HIF-1α stabilization modified by glutaredoxin-1 is critical for intestinal angiogenesis in NEC pathogenesis

Background: Hypoxia inducible factor (HIF-1α) are essential in the pathogenesis of necrotizing enterocolitis (NEC), which is stabilized by Grx1 deletion. Until now, the mechanism of HIF-1α in the intestinal microcirculation in NEC is not well defined. We intend to investigate the role of HIF-1α in the development of NEC in regulating the microcirculation and the following vasodilatory signal, VEGF. Materials and methods: Experimental NEC was induced in full-term C57BL/6 mouse and Grx1-/-pups through the formula gavage and hypoxia technique. The HIF-1α signal was blocked utilizing the HIF-1α inhibitor, YC-1. Intestinal tissues were collected at predetermined time points for the assessment of intestinal microcirculation and the HIF-1α activity involved signal. Results: We found that NEC inducement impaired the intestinal microcirculation, but intestinal blood flow and capillary density were ameliorated in Grx1-/-mice, which was associated with the GSH-protein adducts of HIF-1α in the intestinal tissue. Grx1 ablation could also promote vascular endothelial growth factor (VEGFA) production in the intestinal tissue. This intestinal microvascular improvement was not found in the HIF-1α inhibited mice, suggesting the HIF-1α dependent manner for intestinal microcirculatory perfusion. Conclusion: The current data demonstrated that HIF-1α signaling is involved in the intestinal microvascular modification during the pathogenesis of NEC, suggesting that targeting with HIF-1α might be a promising strategy for NEC treatment.

Sub-therapeutic vasopressin but not therapeutic vasopressin improves gastrointestinal microcirculation in septic rats: A randomized, placebo-controlled, blinded trial

Introduction Sepsis impairs gastrointestinal microcirculation and it is hypothesized that this might increase patient’s mortality. Sub-therapeutic vasopressin improves gastric microcirculation under physiologic conditions whereas a therapeutic dosing regimen seems to be rather detrimental. However, the effects of sub-therapeutic vasopressin on gastrointestinal microcirculation in sepsis are largely unknown. Therefore, we conducted this trial to investigate the effect of sub-therapeutic as well as therapeutic vasopressin on gastrointestinal microcirculation in sepsis. Methods 40 male Wistar rats were randomized into 4 groups. Colon ascendens stent peritonitis (CASP)-surgery was performed to establish mild or moderate sepsis. 24 hours after surgery, animals received either vasopressin with increasing dosages every 30 min (6.75, 13.5 (sub-therapeutic), 27 mU · kg-1 · h-1 (therapeutic)) or vehicle. Microcirculatory oxygenation (μHBO2) of the colon was recorded for 90 min using tissue reflectance spectrophotometry. Intestinal microcirculatory perfusion (total vessel density (TVD; mm/mm2) and perfused vessel density (PVD; mm/mm2)) were measured using incident dark field-Imaging at baseline and after 60 min. Results In mild as well as in moderate septic animals with vehicle-infusion intestinal μHbO2, TVD and PVD remained constant. In contrast, in moderate sepsis, sub-therapeutic vasopressin with 13.5 mU · kg-1 · h-1 elevated intestinal μHBO2 (+ 6.1 ± 5.3%; p < 0.05 vs. baseline) and TVD (+ 5.2 ± 3.0 mm/mm2; p < 0.05 vs. baseline). μHBO2, TVD and PVD were significantly increased compared to moderate sepsis alone. However, therapeutic vasopressin did not change intestinal microcirculation. In mild septic animals sub-therapeutic as well as therapeutic vasopressin had no relevant effect on gastrointestinal microcirculation. Systemic blood pressure remained constant in all groups. Conclusion Sub-therapeutic vasopressin improves gastrointestinal microcirculatory oxygenation in moderate sepsis without altering systemic blood pressure. This protective effect seems to be mediated by an enhanced microcirculatory perfusion and thereby increased oxygen supply. In contrast, therapeutic vasopressin did not show this beneficial effect.

Utility of intestinal intravital microscopy for the study of CNS injury-induced immunodepression syndrome (CIDS)

BACKGROUND: Stroke, traumatic brain injury, or other forms of central nervous system (CNS) injury initiate a local inflammatory response. Compensatory anti-inflammatory pathways are activated to limit secondary damage due to inflammation. The associated release of immunosuppressing neuromodulators can result in system-wide immune dysregulation (CNS injury-induced immune-depression syndrome –CIDS). OBJECTIVE: To establish an experimental stroke model where CIDS can be studied by intravital microscopy (IVM). METHODS: We used the photothrombotic stroke (PTS) model in C57BL/6 mice and studied its effects on peripheral immunity following challenge with lipopolysaccharide (LPS). Leukocyte activation, as well as capillary perfusion of the microcirculation, were assessed using intestinal intravital microscopy (IVM). RESULTS: PTS caused a significant reduction in the number of adhering leukocytes in submucosal venules of the terminal ileum of mice challenged with LPS compared to LPS-challenged animals without stroke. Leukocyte rolling was also impacted by PTS in the submucosal venules. Following stroke, we also observed decreased mucosal functional capillary density (FCD). CONCLUSIONS: Our results suggest that PTS with subsequent LPS challenge poses as a viable model to further study CIDS using intravital microscopy of the intestinal microcirculation.

