Janice Russell

Role of neutrophils in hemorrhagic shock-induced gastric mucosal injury in the rat

Gastric mucosal clearance of 51Cr-labeled red blood cells (51Cr-RBC) was measured in rats during a 30-min control period, a 30-min ischemic period (hemorrhage to 27 mmHg arterial pressure), and a 60-min reperfusion period (reinfusion of shed blood). In untreated (control) rats, a dramatic rise in the leakage of 51Cr-labeled red blood cells into the gastric lumen was observed during the reperfusion period. Treatment with neutrophil antiserum attenuated 51Cr-labeled red blood cell flux into the gastric lumen. Using the radioactive microsphere technique, neutrophil-depleted animals were shown to have higher blood flows in the ischemic period than the untreated rats. Bleeding of untreated rats to a mean arterial pressure of 40 mmHg resulted in blood flows that were not different from those in antiserum-treated rats bled to 27 mmHg and leakage of 51Cr-labeled red blood cells similar to that measured in antiserum-treated rats. The results of this study indicate that neutrophils play an important role in hemorrhagic shock-induced gastric bleeding.

NAD(P)H Oxidase–Derived Superoxide Mediates Hypercholesterolemia-Induced Leukocyte–Endothelial Cell Adhesion

Experimental animals placed on a high-cholesterol diet for 2 or more weeks exhibit an inflammatory response in postcapillary venules. The aims of this study were to determine (1) whether superoxide mediates the hypercholesterolemia-induced inflammatory response and (2) whether leukocyte and/or vessel wall NAD(P)H oxidase contributes to this response. Intravital videomicroscopy was used to quantify leukocyte–endothelial cell adhesion in cremasteric postcapillary venules of wild-type (WT) mice, CuZn-superoxide dismutase transgenic (SOD TgN) mice, and mice heterozygous (p47 phox +/−) or homozygous (p47 phox −/−) for NAD(P)H oxidase placed on either a normal diet or high-cholesterol diet (HCD) for 2 weeks. The number of adherent and emigrated leukocytes in postcapillary venules of WT HCD mice was significantly higher than that detected in venules of their normal-diet counterparts. However, the HCD-induced recruitment of adherent and emigrated leukocytes was not observed in SOD TgN mice. Whereas hypercholesterolemic p47 phox +/− and WT mice exhibited similar inflammatory responses, p47 phox −/− mice did not. Bone marrow chimeras were developed to selectively delete p47 phox from either the vessel wall or circulating leukocytes. Whereas WT marrow transplanted into WT mice produced a normal inflammatory response of venules to HCD, chimeric mice with p47 phox deficiency in either the vessel wall or leukocytes exhibited an attenuated inflammatory response to HCD that was comparable with that observed in p47 phox −/− HCD mice. Our findings indicate that enhanced superoxide production is a critical event that initiates the leukocyte–endothelial cell adhesion in postcapillary venules of HCD mice. NAD(P)H oxidase appears to be an important source of this superoxide.

Apolipoprotein A-IV inhibits experimental colitis

The antiatherogenic properties of apoA-IV suggest that this protein may act as an anti-inflammatory agent. We examined this possibility in a mouse model of acute colitis. Mice consumed 3% dextran sulfate sodium (DSS) in their drinking water for 7 days, with or without daily intraperitoneal injections of recombinant human apoA-IV. apoA-IV significantly and specifically delayed the onset, and reduced the severity and extent of, DSS-induced inflammation, as assessed by clinical disease activity score, macroscopic appearance and histology of the colon, and tissue myeloperoxidase activity. Intravital fluorescence microscopy of colonic microvasculature revealed that apoA-IV significantly inhibited DSS-induced leukocyte and platelet adhesive interactions. Furthermore, apoA-IV dramatically reduced the upregulation of P-selectin on colonic endothelium during DSS-colitis. apoA-IV knockout mice exhibited a significantly greater inflammatory response to DSS than did their WT littermates; this greater susceptibility to DSS-induced inflammation was reversed upon exogenous administration of apoA-IV to knockout mice. These results provide the first direct support for the hypothesis that apoA-IV is an endogenous anti-inflammatory protein. This anti-inflammatory effect likely involves the inhibition of P-selectin-mediated leukocyte and platelet adhesive interactions.

