Diminished lung injury with vascular adhesion molecule-1 blockade in choline-deficient ethionine diet-induced pancreatitis

BACKGROUND: Lung injury in severe acute pancreatitis is mediated by infiltrating leukocytes. Our laboratory has previously demonstrated that acute lung injury in acute pancreatitis results in an up-regulation of vascular adhesion molecule-1 (VCAM-1) cell surface receptor expression on pulmonary vascular endothelium and neutrophil sequestration. The objective of this study was to determine whether blocking expression of VCAM-1 in acute pancreatitis would modify acute pulmonary injury. METHODS: Young female mice were fed a choline-deficient ethionine (CDE) supplemented diet to induce acute pancreatitis. After initiation of the diet, one group (acute pancreatitis treated [n = 18]) was treated with blocking doses (2.35 mg/kg) of monoclonal anti-VCAM-1 receptor antibody (Ab) at 48, 96, and 120 hours. A second group (acute pancreatitis treated control [n = 5]) was treated with a similar dose of an isotypic control for VCAM-1 (nonbinding Ab) at the same time points. A third group (acute pancreatitis untreated [n = 12]) received a CDE diet, and a fourth group (control [n = 11]) received standard food with no Ab treatment. All animals were killed at 144 hours. The dual radiolabeled monoclonal Ab method was used to quantitate VCAM-1 cell surface expression in lung tissue. Lung injury was assessed histologically, and apoptosis was detected by transferase-mediated deoxyuridine triphosphate nick end labeling assay. Pulmonary leukocyte sequestration was determined by myeloperoxidase (MPO) assay and CD18 staining. RESULTS: Pulmonary VCAM-1 cell surface expression was significantly increased in animals with acute pancreatitis when compared to controls (P <.001) and was reduced to near control levels in acute pancreatitis treated animals. On histologic examination, treated animals with acute pancreatitis exhibited significantly less lung injury and apoptosis than did untreated animals with acute pancreatitis. Leukocyte sequestration and MPO activity were significantly reduced in the treated animals with pancreatitis compared to untreated animals with pancreatitis (P <.0001) or acute pancreatitis treated controls (P <.03). CONCLUSIONS: Blocking VCAM-1 on pulmonary vascular endothelium decreases leukocyte adherence and recruitment into the lung, hence reducing lung injury in severe acute pancreatitis. Clinically, VCAM-1 antagonism may be an important adjunct to evolving therapy for distant organ injury in severe acute pancreatitis.     

A role for CD54 (intercellular adhesion molecule-1) in leukocyte recruitment to the lung during the development of experimental idiopathic pneumonia syndrome

BACKGROUND: Idiopathic pneumonia syndrome (IPS) is a frequently fatal complication of allogeneic bone marrow transplantation (BMT). IPS is associated with elevated bronchoalveolar lavage (BAL) fluid levels of tumor necrosis factor-alpha and lipopolysaccharide, both of which are potent activators of endothelial cells (ECs). EC expression of the adhesion molecule CD54 (intercellular adhesion molecule [ICAM]-1) has been shown to be a major regulator of pulmonary inflammation in various experimental models. METHODS: Using a well-established murine BMT system in which lung injury and graft-versus-host disease (GvHD) are induced by minor histocompatibility antigenic differences between donor and host, the RNase Protection Assay, mice deficient in ICAM-1 expression, and a monoclonal blocking antibody to ICAM, we evaluated the role of the pulmonary vascular expression of CD54 in the development of IPS. RESULTS: Enhanced pulmonary vascular expression of ICAM-1 coincided with the development of IPS. When ICAM-1 -/- mice were used as allogeneic BMT recipients, IPS severity (measured by lung histopathology, BAL cellularity, and cytokine expression) was significantly reduced compared with wild-type controls. Similar results were also observed when wild-type recipients were treated with a monoclonal blocking antibody to ICAM-1. Surprisingly, ICAM-1 had differential effects on leukocyte infiltration into GvHD target organs; ICAM-1 deficiency had no impact on intestinal histopathology, whereas ICAM-1-/- BMT recipients had significantly enhanced hepatic injury. CONCLUSIONS: These data demonstrate that although the expression of ICAM-1 is critical for the development of IPS, different mechanisms of leukocyte recruitment are operative in other GvHD target organs.     

