Sphingosine 1-phosphate-induced signal transduction in cat esophagus smooth muscle cells.
We investigated the mechanism of contraction induced by S1P in esophageal smooth muscle cells. Western blot analysis demonstrated that S1P(1), S1P(2), S1P(3), and S1P(5) receptors existed in the cat esophagus. Only penetration of EDG-5 (S1P(2)) antibody into permeabilized cells inhibited S1P-induced contraction. Pertussis toxin (PTX) also inhibited contraction, suggesting that it was mediated by S1P(2) receptors coupled to a PTX-sensitive G(i) protein. Specific antibodies to G(i2), G(q) and G(beta) inhibited contraction, implying that the S1P-induced contraction depends on PTX-insensitive G(q) and G(beta) dimers as well as the PTX-sensitive G(i2). Contraction was not affected by the phospholipase A2 inhibitor DEDA, or the PLD inhibitor rho-chloromer-curibenzoate, but it was abolished by the PLC inhibitor U73122. Incubation of permeabilized cells with PLCb3 antibody also inhibited contraction. Contraction involved the activation of a PKC pathway since it was affected by GF109203X and chelerythrine. Since PKCepsilon antibody inhibited contraction, PKCe may be required. Preincubation of the muscle cells with the MEK inhibitor PD98059 blocked S1P-induced contraction, but the p38 MAP kinase inhibitor SB202190 did not. In addition, co-treatment of cells with GF 109203X and PD98059 did not have a synergistic effect, suggesting that these two kinases are involved in the same signaling pathway. Our data suggest that S1P-induced contraction in esophageal smooth muscle cells is mediated by S1P(2) receptors coupled to PTX-sensitive G(i2) proteins, and PTX-insensitive G(q) and G(beta) proteins, and that the resulting activation of the PLCb3 and PKCepsilon pathway leads to activation of a p44/p42 MAPK pathway.
Song
Hyun Ju
HJ
Department of Pharmacology, College of Pharmacy, Chung Ang University, Seoul 156-756, Korea.
Choi
Tai Sik
TS
Chung
Fa Yong
FY
Park
Sun Young
SY
Ryu
Jung Soo
JS
Woo
Jae Gwang
JG
Min
Young Sil
YS
Shin
Chang Yell
CY
Sohn
Uy Dong
UD
D000818:Animals; D002415:Cats; D004305:Dose-Response Relationship, Drug; D004947:Esophagus / Q000166:cytology*; D020962:Heterotrimeric GTP-Binding Proteins / Q000378:metabolism; D008246:Lysophospholipids / Q000494:pharmacology*; D020930:MAP Kinase Kinase Kinases / Q000037:antagonists & inhibitors; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D008954:Models, Biological; D009119:Muscle Contraction / Q000187:drug effects; D032389:Myocytes, Smooth Muscle / Q000187:drug effects* / Q000378:metabolism*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D049349:Receptors, Lysosphingolipid / Q000378:metabolism; D015398:Signal Transduction / Q000187:drug effects*; D013110:Sphingosine / Q000031:analogs & derivatives* / Q000494:pharmacology; D013997:Time Factors; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors
D008246:Lysophospholipids; D049349:Receptors, Lysosphingolipid; C060506:sphingosine 1-phosphate; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D020930:MAP Kinase Kinase Kinases; D010738:Type C Phospholipases; D020962:Heterotrimeric GTP-Binding Proteins; D013110:Sphingosine