title: C(2)-ceramide-induced circular smooth muscle cell contraction involves PKC-epsilon and p44/p42 MAPK activation in cat oesophagus. Mitogen-activated protein kinase.
abstract: We investigated the mechanism of C(2)-ceramide (C(2))-induced circular smooth muscle cell contraction in cat oesophagus. C(2) produced contraction of smooth muscle cells isolated by enzymatic digestion, peaked at 30 s and was sustained at a plateau at 5 min. The response to C(2) was concentration-dependent. H-7 or chelerythrine inhibited C(2)-induced contraction, while the diacylglycerol (DAG) kinase inhibitor, R59949, had no effect, suggesting that the contraction is protein kinase C (PKC) pathway-dependent. To test if PKC-mediated contraction may be isozyme-specific, we examined the effects of PKC isozymes antibodies on contraction. PKC-epsilon antibody inhibited the contraction by C(2) but not by PKC-betaII or -gamma, suggesting that PKC-epsilon mediates the contraction by C(2). To characterize the specific PKC isozymes that mediate contraction of the smooth muscle cells, we used, as an inhibitor, N-myristoylated peptides (myr-PKC) derived from the pseudosubstrate sequences of PKC-(alpha)(beta)(gamma), -alpha, -delta, or -epsilon. myr-PKC-epsilon only inhibited the contraction, which was concentration-dependent, suggesting that PKC-epsilon isozyme is involved in the contraction. To examine which mitogen-activated protein kinases (MAPKs) are involved in C(2)-induced contraction, specific MAPK inhibitors (MEK inhibitor, PD98059, and p38 MAPK inhibitor, SB202190) are used. Preincubation of PD98059 blocked the contraction induced by C(2) in a concentration-dependent manner. However, SB202190 had no effects on contraction. C(2) increased the intensity of the bands identified by phosphospecific p44/p42 MAPK antibody and preincubation of PD98059 decreased the intensity of bands as compared with C(2)-stimulated cells. In conclusion, C(2) produced the contraction of smooth muscle cells of cat oesophagus. The contraction is mediated by PKC-epsilon, resulting in the activation of p44/p42 MAPK.
authors.affiliation: Department of Pharmacology, College of Pharmacy, Chung Ang University, Seoul, South Korea.
authors.forename: Chang Yell
authors.initials: CY
authors.lastname: Shin
authors.forename: Yul Pyo
authors.initials: YP
authors.lastname: Lee
authors.forename: Tai Sang
authors.initials: TS
authors.lastname: Lee
authors.forename: Hyun Ju
authors.initials: HJ
authors.lastname: Song
authors.forename: Uy Dong
authors.initials: UD
authors.lastname: Sohn
mesh_terms: D000818:Animals; D000906:Antibodies / Q000494:pharmacology; D002415:Cats; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004947:Esophagus / Q000187:drug effects / Q000201:enzymology*; D007527:Isoenzymes / Q000187:drug effects / Q000378:metabolism; D020935:MAP Kinase Signaling System / Q000187:drug effects / Q000502:physiology; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects / Q000201:enzymology*; D010455:Peptides / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D051744:Protein Kinase C-epsilon; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000378:metabolism* / Q000494:pharmacology
chemical_list: D000906:Antibodies; D007527:Isoenzymes; C064769:N-acetylsphingosine; D010455:Peptides; D011493:Protein Kinase C; D051744:Protein Kinase C-epsilon; D048052:Mitogen-Activated Protein Kinase 3; D020928:Mitogen-Activated Protein Kinases; D013110:Sphingosine