Role of CO-releasing molecules liberated CO in attenuating leukocytes sequestration and inflammatory responses in the lung of thermally injured mice

BACKGROUND: Acute lung injury and pulmonary inflammatory responses are important complications most frequently encountered in severely burned patients. Polymorphonuclear leukocyte (PMN) sequestration and the subsequent generation of oxidants and inflammatory mediators play the key roles in the pathogenesis of acute lung injury. In this study, we used CO-releasing molecules (CORM-2) to determine whether the CO-releasing molecules-liberated CO could attenuate leukocytes sequestration and the inflammatory response in the lung of thermally injured mice. MATERIALS AND METHODS: Fifty-four mice were assigned to three groups in three respective experiments. In each experiment, mice in sham group (n=6) underwent sham thermal injury, whereas mice in the burn group (n=6) received 15% total body surface area (TBSA) full-thickness thermal injury and mice in CORM-2 group (n=6) underwent the same thermal injury with immediate administration of CORM-2 (8 mg/kg, i.v.). PMN accumulation (MPO assay) in mice lungs and tumor necrosis factor-alpha and interleukin-1beta in BAL fluid, pulmonary edema formation, and wet/dry weight ratios of lung were determined. Activation of NF-kappaB and expression level of ICAM-1 in the lung was assessed. In in vitro experiment, PMN adhesion to experimental mice serum-stimulated mouse lung endothelial cells (MLEC) was assessed. RESULTS: Treatment of thermally injured mice with CORM-2 attenuated PMN accumulation and prevented activation of NF-kappaB in the lung. This was accompanied by a decrease of the expression of ICAM-1. In parallel, PMN adhesion to MLEC stimulated by CORM-2-treated thermally injured mice serum was markedly decreased. Also, CORM-2 markedly decreased the production of inflammatory mediators in BAL fluid without suppressing the permeability of pulmonary microcirculation. CONCLUSIONS: CORM-released CO attenuates the inflammatory response in the lung of thermally injured mice by decreasing leukocyte sequestration and interfering with NF-kappaB activation, protein expression of ICAM-1, and therefore, suppressing endothelial cells' pro-adhesive phenotype.     

Role of integrins and intracellular adhesion molecule-1 in lung injury after intestinal ischemia-reperfusion

BACKGROUND: We tested the hypothesis that lung injury after intestinal ischemia-reperfusion (IR) requires the activation of CD11/CD18 glycoprotein complex and its ligand, intracellular adhesion molecule-1 (ICAM-1), on pulmonary endothelial surface. METHODS: Rats were assigned to one of six groups including sham operation, intestinal IR (60/120 min) and IR plus treatment with one of the following monoclonal antibodies against CD11a, CD11b, CD18, and ICAM-1. Pulmonary microvascular permeability, neutrophil accumulation, and expression of adhesion molecules were evaluated. RESULTS: Intestinal IR resulted in lung injury characterized by a marked increase in microvascular permeability, neutrophil accumulation and upregulated expression of leukocyte integrins and ICAM-1. The increase in pulmonary microvascular permability and neutrophil accumulation elicited by intestinal reperfusion was effectively prevented by administration of blocking antibodies against ICAM-1, CD11, and CD18. CONCLUSIONS: These results indicate that adhesion molecules contribute to the lung injury after intestinal IR. Immunoneutralization of certain of these adhesion molecules may prevent intestinal IR-induced lung injury.     

Route of nutrition influences intercellular adhesion molecule-1 expression and neutrophil accumulation in intestine.

HYPOTHESIS: The levels of intestinal interleukin 10 and interleukin 4, inhibitors of intercellular adhesion molecule-1 (ICAM-1) expression, decline with total parenteral nutrition (TPN). These cytokine changes induced by lack of enteral nutrition may increase ICAM-1 expression, resulting in polymorphonuclear neutrophil accumulation in intestine. DESIGN: Prospective randomized experimental trials. SETTING: Laboratory. MATERIALS: Male mice. INTERVENTIONS: Sixty-three mice were randomized to chow, intravenous TPN, or intragastric TPN. MAIN OUTCOME MEASURES: Experiment 1: After diet manipulation, iodine 125-labeled anti-ICAM-1 antibody and iodine 131-labeled nonbinding antibody were injected to quantify ICAM-1 expression on endothelial cells in the lung, liver, kidney, and small intestine. Measurement of myeloperoxidase was used to quantify polymorphonuclear neutrophil accumulation in the organs. Experiment 2: Intestine was harvested for both ICAM-1 and myeloperoxidase levels after chow refeeding of mice in the intravenous TPN group. RESULTS: In experiment 1, uninjured mice fed intravenous TPN showed significantly increased intestinal ICAM-1 expression and polymorphonuclear neutrophil accumulation with no significant changes in the lung, liver, or kidney. No changes occurred in mice fed chow or intragastric TPN. In experiment 2, reinstitution of enteral feeding returned intestinal ICAM-1 and myeloperoxidase levels to normal. CONCLUSION: Gut changes associated with lack of enteral feeding induce endothelial changes and an immunologic response, which may influence subsequent responses to injury.     

