Cytokine-induced arginase activity in pulmonary endothelial cells is dependent on Src family tyrosine kinase activity.
We hypothesized that the Src family tyrosine kinases (STKs) are involved in the upregulation of arginase and inducible nitric oxide synthase (iNOS) expression in response to inflammatory stimuli in pulmonary endothelial cells. Treatment of bovine pulmonary arterial endothelial cells (bPAEC) with lipopolysaccharide and tumor necrosis factor-alpha (L/T) resulted in increased urea and nitric oxide (NO) production, and this increase in urea and NO production was inhibited by the STK inhibitor PP1 (10 microM). The STK inhibitors PP2 (10 microM) and herbimycin A (10 microM) also prevented the L/T-induced expression of both arginase II and iNOS mRNA in bPAEC. Together, the data demonstrate a central role of STK in the upregulation of both arginase II and iNOS in bPAEC in response to L/T treatment. To identify the specific kinase(s) required for the induction of urea and NO production, we studied human pulmonary microvascular endothelial cells (hPMVEC) so that short interfering RNA (siRNA) techniques could be employed. We found that hPMVEC express Fyn, Yes, c-Src, Lyn, and Blk and that the protein expression of Fyn, Yes, c-Src, and Lyn could be inhibited with specific siRNA. The siRNA targeting Fyn prevented the cytokine-induced increase in urea and NO production, whereas siRNAs specifically targeting Yes, c-Src, and Lyn had no appreciable effect on cytokine-induced urea and NO production. These findings support our hypothesis that inflammatory stimuli lead to increased urea and NO production through a STK-mediated pathway. Furthermore, these results indicate that the STK Fyn plays a critical role in this process.
Chang
Rossana
R
Center for Perinatal Research, Columbus, OH 43205, USA.
Chicoine
Louis G
LG
Cui
Hongmei
H
Kanagy
Nancy L
NL
Walker
Benjimen R
BR
Liu
Yusen
Y
English
B Keith
BK
Nelin
Leif D
LD
D000818:Animals; D001119:Arginase / Q000235:genetics*; D002417:Cattle; D016207:Cytokines / Q000494:pharmacology*; D017931:DNA Primers; D003907:Dexamethasone / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000187:drug effects / Q000502:physiology*; D004790:Enzyme Induction / Q000187:drug effects*; D009569:Nitric Oxide / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II / Q000235:genetics; D011652:Pulmonary Circulation / Q000187:drug effects / Q000502:physiology*; D012333:RNA, Messenger / Q000235:genetics; D034741:RNA, Small Interfering / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014508:Urea / Q000378:metabolism; D019061:src-Family Kinases / Q000502:physiology*
D016207:Cytokines; D017931:DNA Primers; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D009569:Nitric Oxide; D003907:Dexamethasone; D014508:Urea; D052247:Nitric Oxide Synthase Type II; D019061:src-Family Kinases; D001119:Arginase