Abstract
IntroductionSevere sepsis is characterised by intravascular or extravascular infection with microbial agents, systemic inflammation and microcirculatory dysfunction, leading to tissue damage, organ failure and death. The growth factor angiopoietin (Ang-1) has therapeutic potential but recombinant Ang-1 tends to aggregate and has a short half-life in vivo. This study aimed to investigate the acute effects of the more stable Ang-1 variant matrilin-1-angiopoietin-1 (MAT.Ang-1) on the function of the microcirculation in an experimental model of sepsis, and whether any protection by MAT-Ang-1 was associated with modulation of inflammatory cytokines, angiogenic factors or the endothelial nitric oxide synthase (eNOS)-Akt and vascular endothelial (VE)-cadherin pathways.MethodsAluminium window chambers were implanted into the dorsal skinfold of male C3H/HeN mice (7 to 10 weeks old) to expose the striated muscle microcirculation. Endotoxemia was induced by intraperitoneal injection of lipopolysaccharide (LPS, 1 mg/kg at 0 and 19 hours). MAT.Ang-1 was administered intravenously 20 hours after the onset of sepsis. Microcirculatory function was evaluated by intravital microscopy and Doppler fluximetry.ResultsEndotoxemia resulted in macromolecular leak, which was ameliorated by MAT.Ang-1 post-treatment. LPS induced a dramatic reduction in tissue perfusion, which was improved by MAT.Ang-1. Proteome profiler array analysis of skeletal muscle also demonstrated increased inflammatory and reduced angiogenic factors during endotoxemia. MAT.Ang-1 post-treatment reduced the level of IL-1β but did not significantly induce the expression of angiogenic factors. MAT.Ang-1 alone did not induce leak or increase angiogenic factors but did reduce vascular endothelial growth factor expression in controls.ConclusionAdministration of MAT.Ang-1 after the onset of sepsis protects the microcirculation from endotoxemia-induced vascular dysfunction through reducing inflammation but without pro-angiogenic actions, thus representing a novel, potential pharmacotherapeutic agent for the treatment of sepsis.
Highlights
Severe sepsis is characterised by intravascular or extravascular infection with microbial agents, systemic inflammation and microcirculatory dysfunction, leading to tissue damage, organ failure and death
It maintains the structural integrity of the mature vasculature [8], regulating microvascular permeability via several mechanisms, including: (i) Ang-1bound Tie-2 signalling through RAC - a serine/threonine-protein kinase (Akt) to endothelial nitric oxide synthase, promoting quiescence and survival; (ii) sequestration of non-receptor tyrosine kinase Src through the RhoA downstream target mammalian diaphanous (mDia), which prevents Src-mediated vascular endothelial (VE)-cadherin internalisation, stabilising inter-endothelial junctions [8]
MAT.Ang-1 protects mice against LPS-induced microvascular hyperpermeability LPS caused a significant increase in macromolecular leak compared with controls (LPS group vs. controls, 64.5 ± 4.9 vs. 25.0 ± 6.1 at hours, P < 0.01; 80.7 ± 10.1 vs. 35.1 ± 8.6 at hours, MAT.Ang-1 reduces the inflammatory response during endotoxemia LPS induced a trend of increased expression for the majority of cytokines analysed in skeletal muscle at 24 hours (Table S2 in Additional file 1), including the proinflammatory cytokines IL-1a, IL-1b, IL-6, TNFa, IFNg, soluble intracellular adhesion molecule-1 and triggering receptor expressed on myeloid cells-1 (TREM-1), which were reduced by posttreatment with MAT.Ang-1 (Figure 3)
Summary
Severe sepsis is characterised by intravascular or extravascular infection with microbial agents, systemic inflammation and microcirculatory dysfunction, leading to tissue damage, organ failure and death. Sepsis results from the presence of systemic or localised infections, initiating devastating proinflammatory effects on the microcirculation [1,2,3] These effects lead to increased microvascular permeability and leukocyte activation, along with reduced vascular resistance and blood flow [4], and without treatment they may result in tissue damage, multiple organ failure and death. Ang-1 is essential during angiogenesis, being required for correct organisation and maturation of newly formed vessels [7] It maintains the structural integrity of the mature vasculature [8], regulating microvascular permeability via several mechanisms, including: (i) Ang-1bound Tie-2 signalling through Akt to endothelial nitric oxide synthase (eNOS), promoting quiescence and survival; (ii) sequestration of non-receptor tyrosine kinase Src through the RhoA downstream target mDia, which prevents Src-mediated vascular endothelial (VE)-cadherin internalisation, stabilising inter-endothelial junctions [8]. Being part of the intracellular pathway for nitric oxide (NO)-mediated vasodilation, eNOS modulates blood flow and tissue perfusion [9]
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