Quantitative measurement of P- and E-selectin adhesion molecules in acute pancreatitis: correlation with distant organ injury

OBJECTIVE: To determine whether expression of P- and E-selectin molecules is associated with the development of systemic organ manifestations in acute pancreatitis (AP). SUMMARY BACKGROUND DATA: Overproduction of inflammatory cytokines in AP induces expression of adhesion molecules, which may lead to increased leukocytic infiltration and tissue damage. Understanding the temporal expression of these molecules could afford better measures for therapeutic intervention. METHODS: Acute pancreatitis was induced in 30-day-old female C57/ bI/6J mice by feeding a choline-deficient/ethionine-supplemented diet (n = 95). Mice were divided into three groups. Group I (n = 35) was used to study the biochemical and histologic manifestations of AP and to evaluate the neutrophilic infiltration by myeloperoxidase activity and immunofluorescence. Groups II (n = 35) and III (n = 25) were used to evaluate expression of P- and E-selectin by the dual radiolabeled monoclonal antibody technique. RESULTS: Biochemical and histologic evidence of AP developed in all mice. The inflammatory cytokine tumor necrosis factor-alpha gradually increased in serum as early as 18 hours, reaching more than 800-fold background levels by 72 hours. Biphasic P-selectin expression in the lung was seen with peaks at 24 and 48 hours; E-selectin expression peaked at 48 hours. CD18-positive leukocytes and increased myeloperoxidase activity in the lung were demonstrated at 24 hours, correlating with the onset of selectin upregulation. Histologic scoring of lung tissue demonstrated mild damage at 24 hours, with progressive injury occurring from 48 to 72 hours. CONCLUSIONS: In AP, the production of inflammatory cytokines precedes up-regulation of P- and E-selectin, whose expression coincided with the increased infiltration of CD18-positive cells and neutrophil sequestration in lung tissue. Temporally, these events correlate with evidence of histologic pulmonary injury and underscore the role of adhesion molecules as mediators of pathophysiologic events. This mechanistic pathway may afford novel therapeutic interventions in clinical disease by using blocking agents to ameliorate the systemic manifestations of AP.     

Temporal correlation of tumor necrosis factor-alpha release, upregulation of pulmonary ICAM-1 and VCAM-1, neutrophil sequestration, and lung injury in diet-induced pancreatitis

Lung injury is a major cause of patient morbidity in acute pancreatitis. The purpose of this study was to examine the mechanism of pulmonary infiltration and lung injury in acute pancreatitis. Mice were fed a choline-deficient/ethionine-supplemented (CDE) diet for 144 hours to induce severe acute pancreatitis. Serum samples were collected for measurement of biochemical markers of disease and for the detection of tumor necrosis factor-alpha (TNF-α). Cell surface adhesion molecule expression was quantified by the sensitive radiolabeled dual monoclonal antibody technique. Neutrophil sequestration in lung tissue was measured by the myeloperoxidase assay. Lung injury was determined histologically and lung edema was assessed by wet/dry ratios. Pancreatic injury was demonstrated to occur in all CDE-fed mice, which developed significant hyperamylasemia and hypoglycemia by 48 hours (P <0.0001). Serum TNF-a levels increased significantly by 48 hours over baseline values (P < 0.02). E x p ression of intracellular adhesion molecule (ICAM-1) in pulmonary endothelia was significantly increased above baseline by 30% at 48 hours (P < 0.02) and peaked at 120 hours by 100% (P < 0.000l). Vascular cellular adhesion molecule (XXVI-l) was constitutively expressed at baseline and was upregulated threefold by 48 hours (P < 0.000l). Neutrophil infiltration increased gradually 24 hours after ICAM- and VCAM-1 were upregulated with significant elevation of myeloperoxidase activity over baseline at 72 hours (7.2 ±1.2 vs. 18.1 ±2.2 activity units/gram tissue; P < 0.05). Neutrophil infiltration peaked at 144 hours (26.24 ±10.49 activity units/gram tissue P <0.000l), and its kinetics correlated with the onset and progression of morphologic injury as well as increased lung edema. These results show that acute pancreatitis is associated with a systemic release of inflammatory cytokines, followed by increased expression of pulmonary ICAM- and VCAM-1, neutrophil infiltration, and histologic lung injury. The adhesion molecule axis may be a potential target for practical intervention to ameliorate lung injury and morbidity in acute pancreatitis. Supported by the Transplant Surgical Immunology Laboratory at the University of Tennessee at Memphis and by grant PO I DK-43 785 from the National Institutes of Health (Dr. Granger). Presented at the SSAT/Ross Resident Research Symposium at Howey-in-the-Hills, Fla., (oral presentation), and the Fortieth Annual Meeting of The Society for Surgery of the Alimentary Tract, Orlando, Fla., May 16–19, 1999.     

