Role of protein kinase C and cyclic adenosine monophosphate in the regulation of tissue-type plasminogen activator, plasminogen activator inhibitor-1, and platelet-derived growth factor mRNA levels in human endothelial cells. Possible involvement of proto-oncogenes c-jun and c-fos.
Activation of protein kinase C leads to a strong induction of tissue-type plasminogen activator (t-PA) expression in endothelial cells. Using endothelial cells from human umbilical vein (HUVECs) and human aorta (HAECs), we have studied this regulation of t-PA and its inhibitor, plasminogen activator inhibitor-1 (PAI-1), at the mRNA level and have compared their induction with the expression of platelet-derived growth factors A and B (PDGF-A and PDGF-B) and the proto-oncogenes c-jun and c-fos. Treatment of HUVECs with exogenous bacterial phospholipase C or the synthetic diacylglycerol 1-oleoyl-2-acetylglycerol led to a threefold and a twofold increase, respectively, in t-PA concentrations in 24-hour-conditioned medium. Similarly, the more stable protein kinase C activator 4 beta-phorbol-12-myristate-13-acetate (PMA) caused about a 10-fold increase in t-PA antigen levels. This effect of PMA is maximal between 8 and 16 hours at a concentration of 10 nM and is fully accounted for by parallel increases in t-PA mRNA levels. An increase in intracellular cyclic adenosine monophosphate levels by forskolin (10 microM) slightly diminished t-PA expression but further enhanced the PMA-induced increases in t-PA synthesis and mRNA levels by at least twofold. PMA also enhanced the mRNA levels of two other important endothelium-expressed genes, PDGF-A and PDGF-B, with a time profile similar to that of t-PA, with peak values about fivefold higher than control values. Forskolin did not further stimulate this PMA-induced PDGF expression in HUVECs, which suggests a regulatory mechanism different from that of t-PA. Qualitatively very similar induction patterns of t-PA, PDGF-A, and PDGF-B were seen with HAECs. In contrast to t-PA and PDGF, PAI-1 mRNA and antigen levels increased only slightly after PMA treatment of HUVECs or HAECs; forskolin alone or in combination with PMA diminished the expression of PAI-1. The induction of t-PA mRNA by PMA was dependent on protein synthesis and was preceded by a strong transient increase in c-jun and c-fos mRNA levels; the induction of c-fos but not of c-jun was potentiated by forskolin. Because the products of these two proto-oncogenes form dimeric complexes for which specific binding sites are present in the t-PA promoter region, they may mediate the protein kinase C-dependent increase in t-PA gene expression, including the stimulating action of cyclic adenosine monophosphate.
Kooistra
T
T
Gaubius Laboratory IVVO-TNO, Leiden, The Netherlands.
Bosma
P J
PJ
Toet
K
K
Cohen
L H
LH
Griffioen
M
M
van den Berg
E
E
le Clercq
L
L
van Hinsbergh
V W
VW
D001011:Aorta / Q000187:drug effects; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000502:physiology*; D015342:DNA Probes; D004268:DNA-Binding Proteins / Q000378:metabolism*; D004075:Diglycerides / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D015849:Plasminogen Inactivators / Q000378:metabolism; D010982:Platelet-Derived Growth Factor / Q000378:metabolism; D011493:Protein Kinase C / Q000502:physiology*; D011518:Proto-Oncogene Proteins / Q000378:metabolism*; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D012333:RNA, Messenger / Q000378:metabolism; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D010959:Tissue Plasminogen Activator / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D010738:Type C Phospholipases / Q000494:pharmacology; D014471:Umbilical Veins / Q000187:drug effects
D015342:DNA Probes; D004268:DNA-Binding Proteins; D004075:Diglycerides; D015849:Plasminogen Inactivators; D010982:Platelet-Derived Growth Factor; D011518:Proto-Oncogene Proteins; D016760:Proto-Oncogene Proteins c-fos; D016755:Proto-Oncogene Proteins c-jun; D012333:RNA, Messenger; D014157:Transcription Factors; D005576:Colforsin; C038237:1-oleoyl-2-acetylglycerol; D000242:Cyclic AMP; D011493:Protein Kinase C; D010738:Type C Phospholipases; D010959:Tissue Plasminogen Activator; D013755:Tetradecanoylphorbol Acetate