Hydrogen sulfide triggers late-phase preconditioning in postischemic small intestine by an NO- and p38 MAPK-dependent mechanism.

Hydrogen sulfide (H(2)S) is one of three endogenous gases, along with carbon monoxide (CO) and nitric oxide (NO), that exert a variety of important vascular actions in vivo. Although it has been demonstrated that CO or NO can trigger the development of a preconditioned phenotype in postischemic tissues, it is unclear whether H(2)S may also induce protection in organs subsequently exposed to ischemia-reperfusion (I/R). In light of these observations, we postulated that preconditioning with the exogenous H(2)S donor sodium hydrosulfide (NaHS-PC) would inhibit leukocyte rolling (LR) and adhesion (LA) induced by I/R. We used intravital microscopic techniques to demonstrate that NaHS-PC 24 h, but not 1 h, before I/R causes postcapillary venules to shift to an anti-inflammatory phenotype in wild-type (WT) mice such that these vessels fail to support LR and LA during reperfusion. The protective effect of NaHS-PC on LR was largely abolished by coincident pharmacological inhibition of NO synthase (NOS) in WT animals and was absent in endothelial NOS-deficient (eNOS(-/-)) mice. A similar pattern of response was noted in WT mice treated concomitantly with NaHS plus p38 mitogen-activated protein kinase (MAPK) inhibitors (SB 203580 or SK-86002). Whereas the reduction in LA induced by antecedent NaHS was attenuated by pharmacological inhibition of NOS or p38 MAPK in WT mice, the antiadhesive effect of NaHS was still evident in eNOS(-/-) mice. Thus NaHS-PC prevents LR and LA by triggering the activation of an eNOS- and p38 MAPK-dependent mechanism. However, the role of eNOS in the antiadhesive effect of NaHS-PC was less prominent than its effect to reduce LR.

AICAR preconditioning prevents postischemic leukocyte rolling and adhesion: role of K(ATP) channels and heme oxygenase.

OBJECTIVE: We previously demonstrated that pharmacologic activation of AMP-activated protein kinase (AMPK) with 5-aminoimidazole-4-carboxamide 1-beta-D-ribofuranoside (AICAR) 24 hours prior to (AICAR preconditioning; AICAR-PC) ischemia/reperfusion (I/R) prevents postischemic leukocyte-endothelial cell adhesive interactions (LEI) by a mechanism initiated by endothelial nitric oxide synthase (eNOS)-dependent NO production during the period of AICAR-PC. The major aim of this study was to examine the role of ATP-sensitive potassium (K(ATP)) channels and heme oxygenase as mediators of the antiadhesive effects of AICAR-PC during I/R 24 hours later. METHODS: Intravital fluorescence microscopy was used to quantify LEI in the small intestine of AICAR-preconditioned C57BL/6J mice treated with K(ATP) channel or heme oxygenase inhibitors during I/R 24 hours after AICAR-PC in separate experiments. RESULTS: I/R induced marked increases in LEI relative to sham control mice, proadhesive responses that were prevented by AICAR-PC 24 hours prior to I/R. The effects of AICAR-PC to prevent postischemic LEI were abolished by K(ATP) channel or heme oxygenase inhibition during I/R. DISCUSSION/CONCLUSION: Our results indicate that the antiadhesive effects of AICAR-PC are mediated by K(ATP) channel- and heme oxygenase-dependent mechanisms during I/R.

Angiotensin II mediates postischemic leukocyte-endothelial interactions: role of calcitonin gene-related peptide.

Vascular inflammation and enhanced production of angiotensin II (ANG II) are involved in the pathogenesis of hypertension and diabetes, disease states that predispose the afflicted individuals to ischemic disorders. In light of these observations, we postulated that ANG II may play a role in promoting leukocyte rolling (LR) and adhesion (LA) in postcapillary venules after exposure of the small intestine to ischemia-reperfusion (I/R). Using an intravital microscopic approach in C57BL/6J mice, we showed that ANG II type I (AT(1)) or type II (AT(2)) receptor antagonism (with valsartan or PD-123319, respectively), inhibition of angiotensin-converting enzyme (ACE) with captopril, or calcitonin gene-related peptide (CGRP) receptor blockade (CGRP8-37) prevented postischemic LR but did not influence I/R-induced LA. However, both postischemic LR and LA were largely abolished by concomitant AT(1) and AT(2) receptor blockade or chymase inhibition (with Y-40079). Additionally, exogenously administered ANG II increased LR and LA, effects that were attenuated by pretreatment with a CGRP receptor antagonist or an NADPH oxidase inhibitor (apocynin). Our work suggests that ANG II, formed by the enzymatic activity of ACE and chymase, plays an important role in inducing postischemic LR and LA, effects that involve the engagement of both AT(1) and AT(2) receptors and may be mediated by CGRP and NADPH oxidase.

5'-AMP-activated protein kinase activation prevents postischemic leukocyte-endothelial cell adhesive interactions.