Leukocyte CD18 receptors may be a better target than ICAM-1 ligands for reducing histologic evidence of cellular and vascular rejection in the rabbit

Building evidence suggests that blocking the ICAM-1/CD18 interaction may affect the course of graft rejection. Treatment with monoclonal antibody (mAb) to CD18 was compared to antibody to ICAM-1 in a rabbit heterotopic heart transplant model to determine whether blocking the leukocyte receptor for ICAM-1, CD18, was more effective than antibody targeting of the ligand for ICAM-1. Following transplantation, 28 recipient rabbits were randomized to receive either placebo, mAb to CD18, or mAb to ICAM-1 for 7 days until sacrifice. The cellular rejection grade and percentage of arteries with vascular rejection were significantly lower in animals treated with anti-CD18 than with anti-ICAM-1. As assessed by histology, antibody treatment was more effective in reducing both cellular and vascular rejection when directed at the leukocyte receptor CD18 than the ICAM-1 ligand. These findings suggest that other ICAM ligands may play an active role in the immune response and that CD18 may be important for migration of lymphocytes through myocardium.This work was presented at the 29th Annual Meeting of The International Society for Heart and Lung Transplantation, Boca Raton, Aril 1993.     

Role of tumour necrosis factor in lung injury caused by intestinal ischaemia-reperfusion

BACKGROUND: Despite the well known inflammatory effects of tumour necrosis factor alpha (TNF), the mechanism of TNF-mediated lung injury following ischaemia-reperfusion (I/R) is still unclear. In this study, the role of TNF in the development of acute lung injury following intestinal I/R was investigated. METHODS: Male Wistar rats underwent either sham operation (n = 10), 1 h of superior mesenteric artery occlusion and 2 h of reperfusion (I/R, n = 10), or pretreatment with anti-TNF polyclonal antibody 2 mg/kg and I/R (n = 6). Lung injury was evaluated by Evans blue dye concentration, immunohistochemical staining and morphometric analysis. Intestinal injury was assessed by Evans blue dye concentration and histological examination. RESULTS: Intestinal I/R resulted in lung injury characterized by an increase in Evans blue dye concentration, neutrophil sequestration, and obvious staining for expression of pulmonary CD11b and CD18. Pretreatment of animals with anti-TNF antibody led to a reduction in the sequestration of neutrophils, and a decrease in expression of pulmonary intracellular adhesion molecule 1 and CD18. Anti-TNF antibody pretreatment also reduced the intestinal microvascular injury but not histological grade after intestinal I/R. CONCLUSION: Treatment with an anti-TNF antibody resulted in a significant attenuation of lung injury following intestinal I/R. The data indicate that TNF is an important trigger for upregulation of pulmonary endothelial and neutrophil adhesion molecules after intestinal I/R.     

Administration of either anti-intercellular adhesion molecule-1 or a nonspecific control antibody improves recovery after traumatic brain injury in the rat

Intercellular adhesion molecule-1 (ICAM-1) is an endothelial protein that facilitates invasion of leukocytes into the CNS in response to injury or inflammation. ICAM-1 expression correlates with the severity of clinical head injuries, but its importance in secondary injury events is not fully understood. Therefore, we evaluated ICAM-1 expression and the effect of anti-ICAM-1 treatment on motor recovery and neutrophil invasion after traumatic brain injury induced via the lateral fluid-percussion method in the rat. ICAM-1 was expressed in large and small blood vessels within the injured cortex at 10 and 24 h after injury. Repeated administration of anti-ICAM-1 antibody (clone 1A29) at 1, 10, and again at 24 h after injury significantly improved performance in two of three motor tests, compared to saline controls. Equal doses of nonspecific control antibody (IgG) also significantly improved motor test scores, compared to saline controls. Cortical myeloperoxidase activity, an indicator of neutrophil invasion, was significantly reduced 26 h after injury in animals treated with anti-ICAM-1. Animals treated with IgG showed a trend toward reduction that did not reach significance. These data suggest that ICAM-1 may be involved in neutrophil invasion and neurological dysfunction after TBI, but also implicate a role for a nonspecific antibody effect in improved functional outcome.     

