摘要:

ABSTRACT—Sepsis induces extensive lymphocyte cell death that may contribute to immune depression and morbidity/mortality in the disorder.bcl‐2 is a member of a new class of oncogenes that prevents cell death from an array of noxious stimuli. Transgenic mice that overexpress BCL‐2 in T lymphocytes are resistant to sepsis‐induced T cell apoptosis, and mortality was decreased in sepsis. The purpose of this study was to identify key initiator and executioner “caspases” involved in sepsis‐induced lymphocyte apoptosis and to determine if BCL‐2 actspriorto caspase activation. Thymi were removed 5‐22 h post‐cecal ligation and puncture (CLP) or sham surgery. Apoptosis was evaluated in thymocytes by annexin‐V FITC labeling and flow cytometry. Caspase‐1 activity was determined by western blot analysis of the procaspase protein and p20 subunit of the activated caspase; activities of caspases ‐2, ‐6, and ‐9 were determined by colorimetric assays using specific substrates conjugated to a color reporter molecule. Caspase‐3 activity was determined both by western blot and by a fluorogenic assay in which a fluorescent compound was generated. Thymocytes from CLP mice had markedly increased apoptosis and activation of caspases ‐2, ‐3, ‐6, and ‐9 in comparison with thymocytes of sham‐operated mice. Caspase‐1 was not activated. BCL‐2 prevented sepsis‐induced thymocyte apoptosis and inhibited activation of all caspases. We conclude that sepsis causes activation of multiple caspases and that BCL‐2 acts upstream as an inhibitor of caspase activation. The pattern of caspase activation suggests a mitochondrial mediated pathway.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

ABSTRACT—IL‐1&bgr; stimulation of cultured epithelial cells induces the degradation of I&kgr;B&agr; and the consequent nuclear translocation of NF‐&lgr;B, a critical proinflammatory transcription factor in the mucosal host immune response. The role of reactive oxygen intermediates, serine protease activity, and tyrosine kinase activity in the activation of NF‐&kgr;B is weakly conserved across various cell lineages and has not been defined in human enterocytes, a major target of oxidant stress in sepsis, thermal injury, and hemorrhagic shock. We report here that in Caco‐2BBe cells, a transformed human colon cancer cell line with features of small intestinal epithelial cells in culture, exposure to oxidant stress (hydrogen peroxide 1‐10 mM) did not induce NF‐&kgr;B activation. Similarly, scavenging of free radicals and oxidants by pyrrolidine dithiocarbamate and dimethyl sulfoxide did not block IL‐1&bgr;‐induced I&kgr;B&agr; degradation and NF‐&kgr;B activation. Genistein, a nonspecific tyrosine kinase inhibitor, also had no effect on IL‐1&bgr;‐mediated effects on NF‐&kgr;B. Serine protease inhibition by tosyl‐lysine‐chloromethylketone and tosyl‐phenylalanine‐chloromethylketone inhibited I&kgr;B&agr; degradation and NF‐&kgr;B activation stimulated by IL‐1&bgr;. Our data highlight the strong divergence between epithelial and mononuclear cells in the signal transduction pathways relating IL‐1&bgr; stimulation and NF‐&kgr;B nuclear translocation.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

ABSTRACT—This study investigates the effects of N‐acetylcysteine (NAC) and rutin on the lung oxidative burden of patients with early adult respiratory distress syndrome (ARDS). The protection was evaluated by measuring expired ethane and malondialdehyde (MDA), and oxidized (GSSG) and reduced glutathione (GSH) in the epithelial lining fluid of 36 patients who developed ARDS less than 24 hours before enrollment in the study. The patients were randomly assigned to 3 groups, receiving 250 mL 5% dextrose in water (group 1), NAC 50 mg/kg body weight in 5% dextrose (group 2), and NAC 50 mg/kg + rutin 5 mg/kg in 5% dextrose (group 3). Ethane and MDA concentrations were significantly reduced in the treatment groups after day 6. GSH was 30% increased in the treatment groups. No significant variations were observed in the control group until day 9. The trial confirms that NAC and rutin are efficient in protecting the lungs of patients with ARDS.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