Molecular Determinants of the Prothrombogenic and Inflammatory Phenotype Assumed by the Postischemic Cerebral Microcirculation

Background and Purpose— Circulating blood cells have been implicated in the pathogenesis of cerebral ischemia/reperfusion (I/R) injury and stroke. The objective of this study was to define the magnitude and molecular determinants of the platelet- and leukocyte–endothelial cell adhesive interactions induced by I/R in the mouse brain. Methods— Bilateral common carotid artery occlusion was induced for 1 hour in C57BL/6 mice, followed by either 40 minutes or 4 hours of reperfusion. Fluorescent platelets were administered intravenously, and the frontal brain surface was observed with intravital fluorescence microscopy. Leukocyte–endothelial cell adhesion was monitored with the use of rhodamine-6G. Results— Ischemia followed by 40 minutes of reperfusion resulted in the rolling (125.1±23.6/mm2) and firm adhesion (109.5±25.8/mm2) of leukocytes but not platelets in venules. However, with 4 hours of reperfusion, rolling (138.8±24.6/mm2) and firm adhesion (153.7±22.3/mm2) of platelets were detected, and this was accompanied by a more intense recruitment of rolling (374.5±54.6/mm2) and adherent (445.2±57.1/mm2) leukocytes. In mice deficient in either P-selectin (P-selectin−/−) or intercellular adhesion molecule-1 (ICAM-1) (ICAM-1−/−), the I/R-induced platelet–endothelial cell (by 80% and 60%, respectively) and leukocyte–endothelial cell (by 84% and 78%, respectively) interactions were significantly blunted compared with those of wild-type mice. Conclusions— These findings indicate that I/R promotes the adhesion of both platelets and leukocytes in cerebral venules, with the accumulation of adherent leukocytes preceding the recruitment of platelets. Both P-selectin and ICAM-1 contribute to the inflammatory and prothrombogenic state induced by cerebral I/R.

Blood Cell-Derived RANTES Mediates Cerebral Microvascular Dysfunction, Inflammation, and Tissue Injury After Focal Ischemia–Reperfusion

Background and Purpose— Although chemokines have been implicated in cardiovascular diseases, few studies have addressed the role of these inflammatory mediators in ischemic stroke. This study tested the hypothesis that RANTES (CCL5; regulated on activation, normal T-cell expressed and secreted) mediates the cerebral microvascular dysfunction, inflammation, and tissue injury induced by brain ischemia and reperfusion. Methods— After 60-minute middle cerebral artery occlusion and reperfusion, the adhesion of leukocytes and platelets in cerebral venules, infarct volume, and blood–brain barrier permeability were measured in wild-type mice (WT), RANTES-deficient mice (RANTES−/−), WT mice transplanted with RANTES−/− bone marrow (RANTES>WT), and control bone marrow chimeras (WT>WT). The concentration of RANTES and several cytokines was also measured by enzyme-linked immunosorbent assay and a cytometric bead array. Results— The enhanced leukocyte and platelet adhesion, increased blood–brain barrier permeability, and tissue infarction elicited in WT and WT>WT mice after middle cerebral artery occlusion and reperfusion were significantly blunted in RANTES−/− mice. Similar attenuation of the middle cerebral artery occlusion and reperfusion-induced responses were noted in RANTES>WT chimeras. Although RANTES deficiency did not alter the changes in tissue cytokine levels elicited by middle cerebral artery occlusion and reperfusion, plasma concentrations interleukin-6, interleukin-10, and interleukin-12 were all reduced. Conclusions— These findings implicate blood cell-derived RANTES in the microvascular, inflammatory, and tissue injury responses of the brain to ischemia and reperfusion.

Effects of chronic arterial hypertension on constitutive and induced intercellular adhesion molecule-1 expression in vivo.

Recent reports indicate that bacterial endotoxin (lipopolysaccharide) and cytokines elicit a more profound increase in the surface expression of intercellular adhesion molecule-1 (ICAM-1) in cultured endothelial cells derived from spontaneously hypertensive (SHR) versus normotensive Wistar-Kyoto rats (WKY). Our objective in this study was to characterize and compare in vivo ICAM-1 expression in SHR and WKY under basal conditions and after 5 hours of endothelial cell activation with either lipopolysaccharide (5 mg/kg i.p.) or tumor necrosis factor-alpha (TNF-alpha; 1, 5, and 10 micrograms/kg i.p.). ICAM-1 expression was quantified in different tissues by the double-radiolabeled monoclonal antibody technique. When constitutive (baseline) ICAM-1 expression was corrected for endothelial cell surface area, significantly higher values were noted in SHR than WKY but only in splanchnic organs. Lipopolysaccharide and TNF-alpha elicited significant increases in ICAM-1 expression in all tissues of both WKY and SHR. However, the magnitude of the lipopolysaccharide-induced ICAM-1 upregulation in heart, stomach, skeletal muscle, and brain was significantly lower in SHR than WKY. A similar blunted ICAM-1 upregulation was noted in the stomach of SHR after administration of 5 micrograms/kg TNF-alpha. The differences in induced ICAM-1 expression between SHR and WKY do not appear to be due to differences in endothelial cell surface area or plasma glucocorticoid levels. These results suggest that chronic arterial hypertension results in altered ICAM-1 expression on the endothelium, which may contribute to the abnormal inflammatory responses associated with this disease.

