Overexpression of 12-lipoxygenase causes cardiac fibroblast cell growth.
Evidence suggests that leukocyte type 12-lipoxygenase (12-LO) plays an important role in cell growth. However, the role of 12-LO in cardiac cell growth has not been tested. We have now stably overexpressed 12-LO cDNA in rat fetal cardiac fibroblasts to evaluate the role of the 12-LO pathway in cardiac cell growth. Overexpression of 12-LO increased cell [(3)H]leucine incorporation by 2.1+/-0.1-fold (P<0.01) and cell protein content by 2.2+/-0. 3-fold (P<0.01) over mock-transfected cells. These findings were confirmed in additional clones. Baicalein, a 12-LO enzyme inhibitor, dose-dependently inhibited serum-induced leucine incorporation in cardiac fibroblast cells as well as partially inhibited leucine incorporation in cells overexpressing 12-LO. 12-LO overexpression also caused cell [(3)H]thymidine incorporation to increase by 3.4+/-0.3-fold (P<0.01). Cell flow cytometry analysis showed that the size of 12-LO-overexpressing cells was markedly enlarged compared with that of mock-transfected cells. The fibronectin content of the 12-LO-overexpressing cardiac fibroblasts was also significantly increased. We next evaluated the effects of 12-LO RNA overexpression on kinase pathways linked to cellular growth. The overexpression of 12-LO enhanced extracellular signal-regulated kinase activity (4. 1+/-0.5-fold), c-Jun NH(2)-terminal kinase activity (2.9+/-0.5-fold), and p38 mitogen-activated protein kinase activity (2.2+/-0.3-fold). Pretreatment with SB202190 (100 nmol/L), a specific inhibitor of p38, prevented the increases in protein content of 12-LO-overexpressing cardiac fibroblast cells. These data clearly demonstrate that the overexpression of 12-LO causes cell growth of cardiac fibroblasts, thus supporting the role of 12-LO as a novel growth-promoting pathway in the heart.
Wen
Y
Y
Division of Endocrinology and Metabolism, University of Virginia Health Sciences Center (Charlottesville), USA.
Gu
J
J
Liu
Y
Y
Wang
P H
PH
Sun
Y
Y
Nadler
J L
JL
D000818:Animals; D001092:Arachidonate 12-Lipoxygenase / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D002455:Cell Division / Q000235:genetics; D048429:Cell Size / Q000235:genetics; D002478:Cells, Cultured; D004274:DNA, Recombinant / Q000235:genetics / Q000378:metabolism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005333:Fetus; D005347:Fibroblasts / Q000166:cytology* / Q000201:enzymology; D005353:Fibronectins / Q000378:metabolism; D044950:Flavanones; D005419:Flavonoids / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D007930:Leucine / Q000187:drug effects / Q000378:metabolism; D007962:Leukocytes / Q000201:enzymology; D016859:Lipoxygenase Inhibitors; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D009206:Myocardium / Q000166:cytology*; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D013936:Thymidine / Q000378:metabolism; D014162:Transfection; D014316:Tritium
D004274:DNA, Recombinant; D004791:Enzyme Inhibitors; D005353:Fibronectins; D044950:Flavanones; D005419:Flavonoids; D007093:Imidazoles; D016859:Lipoxygenase Inhibitors; D011725:Pyridines; D014316:Tritium; C006680:baicalein; D001092:Arachidonate 12-Lipoxygenase; D020928:Mitogen-Activated Protein Kinases; D007930:Leucine; C090942:4-(4-fluorophenyl)-2-(4-hydroxyphenyl)-5-(4-pyridyl)imidazole; D013936:Thymidine