Endothelial cell apoptosis is an early event in the development of acute lung injury (ALI). We have previously found that the Src family tyrosine kinase (STK) Yes activates caspase-3, whereas the STK Fyn inhibits caspase-3 activation in cultured pulmonary endothelial cells. We hypothesized that deficiency in Yes or Fyn in mice would have differential effects on lipopolysaccharide (LPS)-induced ALI. Mice were treated with LPS (10 mg/kg ip) for 24 h. Histological evidence of lung injury was greater in LPS-treated wild-type mice than in vehicle-treated wild-type mice, and the LPS-induced histological evidence of lung injury was attenuated in yes-/- mice and enhanced in fyn-/- mice. In wild-type or fyn-/- mice, LPS resulted in greater lung wet-to-dry weight ratios than in controls, whereas in yes-/- mice lung, wet-to-dry weight was similar between LPS and controls. LPS-exposed fyn-/- mice had greater respiratory system resistance and lower respiratory system compliance than did LPS-exposed wild-type mice. TUNEL positive cells in the lung following LPS treatment were greater in the fyn-/- mice and lower in the yes-/- mice compared with that in the wild-type mice. Following LPS treatment lung protein levels of PECAM-1 were lower in fyn-/- mice than in controls or yes-/- mice. LPS treatment increased cleaved caspase-3 protein levels in wild-type mice, whereas LPS-induced caspase-3 activation was attenuated in yes-/- mice and enhanced in fyn-/- mice. These results indicate that LPS-induced ALI is positively mediated via Yes-related mechanisms and negatively mediated by Fyn-related mechanisms.