Acute lung injury following reperfusion after ischemia in the hind limbs of rats.

In this study, we proposed that oxygen free radicals participate in the acute pulmonary injury that follows limb ischemia/reperfusion. Using an established model of hind limb ischemia, reproducible lung injury occurred after reperfusion. Lung microvascular permeability was measured with 125I-BSA and increased two-fold after 30 minutes of reperfusion. Pulmonary injury was blocked with DMSO, DMTU, allopurinol, indomethacin, and SOD plus catalase. The degree of pulmonary neutrophil sequestration as assessed by tissue myeloperoxidase activity was significantly diminished in animals pretreated with antioxidants. Pretreatment with indomethacin did not attenuate the neutrophil sequestration within the pulmonary parenchyma. These data suggest that increased lung microvascular permeability and neutrophil accumulation occur following hind limb ischemia/reperfusion. Therapeutic interventions with oxygen radical inhibitors blocked this process, while the prostaglandin inhibitor, indomethacin, only reduced lung permeability.     

Pentoxifylline reduces acute lung injury in chronic endotoxemia

BACKGROUND: Pentoxifylline (PTX) attenuates end-organ injury in models of sepsis and hemorrhage. PTX is thought to act by inhibiting phosphodiesterase, thus increasing cAMP and decreasing tumor necrosis factor-alpha (TNF-alpha) synthesis. The effects of PTX on neutrophil and endothelial cell adhesion molecules and, ultimately, organ injury in a chronic endotoxemia model have not been studied. We hypothesized that continuous infusion of PTX reduces acute lung injury (ALI) caused by chronic lipopolysaccharide (LPS) exposure. MATERIALS AND METHODS: Male Sprague-Dawley rats were given continuous infusion of LPS, PTX + LPS combined, or saline (sham) by implantable pumps. Neutrophil CD11b expression, lung histopathology, lung intercellular adhesion molecule-1 (ICAM-1) expression assessed by immune staining, serum TNF-alpha, serum interleukin-6 (IL-6), and bronchoalveolar lavage (BAL) IL-8 were evaluated at different time points. Lung injury was graded in a blinded fashion from 0 (normal) to 4 (severe) for interstitial inflammation, neutrophil infiltration, congestion, and edema. Total lung injury score (TLIS) was calculated by adding listed categories. White cell count in the peripheral blood and in the BAL was also performed. RESULTS: Animals treated with PTX + LPS showed a significant reduction in lung injury score, a marked decrease in ICAM-1 expression, and a significant decrease in IL-8 levels in the BAL and serum IL-6 levels when compared with LPS-treated animals. CONCLUSIONS: Continuous infusion of PTX reduces ALI caused by chronic endotoxemia. The effect seems to be a result of decreased expression of endothelial and epithelial ICAM-1 and modulation of proinflammatory cytokine synthesis.     

急性坏死性胰腺炎肺组织炎性介质mRNA表达与肺损伤的关系

目的 探讨急性坏死性胰腺炎(ANP)大鼠肺组织白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)及细胞间黏附分子(ICAM-1)等炎性介质mRNA表达与肺损伤的关系.方法 33只Wistar大鼠随机分为正常对照和胰腺炎不同时间点(1、4、12和24 h)各组,应用3.5%牛磺胆酸钠逆行胰胆管注射制备ANP模型.采用RT-PCR法检测ANP肺组织IL-6、TNF-α及ICAM-1 mRNA表达,同时观察血淀粉酶及脂肪酶、胰腺和肺组织湿/干重比率及病理改变.结果 造模ANP 1 h后肺组织IL-6、TNF-α及ICAM-1 mRNA水平(1.25±0.16、0.33±0.09及082±0.03)较正常对照组(0.07±0.02、0.06±0.02及0.41±0.04)表达增高(P<0.05),并持续升高至12及24 h(分别为1.674±0.14、0.99±0.11、1.17士0.05及1.87±0.05、0.96士0.06、1.11士0.04),同时伴有肺组织病理损害,其严重程度与肺TNF-α及ICAM-1 mRNA表达、肺组织湿/干重比率与TNF-α、IL-6、ICAM-1 mRNA表达的相关系数分别为0.93及0.70(P<0.05).结论 大鼠ANP早期肺组织IL-6、TNF-α及ICAM-1mRNA即过度表达,肺IL-6、TNF-α及ICAM-1mRNA过度表达是ANP肺损害发生的原因之一,肺损伤严重程度与IL-6、TNF-α及ICAM-1mRNA表达的高低有关.     