ABSTRACT—Induction of the heat shock response may improve outcome from pathophysiological disturbances. This improvement is associated with and believed to result from expression of heat shock protein (HSP)‐70. Therefore, we examined the temporal expression of HSP‐70 in an animal model of acute respiratory distress syndrome (ARDS) secondary to fecal peritonitis. Specifically, we hypothesize that sepsis in rats impairs pulmonary HSP‐70 expression. ARDS was induced in adolescent rats via cecal ligation and double puncture (2CLP). Sham‐operated animals served as controls. Lung tissue was collected 0, 3, 6, 16, 24, and 48 h after 2CLP and sham operation. Northern blot hybridization analysis was performed to detect steady‐state HSP‐70 messenger ribonucleic (mRNA) levels. HSP‐70 protein levels were determined via immunoblotting and immunohistochemistry. Mortality after 2CLP was 50% at 24 h and 75% at 48 h. Northern blot hybridization analysis revealed no significant change in steady‐state HSP‐70 mRNA levels in lung at any time after 2CLP. HSP‐70 steady‐state mRNA levels increased after sham operation and was higher than values in 2CLP at 6, 16, and 24 h. HSP‐70 protein levels did not change over time in either group. Thus, the expression of HSP‐70 does not change after 2CLP. Although lack of an increase in protein levels may be adaptive after sham operation, it is not appropriate after 2CLP. Therefore, failed HSP‐70 expression represents a form of pulmonary epithelial dysfunction that may contribute to lung injury in sepsis.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

ABSTRACT—Reactive oxygen species and peroxidative damage are implicated in the pathophysiology of sepsis. Magnolol is a compound extracted from the Chinese medicinal herbMagnolia officinalisand has multiple pharmacological effects, notably antioxidant functions. To determine whether magnolol can modulate the course of sepsis, survival rate and biochemical parameters were analyzed in rats with sepsis with various treatment protocols. Magnolol at doses ranging from 10‐9g/kg to 10‐5g/kg was administered either before or after induction of sepsis by cecal ligation and puncture. Magnolol did not modulate the course of sepsis induced by two cecal punctures. When one cecal puncture was performed, a moderately evolving type of sepsis was induced, and the survival rate of affected rats was significantly improved by pretreatment with 10‐7g/kg magnolol. The beneficial effect was partially retained if magnolol was administered 6 hours after onset of sepsis when a higher dose (10‐5g/kg) was used. The intensity of lipid peroxidation in plasma, liver, and lung of septic rats was also attenuated in a treatment‐dependent manner. Magnolol at this dose range exerted these beneficial effects probably through its antioxidant efficacy. These significant results may suggest magnolol as a candidate agent for the treatment of sepsis.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

ABSTRACT—Packed red blood cell (PRBC) transfusion has been invoked previously with immunosuppression and increased infections, but it has now been demonstrated that stored PRBCs (>14 days) can prime PMNs and provoke multiple organ failure. Recently, the role of PMNs in the genesis of MOF has been extended to their release of inflammatory cytokines, notably IL‐1, IL‐8, TNF&agr;, and secretory phospholipase A2(sPLA2). We hypothesize that stored PRBCs can act as a second event via stimulating the release of inflammatory cytokines from PMNs. Isolated human PMNs were incubated for 24 h in RPMI with either 20% fresh plasma or plasma from 42 day old PRBC (day of outdate) and release of IL‐8, IL‐1&bgr;, TNF&agr;, and sPLA2were measured. Plasma from stored PRBCs contained small amounts of IL‐8, sPLA2, and TNF&agr; (102.1 ± 5.6 pg/ml, 87.6 ± 6.0 pg/ml and 9.7 ± .7 pg/ml). Levels of IL‐1&bgr; were below detection (<1 pg/ml). Day 42 PRBC plasma stimulated significant PMN release of both IL‐8 and sPLA2as compared to both control and day 0 plasma (*P< .05), but PRBC plasma did not stimulate PMN release of either IL‐1&bgr; or TNF&agr;. Transfused blood is emerging as an inflammatory agent that is capable of producing PMN priming. In this study we have demonstrated that PRBC plasma selectively activates PMNs to release both IL‐8 and sPLA2. Thus, transfusion of PRBCs may represent a preventable inflammatory insult via modification of both blood banking and transfusion practices.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