Tissue factor: a mediator of inflammatory cell recruitment, tissue injury, and thrombus formation in experimental colitis.

There is growing evidence for an interplay between inflammatory and coagulation pathways in acute and chronic inflammatory diseases. However, it remains unclear whether components of the coagulation pathway, such as tissue factor (TF), contribute to intestinal inflammation, and whether targeting TF will blunt the inflammatory cell recruitment, tissue injury, and enhanced thrombus formation that occur in experimental colitis. Mice were fed 3% dextran sodium sulfate (DSS) to induce colonic inflammation, with some mice receiving a mouse TF-blocking antibody (muTF-Ab). The adhesion of leukocytes and platelets in colonic venules, light/dye-induced thrombus formation in cremaster muscle microvessels, as well as disease activity index, thrombin–antithrombin (TAT) complexes in plasma, and histopathologic changes in the colonic mucosa were monitored in untreated and muTF-Ab–treated colitic mice. In untreated mice, DSS elicited the recruitment of adherent leukocytes and platelets in colonic venules, caused gross and histologic injury, increased plasma TAT complexes, and enhanced thrombus formation in muscle arterioles. muTF-Ab prevented elevation in TAT complexes, reduced blood cell recruitment and tissue injury, and blunted thrombus formation in DSS colitic mice. These findings implicate TF in intestinal inflammation and support an interaction between inflammation and coagulation in experimental colitis.

Obesity Exacerbates Sepsis-Induced Inflammation and Microvascular Dysfunction in Mouse Brain

Objective: Obese patients with sepsis have higher morbidity and mortality than lean counterparts, but the mechanisms involved are unknown. The authors examined the inflammatory and thrombogenic responses of the cerebral microvasculature to sepsis induced by cecal ligation and perforation in obese and lean wild‐type mice.

Endothelial cell adhesion molecule expression in gene-targeted mice

Gene-targeted mice are now routinely employed as tools for defining the contribution of different leukocyte and endothelial cell adhesion molecules to the leukocyte recruitment and tissue injury associated with acute and chronic inflammation. The objective of this study was to determine whether gene-targeted mice that are deficient in CD11/CD18, intracellular adhesion molecule-1 (ICAM-1), or P-selectin exhibit an altered constitutive or induced expression of the endothelial cell adhesion molecules E- and P-selectin. The gene-targeted mice were all developed in the 129Sv mouse strain and backcrossed into C57Bl/6J mice. The number of backcrosses ranged between 8 (P-selectin) and 10 (CD18 and ICAM-1) generations. The dual-radiolabeled monoclonal antibody technique was used to quantify E- and P-selectin expression in different vascular beds. In the unstimulated state, E-selectin expression was significantly elevated (relative to wild-type mice) in the stomach, large intestine, and brain of mutants deficient in ICAM-1. In general, constitutive expression of P-selectin did not differ between wild-type, ICAM-1-deficient, and CD11/CD18-deficient mutants. In CD11/CD18-deficient mice, tumor necrosis factor-α (TNF-α) administration elicited a more profound upregulation of P-selectin in several vascular beds, compared with wild-type and ICAM-1-deficient mice. E-selectin expression in brain of TNF-α-stimulated, ICAM-1-deficient, and P-selectin-deficient mice was attenuated compared with wild-type mice. These findings indicate that chronic deficiency of some of the adhesion glycoproteins that mediate leukocyte recruitment alters basal and induced surface expression of other adhesion molecules on endothelial cells.

Role of Platelets in Hypercholesterolemia-Induced Leukocyte Recruitment and Arteriolar Dysfunction

Objective: To define the contribution of platelets, specifically platelet‐associated P‐selectin, to the altered venular and arteriolar responses induced by hypercholesterolemia.