HIF-1α stabilization modified by glutaredoxin-1 is critical for intestinal angiogenesis in NEC pathogenesis

BackgroundHypoxia inducible factor (HIF-1α) are essential in the pathogenesis of necrotizing enterocolitis (NEC), which is stabilized by Grx1 deletion. Until now, the mechanism of HIF-1α in the intestinal microcirculation in NEC is not well defined. We intend to investigate the role of HIF-1α in the development of NEC in regulating the microcirculation and the following vasodilatory signal, VEGF.Materials and methodsExperimental NEC was induced in full-term C57BL/6 mouse and Grx1-/- pups through the formula gavage and hypoxia technique. The HIF-1α signal was blocked utilizing the HIF-1α inhibitor, YC-1. Intestinal tissues were collected at predetermined time points for the assessment of intestinal microcirculation and the HIF-1α activity involved signal.ResultsWe found that NEC inducement impaired the intestinal microcirculation, but intestinal blood flow and capillary density were ameliorated in Grx1-/- mice, which was associated with the GSH-protein adducts of HIF-1α in the intestinal tissue. Grx1 ablation could also promote vascular endothelial growth factor (VEGFA) production in the intestinal tissue. This intestinal microvascular improvement was not found in the HIF-1α inhibited mice, suggesting the HIF-1α dependent manner for intestinal microcirculatory perfusion.ConclusionThe current data demonstrated that HIF-1α signaling is involved in the intestinal microvascular modification during the pathogenesis of NEC, suggesting that targeting with HIF-1α might be a promising strategy for NEC treatment.

Unfractionated Heparin Improves the Intestinal Microcirculation in a Canine Septic Shock Model

Background. Alterations of microcirculation are associated with organ hypoperfusion and high mortality in septic shock. This study is aimed at investigating the effects of unfractionated heparin (UFH) on intestinal microcirculatory perfusion and systemic circulation in a septic shock model. Methods. Twenty-four beagle dogs were randomly allocated into four groups: (a) sham group: healthy controls, (b) shock group: septic shock induced by Escherichia coli, (c) basic therapy group: septic shock animals treated with antibiotics and 10 ml/kg/h saline, and (d) heparin group: septic shock animals treated with basic therapy plus UFH. Hemodynamic variables were measured within 24 h after E. coli administration. The intestinal microcirculation was simultaneously investigated with a sidestream dark-field imaging technique. Additionally, the function of vital organs was evaluated at 12 h postadministration (T12). Results. E. coli induced a progressive septic shock in which the mean arterial pressure (MAP) decreased and lactate levels sharply increased, accompanied by deteriorated microvessel perfusion. While basic therapy partially improved the microvascular flow index and the perfused microvessel density in the jejunal villi, UFH significantly restored major microcirculation variables at T12. Physiological variables, including MAP, urine output, and lactate levels, were improved by UFH, whereas some hemodynamic indices were not affected by UFH. With respect to organ function, UFH increased the platelet count and decreased the creatinine level. Conclusions. UFH improves microcirculatory perfusion of the small intestine independently of the changes in systemic hemodynamic variables in a canine model of septic shock, thereby improving coagulation and renal function.

Sodium Thiosulfate Improves Intestinal and Hepatic Microcirculation Without Affecting Mitochondrial Function in Experimental Sepsis