Trigger for intercellular adhesion molecule-1 expression in rat lungs transplanted from non-heart-beating donors

BACKGROUND: Lung transplantation from non-heart-beating donors causes ischemia-reperfusion injury. We sought to determine the trigger for expression of intercellular adhesion molecule-1 (ICAM-1) caused by ischemia-reperfusion injury. METHODS: Thirty-six Sprague-Dawley rats underwent left lung transplant (six groups of 6). Lungs were transplanted immediately after arrest, or from non-heart-beating donors after 2 hours of oxygen-ventilation or no ventilation. Recipients were reperfused for 4 or 6 hours, then lungs were stained with a mouse anti-rat ICAM-1 monoclonal antibody, developed with avidin-biotin peroxidase to a biotinylated anti-mouse immunoglobin G antibody. Intercellular adhesion molecule-1 expression was graded by two masked observers as 0 = absent, 1 = weak, or 2 = strong in alveoli, arterioles, and venules. Explanted recipient left lungs served as negative controls, and positive controls were generated 6 hours after intraperitoneal injection of endotoxin. Intercellular adhesion molecule-1 expression above baseline among groups was compared by Fisher's exact test. RESULTS: Constitutive expression of ICAM-1 was present in rat lung alveoli, with 24 of 35 controls staining weakly and 4 of 35 strongly positive in alveolar areas. Intercellular adhesion molecule-1 expression was not increased in transplanted lungs evaluated after 4 hours of reperfusion, even lungs retrieved from non-heart-beating donors. But when non-heart-beating donor lungs were assessed 6 hours after onset of reperfusion, ICAM-1 expression was significantly more apparent in alveolar and arteriolar areas, compared with controls and lungs transplanted immediately after arrest. CONCLUSIONS: Lungs transplanted immediately after circulatory arrest do not sustain sufficient ischemia-reperfusion injury to upregulate ICAM-1. Onset of reperfusion is the signal for ICAM-1 expression, not the onset of ischemia or the total duration of ischemic and reperfusion time together. Strategies at reperfusion may minimize ICAM-1 expression.     

Hemorrhagic shock induced up-regulation of P-selectin expression is mediated by factors in mesenteric lymph and blunted by mesenteric lymph duct interruption

BACKGROUND: Previous studies have shown that mesenteric lymph duct interruption prevents lung injury and decreases lung neutrophil sequestration after hemorrhagic shock (HS). Since endothelial cells rapidly express P-selectin after ischemia/reperfusion injury and HS-induced lung injury appears to involve neutrophil-endothelial cell interactions, we tested the following two hypotheses. First, that HS increases endothelial cell P-selectin expression and that interruption of mesenteric lymph flow in vivo would diminish this expression. Second, that incubation of human umbilical vein endothelial cells with post-HS mesenteric lymph but not sham shock (SS) lymph or postshock portal vein plasma would up-regulate P-selectin expression. METHODS: Pulmonary microvascular P-selectin expression was measured in male rats subjected to 90 minutes of HS (30 mm Hg), SS, or HS with lymphatic ligation, with a dual radiolabeled monoclonal antibody technique. The lungs from these animals were subsequently harvested and P-selectin expression was expressed as mean +/- SEM nanograms of monoclonal antibody per gram of tissue. RESULTS: Pulmonary P-selectin expression was 2.0 +/- 0.4 after SS, 9.7 +/- 3.0 after HS, but decreased to 2.3 +/- 0.3 after HS with lymph interruption (p < 0.05 HS vs. SS or HS plus lymph ligation). Incubation of human umbilical vein endothelial cells with shock lymph collected 3 to 4 hours after shock resulted in a nearly fivefold increase in P-selectin expression (p < 0.001) as compared with SS lymph, lymph collected 6 hours after shock, or postshock portal vein plasma. CONCLUSION: These results support the concept that gut-derived lymph promotes HS-induced lung injury through up-regulation of microvascular adhesion molecules and that intestinal lymph duct interruption may prevent distant organ injury by blunting the expression of these molecules.     

