The role of the Src family of tyrosine kinases after oxidant-induced lung injury in vivo.
BACKGROUND
Patients sustaining major trauma are predisposed to the development of organ dysfunction. We have shown that oxidant stress generated by hemorrhagic shock/resuscitation (S/R) in rodents increases lipopolysaccharide (LPS)-induced lung injury and translocation of nuclear factor kappa B (NF-kappaB) in alveolar macrophages (AMs). In addition, using a cellular model, we have shown that priming with oxidants reprograms LPS signaling through an Src-dependent pathway. In the present studies, we hypothesize that oxidant priming by S/R in vivo involves Src family kinases.


METHODS
Rats were bled to a mean arterial pressure of 40 mmHg and maintained for 1 hour, then resuscitated with shed blood and equal volume of Ringer's lactate. In some studies, animals received the antioxidant NAC (0.5 g/kg) or a Src family inhibitor, PP2 (0.1 or 0.2 mg/kg), before resuscitation. LPS was given intratracheally (30 mg/kg) for 4 hours. AMs were lavaged, and total cell counts were determined. AMs were also obtained at end resuscitation and exposed to LPS (0.1 microg/mL) from 0 to 60 minutes. Activation of Hck, an Src family kinase, was analyzed by Western blot using a phosphospecific antibody. Nuclear extracts were obtained to examine NF-kappaB translocation.


RESULTS
S/R caused a rise in Src family activity compared with sham animals as shown by the phosphorylation of Hck. This was prevented by treating the animals during resuscitation with NAC. The LPS-induced NF-kappaB translocation in AMs after shock/resuscitation was 3-fold higher than in sham AMs treated with LPS. This augmented translocation was prevented by pretreating the animals with PP2 before resuscitation. In a parallel fashion, PP2 pretreatment reduced the absolute lung neutrophil sequestration.


CONCLUSIONS
Oxidant stress generated during S/R in vivo causes Src family kinase activation in AMs. Inhibition of Src activation by PP2 attenuates AM priming for increased LPS responsiveness after hemorrhagic shock and causes a modest reduction in lung injury. Inhibition of the Src family kinases may be a novel approach for the treatment of lung injury after trauma.
Department of Surgery, University Health Network, and University of Toronto, Ontario, Canada.
Rachel G
RG
Khadaroo
Ruijuan
R
He
Jean
J
Parodo
Kinga A
KA
Powers
John C
JC
Marshall
Andras
A
Kapus
Ori D
OD
Rotstein
D000818:Animals; D008070:Lipopolysaccharides / Q000633:toxicity; D008297:Male; D016328:NF-kappa B / Q000378:metabolism; D009504:Neutrophils / Q000502:physiology; D018384:Oxidative Stress; D010766:Phosphorylation; D021381:Protein Transport; D011743:Pyrimidines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D012128:Respiratory Distress Syndrome / Q000209:etiology*; D012771:Shock, Hemorrhagic / Q000201:enzymology; D014443:Tyrosine / Q000378:metabolism; D019061:src-Family Kinases / Q000502:physiology*
C412373:AG 1879; D008070:Lipopolysaccharides; D016328:NF-kappa B; D011743:Pyrimidines; D014443:Tyrosine; D019061:src-Family Kinases