IntroductionIn the immunology of sepsis microcirculatory and mitochondrial dysfunction in the gastrointestinal system are important contributors to mortality. Hydrogen sulfide (H2S) optimizes gastrointestinal oxygen supply and mitochondrial respiration predominantly via K(ATP)-channels. Therefore, we tested the hypothesis that sodium thiosulfate (STS), an inducer of endogenous H2S, improves intestinal and hepatic microcirculation and mitochondrial function via K(ATP)-channels in sepsis.MethodsIn 40 male Wistar rats colon ascendens stent peritonitis (CASP) surgery was performed to establish sepsis. Animals were randomized into 4 groups (1: STS 1 g • kg-1 i.p., 2: glibenclamide (GL) 5 mg • kg-1 i.p., 3: STS + GL, 4: vehicle (VE) i.p.). Treatment was given directly after CASP-surgery and 24 hours later. Microcirculatory oxygenation (µHBO2) and flow (µflow) of the colon and the liver were continuously recorded over 90 min using tissue reflectance spectrophotometry. Mitochondrial oxygen consumption in tissue homogenates was determined with respirometry. Statistic: two-way ANOVA + Dunnett´s and Tukey post - hoc test (microcirculation) and Kruskal-Wallis test + Dunn’s multiple comparison test (mitochondria). p &lt; 0.05 was considered significant.ResultsSTS increased µHbO2 (colon: 90 min: + 10.4 ± 18.3%; liver: 90 min: + 5.8 ± 9.1%; p &lt; 0.05 vs. baseline). Furthermore, STS ameliorated µflow (colon: 60 min: + 51.9 ± 71.1 aU; liver: 90 min: + 22.5 ± 20.0 aU; p &lt; 0.05 vs. baseline). In both organs, µHbO2 and µflow were significantly higher after STS compared to VE. The combination of STS and GL increased colonic µHbO2 and µflow (µHbO2 90 min: + 8.7 ± 11.5%; µflow: 90 min: + 41.8 ± 63.3 aU; p &lt; 0.05 vs. baseline), with significantly higher values compared to VE. Liver µHbO2 and µflow did not change after STS and GL. GL alone did not change colonic or hepatic µHbO2 or µflow. Mitochondrial oxygen consumption and macrohemodynamic remained unaltered.ConclusionThe beneficial effect of STS on intestinal and hepatic microcirculatory oxygenation in sepsis seems to be mediated by an increased microcirculatory perfusion and not by mitochondrial respiratory or macrohemodynamic changes. Furthermore, the effect of STS on hepatic but not on intestinal microcirculation seems to be K(ATP)-channel-dependent.

Luteolin ameliorates lipopolysaccharide-induced microcirculatory disturbance through inhibiting leukocyte adhesion in rat mesenteric venules

Background Microcirculatory disturbance is closely associated with multiple diseases such as ischemic and septic stroke. Luteolin (3,4,5,7-tetrahydroxyflavone) is a vascular protective flavonoid present in several dietary foods. However, how luteolin plays a role in microcirculatory disturbance is still unknown. The purpose of this study was to find out the influence of luteolin on the lipopolysaccharide (LPS)-induced microcirculatory disturbance, focusing on its effect on leukocyte adhesion and the underlying mechanism of this effect. Methods After injecting LPS into rats, we used an inverted intravital microscope to observe the velocity of red blood cells in venules, numbers of leukocytes adherent to and emigrated across the venular wall, hydrogen peroxide production in venular walls and mast cell degranulation. Intestinal microcirculation blood flow was measured by High-resolution Laser Doppler Perfusion Imaging. Histological changes of small intestine and mesenteric arteries were evaluated. Additionally, cell adhesion stimulated by LPS was tested on EA.hy926 and THP-1 cells. The production of pro-inflammatory cytokines, adhesion molecules and the activation of TLR4/Myd88/NF-κB signaling pathway were determined. Results The results showed luteolin significantly inhibited LPS-induced leukocyte adhesion, hydrogen peroxide production and mast cell degranulation, and increased intestinal microcirculation blood flow and ameliorated pathological changes in the mesenteric artery and the small intestine. Furthermore, luteolin inhibited the release of pro-inflammatory cytokines, the expression of TLR4, Myd88, ICAM-1, and VCAM-1, the phosphorylation of IκB-α and NF-κB/p65 in LPS stimulated EA.hy926. Conclusions Our findings revealed that it is likely that luteolin can ameliorate microcirculatory disturbance. The inhibitory effects of luteolin on the leukocyte adhesion stimulated by LPS, which participates in the development of microcirculatory disturbance, are mediated through the regulation of the TLR4/Myd88/NF-κB signaling pathway.

Imaging of the Intestinal Microcirculation during Acute and Chronic Inflammation

Because of its unique microvascular anatomy, the intestine is particularly vulnerable to microcirculatory disturbances. During inflammation, pathological changes in blood flow, vessel integrity and capillary density result in impaired tissue oxygenation. In severe cases, these changes can progress to multiorgan failure and possibly death. Microcirculation may be evaluated in superficial tissues in patients using video microscopy devices, but these techniques do not allow the assessment of intestinal microcirculation. The gold standard for the experimental evaluation of intestinal microcirculation is intravital microscopy, a technique that allows for the in vivo examination of many pathophysiological processes including leukocyte-endothelial interactions and capillary blood flow. This review provides an overview of changes in the intestinal microcirculation in various acute and chronic inflammatory conditions. Acute conditions discussed include local infections, severe acute pancreatitis, necrotizing enterocolitis and sepsis. Inflammatory bowel disease and irritable bowel syndrome are included as examples of chronic conditions of the intestine.