Lack of enteral feeding increases expression of E-selectin after LPS challenge

BACKGROUND: Total parenteral nutrition (IV-TPN) increases neutrophil accumulation in the small intestine, expression of intestinal ICAM-1 and P-selectin, and upregulates E-selectin expression in the lung. Endothelial activation induced by lack of enteral nutrition may change the response to injury or infection. This study investigated whether nutrition influenced the expression of the adhesion molecule, E-selectin and ICAM-1, following endotoxin challenge. MATERIALS AND METHODS: Forty-three mice were injected with saline, 2, 20, 200, 2000, or 10000 microg/kg lipopolysaccharide (LPS) intraperitoneally. E-selectin expression in the lung, small intestine, and heart was quantified at 3 h after challenge, while ICAM-1 was measured at 5 h, using the dual-radiolabeled monoclonal antibody technique. Next, 80 mice were fed chow, intragastric (IG)-TPN, or IV-TPN for 5 days, and then received intraperitoneal 2 or 200 microg/kg LPS. E-selectin and ICAM-1 expression in organs was measured at 3 and 5 h after endotoxin, respectively. RESULTS: E-selectin expression in organs increased LPS dose dependently. ICAM-1 levels reached early peaks in the lung and in the intestine. Also, IV-TPN significantly increased E-selectin expression in the small intestine and tended to increase pulmonary E-selectin, when compared to chow or IG-TPN animals. There were no significant differences in E-selectin expression among three diet groups after 200 microg/kg LPS challenge. No differences in ICAM-1 expression were observed in any organ among the three groups after 2 or 200 microg/kg LPS injection. CONCLUSIONS: E-selectin rather than ICAM-1, because of the expression pattern after various dosages of LPS challenge, may be a determining factor for the degree of LPS-induced inflammation at the early phase. Lack of enteral nutrition may increase inflammatory response through enhanced gut E-selectin levels after a small dose of LPS.     

肠缺血再灌注损伤大鼠小肠运动功能的变化与细胞间黏附分子1表达的关系

目的 探讨肠缺血再灌注(intestinal ischemia reperfusion,IIR)损伤大鼠小肠运动功能的变化及其与细胞间黏附分子1(intercellular adhesion molecule-1,ICAM-1)表达的关系.方法 将Wistar大鼠30只随机分为ⅡR组、ICAM-1单抗组和假手术组,ⅡR组采用夹闭-放开肠系膜上动脉的方法建立IIR损伤模型,ICAM-1单抗组于阻断肠系膜上动脉血供同时静脉给予ICAM-1单克隆抗体1 mg/kg.用荧光分光光度法测定各组大鼠小肠传输功能,多导生理仪测定环行肌条收缩能力的变化,RT-PCR法检测小肠肌层ICAM-1 mRNA的表达量,免疫组化法检测ICAM-1蛋白表达,图像分析系统计数小肠肌层浸润白细胞数量,比色法测定小肠肌层组织髓过氧化物酶(myeloperoxidase,MPO)的活性.结果 ICAM-1单抗组较ⅡR组大鼠小肠传输功能明显改善,几何中心分别为8.4±0.7和3.4±0.5;两组环形平滑肌条收缩力为(1.81±0.28)g/mm2·s-1和(0.52±0.09)g/mm2·s-1;浸润白细胞数量为(5.6±2.2)c/f和(18.2±3.1)c/f;MPO活性为(2.03±0.56)U/g和(6.41±1.25)U/g,两组各项指标之间比较差异均有统计学意义(P＜O.05).两组小肠肌层ICAM-1 mRNA的表达量为1.69±0.57和1.76±0.32,差异无统计学意义(P＞0.05).IIR组和ICAM-1单抗组小肠肌层均有大量棕色颗粒样染色.结论IIR损伤上调小肠肌层ICAM-1 mRNA的表达,介导中性粒细胞的黏附和浸润,导致小肠运动功能的障碍.

Abstract：

Objective To evaluate the relationship between the intestinal motility alterations and intercellular adhesion molecule-1 ( ICAM-1 ) expression within the rat intestinal muscularis after ischemia reperfusion. Methods Thirty Wistar rats were divided randomly into IIR, monoclanal anti-ICAM-1 and sham group (n = 10), HR models were established by clamping-releasing the superior mesenteric artery. Gut transit was measured in vivo and intestinal circular muscle contractions were measured in an organ bath. The expression of ICAM-1 mRNA was detected by RT-PCR and immunohistochemisty. Leukocyte infiltration and myeloperoxidase (MPO) activity were quantified in sham and ischemia/reperfusion rats with and without monoclonal anti-ICAM-1 antibody treatment. Results The gut transit of monoclonal anti-ICAM-1 group was improved obviously as compared with IIR group. Circular smooth muscle contractility stimulated by ICAM-1 mRNA expression was 1.69 ± 0. 57 and 1.76 ± 0. 32 within the muscularis; Leukocyte infiltration into the muscularis was (5.6 ±2. 2) c/f and ( 18. 2 ±3. 1 ) c/f. MPO activity was (2. 03 ±0. 56) U/g and (6. 41 ± 1.25 ) U/g respectively. The differences were all statistically significant between IIR and treatment groups ( P ＜ 0. 05 ). The expression of ICAM-1 protein in IIR and anti-ICAM-1 groups was not significantly different (P ＞ 0. 05). Conclusions The up-regulated expression of ICAM-1 after ⅡR injury calls forth local infiltration of PMN within the intestinal muscularis, leading to intestinal motility dysfunction.     

