Possible role of the protein kinase C/CPI-17 pathway in the augmented contraction of human myometrium after gestation.
1. Activation of protein kinase C (PKC) by phorbol 12,13-dibutylate (PDBu, 1 microm) induced sustained contractions with no increase in [Ca2+]i in nonpregnant and pregnant human myometria. The contractile effects of PDBu in pregnant myometrium were much greater than those in nonpregnant myometrium, and the contractions in pregnant myometrium were accompanied by an increase in myosin light chain (MLC) phosphorylation at Ser19. 2. The contraction induced by PDBu in pregnant myometrium was inhibited by the inhibitors of conventional PKC isoforms, bisindolylmaleimides and indolocarbazole, such as Go6976, Go6983, and Go6850 (1 microM). LY333531 (1 microM), a specific inhibitor of PKC beta, also inhibited the PDBu-induced contraction in the pregnant myometrium. 3. In the pregnant myometrium permeabilized with alpha-toxin, PDBu increased the contractions induced at fixed Ca2+ concentration (0.3 microM) both in nonpregnant and pregnant myometria, indicating Ca2+ sensitization of contractile elements. 4. Western immunoblot analysis indicated that pregnant myometrium contained PKC isozymes such as conventional PKC (alpha, beta, gamma), novel PKC (delta, epsilon, theta), and atypical PKC (zeta but not iota and lambda). RT-PCR and real-time RT-PCR analysis indicated that, among the conventional PKC, the levels of mRNA of beta isoform in pregnant human myometrium were greater than those in nonpregnant myometrium. 5. CPI-17 is a substrate for PKC, and the phosphorylated CPI-17 is considered to inhibit myosin phosphatase. The levels of CPI-17 mRNA and protein expression were also greater in the pregnant myometrium. 6. These results suggest that the PKC-mediated contractile mechanism is augmented in human myometrium after gestation, and that this augmentation may be attributable to the increased activity of the beta PKC isoform and CPI-17.
Ozaki
Hiroshi
H
Department of Veterinary Pharmacology, Graduate School of Agriculture and Life Sciences, The University of Tokyo, Bunkyo-ku, Yayoi 1-1-1, Tokyo 113-8657, Japan. aozaki@mail.ecc.u-tokyo.ac.jp
Yasuda
Katsuhiko
K
Kim
Yoon-Sun
YS
Egawa
Makoto
M
Kanzaki
Hideharu
H
Nakazawa
Hiroshi
H
Hori
Masatoshi
M
Seto
Minoru
M
Karaki
Hideaki
H
D000328:Adult; D015153:Blotting, Western; D002118:Calcium / Q000378:metabolism; D002227:Carbazoles / Q000494:pharmacology; D020536:Enzyme Activators / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006801:Humans; D007211:Indoles / Q000494:pharmacology; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000378:metabolism; D008301:Maleimides / Q000494:pharmacology; D009119:Muscle Contraction / Q000187:drug effects* / Q000502:physiology; D009124:Muscle Proteins / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D009215:Myometrium / Q000187:drug effects / Q000201:enzymology* / Q000502:physiology; D018994:Myosin Light Chains / Q000187:drug effects; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010749:Phosphoprotein Phosphatases; D010750:Phosphoproteins / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011247:Pregnancy / Q000502:physiology*; D011263:Pregnancy Trimester, Third; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D012694:Serine / Q000378:metabolism; D010738:Type C Phospholipases / Q000494:pharmacology; D014590:Uterine Contraction / Q000187:drug effects*
C465664:2-(1-(3-dimethylaminopropyl)-5-methoxyindol-3-yl)-3-(1H-indol-3-yl)maleimide; D002227:Carbazoles; D020536:Enzyme Activators; D004791:Enzyme Inhibitors; D007211:Indoles; D047908:Intracellular Signaling Peptides and Proteins; D007527:Isoenzymes; D008301:Maleimides; D009124:Muscle Proteins; D018994:Myosin Light Chains; C107507:PPP1R14A protein, human; D010750:Phosphoproteins; D012333:RNA, Messenger; C081021:Go 6976; D015240:Phorbol 12,13-Dibutyrate; D012694:Serine; C099154:ruboxistaurin; D011493:Protein Kinase C; D010749:Phosphoprotein Phosphatases; D010738:Type C Phospholipases; C070515:bisindolylmaleimide I; D002118:Calcium