by PD98059. We conclude that Ca2+ stores of HSCM cells may be reduced in ulcerative colitis and that the signal transduction pathway of neurokinin A-induced contraction switches from calmodulin- and protein kinase C-dependent in normal cells to protein kinase C-dependent in ulcerative colitis cells. IL-1β reproduces these changes, possibly by production of H2O2 via sequential activation of MAPKs (ERK1/ERK2) and cPLA2.
Footnotes
-
Tis work was supported by Lifespan Research Funds (to W.C.) and National Institute of Diabetes and Digestive and Kidney Diseases Grant R21 DK62775-01 (to W.C.). These data were presented in part at the 104th annual meeting of the American Gastroenterological Association, in Orlando, FL, in May 2003.
-
doi:10.1124/jpet.104.068023.
-
ABBREVIATIONS: NK, neurokinin; IL, interleukin; IL-1ra, IL-1 receptor antagonist; CGS9343B, 1,3-dihydro-1-[1-[(4-methyl-4H,6H-pyrrolo[1,2-a][4,1]-benzoxazepin-4-yl)methyl]-4-piperidinyl]-2H-benzimidazol-2-one (1:1) maleate; BAPTA, 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid; MAPK, mitogen-activated protein kinase; MEK, mitogen-activated protein kinase kinase; PD98059, 2′-amino-3′-methoxyflavone; SB203580, 4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole; cPLA2, cytosolic phospholipase A2; AACOCF3, arachidonyltrifluoromethyl ketone; NF-κB, nuclear factor-κB; SN50, H-Ala-Ala-Val-Ala-Leu-Leu-Pro-Ala-Val-Leu-Leu-Ala-Leu-Leu-Ala-Pro-Val-Gln-Arg-Lys-Arg-Gln-Lys-Leu-Met-Pro-OH; ERK, extracellular signal-regulated kinase; ANOVA, analysis of variance.
- Received March 8, 2004.
- Accepted June 17, 2004.
- The American Society for Pharmacology and Experimental Therapeutics
JPET articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|