粘附分子基因表达在大鼠急性坏死性胰腺炎肺损伤中的作用

目的　研究急性坏死性胰腺炎 (ANP)大鼠肺组织细胞间粘附分子 1(ICAM 1)mRNA的表达情况及其与肺损伤的关系。方法　将 33只Wistar大鼠随机分为正常对照组和胰腺炎不同时间点 (1、4、12和 2 4h)各组 ,用逆行胰胆管注射方法制备ANP模型。采用逆转录聚合酶链反应法检测ANP大鼠肺组织中ICAM 1mRNA表达 ,同时观察肺组织髓过氧化物酶 (MPO)及病理改变。结果造模ANP 1h后大鼠肺组织中ICAM 1mRNA(0 82± 0 0 3)比正常对照组 (0 41± 0 0 4)的表达水平高 (P <0 0 5 ) ,并持续升高至 12h及 2 4h(分别为 1 17± 0 0 5及 1 11± 0 0 4) ,同时伴有肺组织病理损害 ,其严重程度与ICAM 1mRNA表达、MPO及肺MPO与ICAM 1mRNA表达均呈正相关 ,相关系数分别为0 85、0 85及 0 96 (P <0 0 5 )。结论　ANP大鼠早期肺组织中ICAM 1mRNA呈过度表达 ,肺损伤严重程度与ICAM 1mRNA表达的高低有关。肺组织中ICAM 1过度表达及中性粒细胞浸润是ANP肺损害发生的原因之一。     

肠缺血-再灌注大鼠不同组织ICAM-1表达的规律

目的:了解肠缺血-再灌流过程中不同脏器 ICAM-1表达的时间、空间规律。方法:用 RT-PCR 和免疫组化的方法检测肠缺血-再灌流大鼠肠、肝和肺组织中 ICAM-1mRNA 和蛋白表达的变化。结果:肠组织中的ICAM-1 mRNA 和蛋白在肠缺血期及再灌流1h 表达增加,肝脏和肺脏毛细血管内皮上的 ICAM-1表达增加在再灌流后2h 和6h,晚于肠组织,并且与组织中性白细胞的聚集增加一致。结论:肠缺血-再灌流大鼠不同组织的ICAM-1表达的变化呈现序贯性。     

Pulmonary MnSOD is nitrated following hepatic ischemia-reperfusion

BACKGROUND: Ischemia-reperfusion (I/R) of remote organs is a common cause of lung injury. We observed that lung injury after partial hepatic I/R in mice coincides with the appearance of 3-nitrotyrosine (NT) in the lung tissue, a marker of peroxynitrite involvement and oxidant stress. Peroxynitrite can cause mitochondrial dysfunction by inactivation of manganese superoxide dismutase (MnSOD), the major antioxidant enzyme in mitochondria. Our aims were to examine whether pulmonary MnSOD is a target of nitration following hepatic I/R and whether nitrated MnSOD (N-MnSOD) correlates with acute lung injury. METHODS: Five 20-25-g male C57BL/6 mice underwent laparotomy, and atraumatic occlusion of the portal and arterial blood supply to the upper three lobes of the liver for 90 min. This warm ischemic period was followed by 4 h of reperfusion, and the animals were then euthanized. Lung injury was assessed by LDH and protein levels in bronchoalveolar lavage (BAL) fluid. Pulmonary MnSOD activity in pulmonary homogenates was measured by the cytochrome c reduction method. The presence of N-MnSOD was determined by immunoprecipitation (IP) and Western Blot analysis. Controls (N = 5) underwent sham operation. RESULTS: Elevated plasma transaminases confirmed hepatic injury. Lung injury was demonstrated by elevation in BAL protein and LDH levels (495.7 (48.4) versus 644.9 (37.3) [p < 0.05] and 56.5 (11.8) versus 345.2 (80) [p < 0.01], respectively). Immunoprecipitation and Western blot demonstrated N-MnSOD in the lung tissue of I/R animals but not controls. MnSOD activity decreased following I/R (8.1 (0.7) versus 10.8 (0.3) [p < 0.05]). CONCLUSIONS: Pulmonary MnSOD is both nitrated and inactivated following hepatic I/R and is associated with acute lung injury. These findings suggest that MnSOD incapacitance may contribute to I/R-induced lung injury and provide a therapeutic target in attenuating multisystem injury following hepatic I/R.