Preconditioning (PC) with nitric oxide (NO) donors or agents that increase endothelial NO synthase (eNOS) activity 24 h before ischemia-reperfusion (I/R) prevents postischemic leukocyte rolling (LR) and stationary leukocyte adhesion (LA). Since 5'-AMP-activated protein kinase (AMPK) phosphorylates eNOS at Ser1177, resulting in activation, we postulated that AMPK activation may trigger the development of a preconditioned anti-inflammatory phenotype similar to that induced by NO donors. Wild-type (WT) C57BL/6J and eNOS(-/-) mice were treated with the AMPK agonist 5-aminoimidazole-4-carboxamide 1-beta-d-furanoside (AICAR) 30 min (early AICAR PC) or 24 h (late AICAR PC) before I/R; LR and LA were quantified in single postcapillary venules in the jejunum using intravital microscopy. I/R induced comparable marked increases in LR and LA in WT and eNOS(-/-) mice relative to sham-operated (no ischemia) animals. Late AICAR PC prevented postischemic LR and LA, whereas early AICAR PC prevented LA in WT mice. Late AICAR PC was ineffective in preventing I/R-induced LR but not LA in the eNOS(-/-) mice, and the same pattern was seen in WT animals treated with the NOS inhibitor N(omega)-nitro-l-arginine. Early AICAR PC remained effective in preventing LA in eNOS(-/-) mice. Our results indicate that both early and late PC with an AMPK agonist produces an anti-inflammatory phenotype in postcapillary venules. Since the protection afforded by late AICAR PC on postischemic LR was prevented by NOS inhibition in WT mice and absent in eNOS-deficient mice, it appears that eNOS triggers this protective effect. In stark contrast, antecedent AMPK activation prevented I/R-induced LA by an eNOS-independent mechanism.

Role of eNOS-derived NO in the postischemic anti-inflammatory effects of antecedent ethanol ingestion in murine small intestine.

Ingestion of low levels of ethanol 24 h before [ethanol preconditioning (EPC)] ischemia and reperfusion (I/R) prevents postischemic leukocyte rolling (LR) and adhesion (LA), effects that were abolished by adenosine A(2) receptor (ADO-A(2)R) antagonists or nitric oxide (NO) synthase (NOS) inhibitors. The aims of this study were to determine whether NO derived from endothelial NOS (eNOS) during the period of ethanol exposure triggered entrance into this preconditioned state and whether these events were initiated by an ADO-A(2)R-dependent mechanism. Ethanol or distilled water vehicle was administered to C57BL/6J [wild type (WT)] or eNOS-deficient (eNOS-/-) mice by gavage. Twenty-four hours later, the superior mesenteric artery was occluded for 45 min. LR and LA were quantified by intravital microscopy after 30 and 60 min of reperfusion. I/R increased LR and LA in WT mice, effects that were abolished by EPC or NO donor preconditioning (NO-PC). NO-PC was not attenuated by coincident administration of an ADO-A(2)R antagonist. I/R increased LR and LA in eNOS-/- mice to levels comparable with those noted in WT animals. However, EPC only slightly attenuated postischemic LR and LA, whereas NO-PC remained effective as a preconditioning stimulus in eNOS-/- mice. Preconditioning with an ADO-A(2)R agonist (which we previously demonstrated prevents I/R-induced LR and LA in WT animals) failed to attenuate these postischemic adhesive responses in eNOS-/- mice. Our results indicate that EPC is triggered by NO formed secondary to ADO-A(2)R-dependent eNOS activation during the period of ethanol exposure 24 h before I/R.

Role of calcitonin gene-related peptide in the postischemic anti-inflammatory effects of antecedent ethanol ingestion.

The aim of this study was to determine the role of calcitonin gene-related peptide (CGRP) in the postischemic anti-inflammatory effects of antecedent ethanol ingestion. Ethanol was administered to wild-type C57BL/6 mice on day 1 as a bolus by gavage at a dose that produces a peak plasma ethanol of 45 mg/dl 30 min after administration. Twenty-four hours later (day 2), the superior mesenteric artery was occluded for 45 min followed by 70 min of reperfusion (I/R). Intravital fluorescence microscopy was used to quantify the numbers of rolling (LR) and adherent (LA) leukocytes labeled with carboxyfluorescein diacetate succinimidyl ester in postcapillary venules of the small intestine. I/R increased LR and LA, effects that were prevented by antecedent ethanol. The postischemic anti-inflammatory effects of ethanol consumption were abolished by administration of a specific CGRP receptor antagonist [CGRP-(8-37)] or after sensory nerve neurotransmitter depletion using capsaicin administered 4 days before ethanol ingestion, which initially induces rapid release of CGRP from sensory nerves, thereby depleting stored neuropeptide. Administration of exogenous CGRP or induction of endogenous CGRP release by treatment with capsaicin 24 h before I/R mimicked the postischemic anti-inflammatory effects of antecedent ethanol ingestion. Preconditioning with capsaicin 24 h before I/R was prevented by coincident treatment with CGRP-(8-37), while exogenous CGRP induced an anti-inflammatory phenotype in mice depleted of CGRP by capsaicin administration 4 days earlier. Our results indicate that the effect of antecedent ethanol ingestion to prevent postischemic LR and LA is initiated by a CGRP-dependent mechanism.

Polyphenols in cerebral ischemia: novel targets for neuroprotection.

Plant polyphenols are dietary components that exert a variety of biochemical and pharmacological effects. Recently, considerable interest has been focused on polyphenols because of their antioxidant, anti-inflammatory, and antiproliferative activities. Oxidative stress is thought to be a key event in the pathogenesis of cerebral ischemia. Overproduction of reactive oxygen species during ischemia/reperfusion could cause an imbalance between oxidative and antioxidative processes. Reactive oxygen species can damage lipids, proteins, and nucleic acids, thereby inducing apoptosis or necrosis. There is increasing evidence supporting the hypothesis that plant polyphenols can provide protection against neurodegenerative changes associated with cerebral ischemia. This article reviews the neuroprotective effects of plant extracts and their constituents that have been used in animal models of cerebral ischemia. The use of polyphenols as therapeutic agents in stroke has been suggested.