Lysophosphatidic acid stimulates protein kinase C isoforms alpha, beta, epsilon, and zeta in a pertussis toxin sensitive pathway in vascular smooth muscle cells.
The natural phospholipid lysophosphatidic acid (LPA) has been characterized as an important vascular smooth muscle cell (VSMC) mitogen whose effects are mainly mediated by pertussis toxin (PTX)-sensitive guanosine triphosphate (GTP)-binding protein (Gi-protein). Protein kinase C (PKC) isoforms play an important role in intracellular signaling cascades and in growth of VSMC. In the present study we investigated the effect of LPA on activation of PKC isoforms alpha, beta, epsilon, and zeta in VSMC by Western blot of cytosolic and membrane fractions. Furthermore, we examined the role of PKC activation on LPA-induced growth of VSMC using PKC inhibitor 19-27. Stimulation of VSMC by 5 microg/mL LPA for 10 min increased the amount of PKC alpha, beta, epsilon, and zeta in the particulate fraction by 689%, 285%, 424%, and 510%, respectively, and returned to control level after 30 min. Correspondingly, the amount of PKC alpha, beta, epsilon, and zeta in the cytosolic fraction decreased by 32%, 94%, 44%, and 95%, respectively, compared to control. Furthermore, we could show that LPA-induced activation of PKC alpha, beta, epsilon, and zeta isoforms was PTX sensitive. Incubation of VSMC with nonspecific PKC inhibitor 19-27 (10 micromol/L) for 24 h resulted in a 30% inhibition of LPA-induced DNA synthesis as measured by [3H]thymidine incorporation. In conclusion, in VSMC LPA stimulated translocation of PKC isoforms alpha, beta, epsilon, and zeta in a PTX-sensitive manner. Furthermore stimulation of PKC might be critically involved in LPA-induced mitogenesis in VSMC.
Seewald
S
S
Medizinische Universitäts-Poliklinik, Bonn, Germany.
Schmitz
U
U
Seul
C
C
Ko
Y
Y
Sachinidis
A
A
Vetter
H
H
D000818:Animals; D001013:Aorta, Thoracic / Q000166:cytology / Q000187:drug effects / Q000201:enzymology; D015153:Blotting, Western; D003600:Cytosol / Q000201:enzymology; D004247:DNA / Q000096:biosynthesis / Q000187:drug effects; D004591:Electrophoresis, Polyacrylamide Gel; D007527:Isoenzymes / Q000378:metabolism; D008246:Lysophospholipids / Q000494:pharmacology*; D009131:Muscle, Smooth, Vascular / Q000187:drug effects* / Q000201:enzymology; D037342:Pertussis Toxin; D011493:Protein Kinase C / Q000378:metabolism*; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051744:Protein Kinase C-epsilon; D051381:Rats; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*; D010566:Virulence Factors, Bordetella / Q000494:pharmacology*; D000099221:Protein Kinase C zeta
D004247:DNA; D007527:Isoenzymes; D008246:Lysophospholipids; D037342:Pertussis Toxin; D011493:Protein Kinase C; D064546:Protein Kinase C beta; D051571:Protein Kinase C-alpha; D051744:Protein Kinase C-epsilon; D010566:Virulence Factors, Bordetella; D000099221:Protein Kinase C zeta; C497257:Prkce protein, rat