Remote trauma sensitizes hepatic microcirculation to endothelin via caveolin inhibition of eNOS activity

This study addresses the microvascular mechanisms by which a remote, mild stress such as blunt trauma sensitizes the liver to injury. Rats received closed femur fracture (FFx), and 24 h later livers were isolated and perfused at a similar starting flow rate for assessment of vascular response to endothelin-1 (ET-1). Sinusoidal volumetric flow (Q(S)), red blood cell velocity (V-RBC), and sinusoidal diameter (D-s) were determined by intravital microscopy. Baseline portal resistance in livers from FFx rats was not changed. The FFx group showed a lower baseline V-RBC (322.9 +/- 26.4 and 207.3 +/- 17.2 mum/s in sham and FFx(1)) and Q(S) (28.4 +/- 4.2 and 17.6 +/- 2.1 pL/s in sham and FFx, P < 0.05). ET-1 caused a decrease in the V-RBC in sham but no change after FFx. In contrast, D. was unchanged by ET-1 in sham but decreased in FFx (10.3 +/- 0.4 to 10.7 +/- 0.5 vs. 10.6 +/- 0.4 to 9.0 +/- 0.4 pm at 10 min in sham and FFx groups, P < 0.05). The overall result of these changes was a greater decrease in sinusoidal flow in FFx compared with sham. There was no significant change in mRNA for ET-1, endothelin A (ETA) receptor, or NOS (inducible nitric oxide synthase) in FFx compared with sham. However, endothelin B (ETB) receptor mRNA and eNOS (endothelial nitric oxide synthase) mRNA were increased in the FFx group (ETB, 54.81 +/- 8.08 in sham vs. 83.28 +/- 8.19 in FFx; eNOS, 56.11 +/- 2.53 in sham vs. 83.31 +/- 5.51 in FFx; P < 0.05) while the levels of these proteins remained unchanged. Caveolin-1 (cav-1) protein levels were elevated in FFx, and coimmunoprecipitation with both ETB and eNOS showed increased associations with these proteins, suggesting a possible inactivation of eNOS. The eNOS activity was also blunted in FFx animals in the presence of increased cav-1 expression. Taken together, these results demonstrate that remote trauma sensitizes the liver to the sinusoidal constrictor effect of ET-1. We propose that this hyperresponsiveness occurs as a result of uncoupling of the ETB receptor from eNOS activity mediated by interaction of eNOS and possibly the ETB, receptor with increased caveolin-1. This vascular sensitization that occurs after IFFx may contribute to the exacerbation of injury during subsequent stresses.