Phytanic acid activates NADPH oxidase through transactivation of epidermal growth factor receptor in vascular smooth muscle cells.
BACKGROUND
Phytanic acid (PA) has been implicated in development of cancer and its defective metabolism is known to cause life-threatening conditions, such as Refsum disease, in children. To explore molecular mechanisms of phytanic acid-induced cellular pathology, we investigated its effect on NADPH oxidase (NOX) and epidermal growth factor receptor (EGFR) in rat aortic smooth muscle cells (RASMC).


METHODS
Smooth muscle cells were isolated from rat aortae using enzymic digestion with collagenase and elastase. Cultured RASMC were treated with varying concentrations (0.5-10 μg/ml) of phytanic acid in the presence/absence of fetal bovine serum (FBS) and/or EGFR inhibitor, AG1478. Following treatment with experimental agents, NOX activity was assayed in RASMC cultures by luminescence method. Gene expression of NOX-1 and p47phox was assessed using RT-PCR. NOX-1, p47phox and, total EGFR protein and its phosphorylated form were measured by Western blotting.


RESULTS
Treatment of RASMC with supraphysiological concentrations (>2.5 μg/ml) of PA significantly (p < 0.01) increased the NOX activity. PA also significantly increased gene/protein expression of NOX-1 and p47phox whereas p22phox and p67phox remained unaffected. Interestingly, PA (2.5-10 μg/ml) markedly (2-3 folds) increased the total and phosphorylated EGFR. Treatment of cells with EGFR inhibitor, AG1478, significantly blocked the PA-induced enhancement of NOX activity.


CONCLUSIONS
Our findings that PA transactivates EGFR and induces NOX activity in vascular smooth muscle cells provide new insights into molecular mechanisms of PA's role in cancer and Refsum disease.
Departments of Pediatrics, Faculty of Medicine, Kuwait University, Kuwait City, Kuwait. dhaunsig@hsc.edu.kw.
Gursev S
GS
Dhaunsi
Departments of Pediatrics, Faculty of Medicine, Kuwait University, Kuwait City, Kuwait.
Mayra
M
Alsaeid
Pharmacology and Toxicology, Faculty of Medicine, Kuwait University, Kuwait City, Kuwait.
Saghir
S
Akhtar
D000818:Animals; D001011:Aorta / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D002417:Cattle; D002469:Cell Separation / Q000379:methods; D003470:Culture Media / Q000737:chemistry / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000494:pharmacology; D066246:ErbB Receptors / Q000037:antagonists & inhibitors / Q000235:genetics* / Q000378:metabolism; D008297:Male; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D032389:Myocytes, Smooth Muscle / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D019255:NADPH Oxidases / Q000235:genetics* / Q000378:metabolism; D010750:Phosphoproteins / Q000235:genetics* / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D010831:Phytanic Acid / Q000494:pharmacology*; D061251:Primary Cell Culture; D011799:Quinazolines / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D015398:Signal Transduction; D015533:Transcriptional Activation / Q000187:drug effects; D020032:Tyrphostins / Q000494:pharmacology
D003470:Culture Media; D004791:Enzyme Inhibitors; D010750:Phosphoproteins; D011799:Quinazolines; D020032:Tyrphostins; C079789:neutrophil cytosol factor 67K; D010831:Phytanic Acid; C101044:RTKI cpd; D019255:NADPH Oxidases; C515768:Cyba protein, rat; C060505:neutrophil cytosolic factor 1; C483803:Egfr protein, rat; D066246:ErbB Receptors
Aortic smooth muscle; EGFR; NADPH oxidase; Phytanic acid