ROCK induced inflammation of the microcirculation during endotoxemia mediated by nitric oxide synthase

Sepsis may be modeled using lipopolysaccharide (LPS), which alters levels of nitric oxide (NO), synthesized via endothelial and inducible nitric oxide synthase (eNOS and iNOS). This study aimed to determine whether the Rho kinase (ROCK) inhibitor fasudil protected against LPS-induced (endotoxemia) macromolecular leak and leukocyte adhesion via NOS pathways. Male Wistar rats (283 +/- 8 g, n = 36) were anaesthetized with thiopental and the mesentery prepared for fluorescent intravital microscopy (IVM). Animals received either (i) LPS alone (150 mu g kg(-1) h(-1) i.v., n = 6): (ii) fasudil (FAS, 3 mg kg(-1) i.v., n = 6) or (iii) fasudil (10 mg kg(-1) i.v., n = 6), immediately prior to LPS administration, (iv) fasudil (FAS, 3 mg kg(-1) i.v., n = 6) alone or (v) fasudil (FAS, 10 mg kg(-1) iv., n = 6) alone, or (vi) saline alone (1 ml kg(-1) h(-1) iv, n = 6) for 4 h (240 min). LPS increased macromolecular leak (cumulative normalized grey levels, arbitrary units) from post capillary venules (< 40 mu m) and this was reduced by 3 mg kg(-1) fasudil, however, 10 mg kg(-1) was less effective (t = 240 min, control: 3.3 +/- 1.7; LPS: 15.1 +/- 2.0; LPS + 3 mg kg(-1) fasudil: 3.3 +/- 1.1 (p < 0.05), LPS + 10 mg kg(-1) fasudil: 8.4 +/- 3.2 NS). The numbers of leukocytes adhering for > 1 min/100 mu m venule were reduced by fasudil (t = 240 min, control: 1.8 +/- 0.7; LPS: 7.0 +/- 1.0; LPS + 3 mg kg(-1) fasudil: 1.75 +/- 0.25, p < 0.05; LPS + 10 mg kg(-1) fasudil: 1.8 +/- 0.8, p < 0.05). Immunohistochemistry demonstrated that fasudil increased endothelial cell expression of eNOS during sepsis, and decreased LPS-induced up-regulation of iNOS. Inhibition of ROCK in rats increases eNOS and decreases iNOS during endotoxemia, concomitantly reducing microvascular inflammation. Thus, targeting the ROCK pathway during sepsis could have therapeutic potential for reducing inflammation via a NO dependent mechanism. (C) 2011 Elsevier Inc. All rights reserved.