Functional role and species-specific contribution of arginases in pulmonary fibrosis.
Lung fibrosis is characterized by increased deposition of ECM, especially collagens, and enhanced proliferation of fibroblasts. l-arginine is a key precursor of nitric oxide, asymmetric dimethylarginine, and proline, an amino acid enriched in collagen. We hypothesized that l-arginine metabolism is altered in pulmonary fibrosis, ultimately affecting collagen synthesis. Expression analysis of key enzymes in the arginine pathway, protein arginine methyltransferases (Prmt), arginine transporters, and arginases by quantitative (q) RT-PCR and Western blot revealed significant upregulation of arginase-1 and -2, but not Prmt or arginine transporters, during bleomycin-induced pulmonary fibrosis in mice. HPLC revealed a concomitant, time-dependent decrease in pulmonary l-arginine levels. Arginase-1 and -2 mRNA and protein expression was increased in primary fibroblasts isolated from bleomycin-treated mice, compared with controls, and assessed by qRT-PCR and Western blot analysis. TGF-beta1, a key profibrotic mediator, induced arginase-1 and -2 mRNA expression in primary and NIH/3T3 fibroblasts. Treatment of fibroblasts with the arginase inhibitor, NG-hydroxy-l-arginine, attenuated TGF-beta1-stimulated collagen deposition, but not collagen mRNA expression or Smad signaling, in fibroblasts. In human lungs derived from patients with idiopathic pulmonary fibrosis, arginase activity was unchanged, but arginase-1 expression significantly decreased when compared with donor lungs. Our results thus demonstrate that arginase-1 is expressed and functionally important for collagen deposition in lung fibroblasts. TGF-beta1-induced upregulation of arginase-1 suggests an interplay between profibrotic agents and l-arginine metabolism during the course of lung fibrosis in the mouse, whereas species-specific regulatory mechanisms may account for the differences observed in mouse and human.
Kitowska
Kamila
K
Department of Medicine II, Univ. of Giessen School of Medicine, Aulweg 123, D-35392 Giessen, Germany.
Zakrzewicz
Dariusz
D
Königshoff
Melanie
M
Chrobak
Izabella
I
Grimminger
Friedrich
F
Seeger
Werner
W
Bulau
Patrick
P
Eickelberg
Oliver
O
D016475:3T3 Cells; D000328:Adult; D000818:Animals; D001119:Arginase / Q000235:genetics / Q000378:metabolism*; D001761:Bleomycin / Q000627:therapeutic use; D017931:DNA Primers; D005260:Female; D006801:Humans; D008168:Lung / Q000201:enzymology / Q000473:pathology; D008297:Male; D051379:Mice; D008875:Middle Aged; D011484:Protein-Arginine N-Methyltransferases / Q000235:genetics / Q000378:metabolism; D011658:Pulmonary Fibrosis / Q000188:drug therapy / Q000201:enzymology* / Q000473:pathology; D012313:RNA / Q000235:genetics / Q000302:isolation & purification; D012016:Reference Values; D020133:Reverse Transcriptase Polymerase Chain Reaction
D017931:DNA Primers; D001761:Bleomycin; D012313:RNA; D011484:Protein-Arginine N-Methyltransferases; D001119:Arginase