ABSTRACT—Protein serine/threonine (ser/thr) phosphorylation is an early signaling event in macrophage activation. We investigated the changes in stress‐activated protein kinase/c‐Jun NH2‐terminal kinase (SAPK/JNK) activity and effects of phosphatase inhibition on alveolar macrophage (AM) function in rats challenged with intratracheal endotoxin. Animals were sacrificed 90 min post intratracheal lipopolysaccharide (LPS, 100 &mgr;g/rat) challenge. AMs were incubated with or without phosphatase inhibitors at 37°C for 30 min. Phagocytosis, CD18 expression, SAPK/JNK and phosphatase activities of AMs were determined. LPS challenge activated SAPK/JNK activity and enhanced phagocytosis of AMs without altering phosphatase activity in these cells. Inhibition of phosphatase 1 and 2A activity with okadaic acid and calyculin A exerted a bi‐phasic effect on AM phagocytic function. Okadaic acid at a concentration of 1 &mgr;M increased the mean channel fluorescence intensity (MCF) and the percentage of cells engaged in phagocytosis (percent phagocytosis) in AMs from saline‐treated rats. This inhibitor at concentrations of 0.5 and 1 &mgr;M enhanced both the MCF and percent phagocytosis of AMs from LPS‐challenged rats. Calyculin A at a concentration of 10 nM increased the MCF phagocytosis of AMs from LPS‐challenged rats. At higher concentrations (20 and 30 nM), calyculin A showed a suppression on both the MCF and percent phagocytosis of AMs in both saline and LPS groups. AM CD18 expression was not altered following LPS challenge. Phosphatase inhibitors at doses that enhanced AM phagocytosis showed either no effect (okadaic acid) or inhibition (calyculin A) of AM CD18 expression. These results suggest that ser/thr phosphorylation and dephosphorylation participate in mediating the phagocytic response of AMs to LPS.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

ABSTRACT—Prostaglandin E2production by tissue‐fixed macrophages (MØ) after severe injury contributes to an enhanced susceptibility to infection and sepsis. The purpose of this study was to investigate the effect of cyclic adenosine monophosphate (cAMP) on prostaglandin (PGE2) production and cyclooxygenase II (COX‐2) gene activation in LPS‐stimulated macrophages (MØ). RAW264.7 cells, a mouse MØ cell line, were exposed to various concentrations of dibutyryl cAMP ± lipopolysaccharide (10 &mgr;g/mL) stimulation. Total MØ ribonucleic acid (RNA) was harvested for the determination of COX‐2 messenger RNA (mRNA) with mouse complementary deoxyribonucleic acid (cDNA) by Northern blot assay. MØ supernatant was collected for the measurement of tumor necrosis factor (TNF) by L929 bioassay and PGE2by enzyme‐linked immunosorbent assay (ELISA), respectively. MØ NF&kgr;B activity was determined by electrophoretic mobility shift assays (EMSA). Dibutyryl cAMP significantly inhibited TNF production by LPS‐stimulated MØ. Dibutyryl cAMP (1 mM) alone induced PGE2production. Dibutyryl cAMP (100 &mgr;M and 1 mM) also augmented PGE2production by LPS‐stimulated MØ. Dibutyryl cAMP had similar effect on MØ COX‐2 mRNA expression and NF&kgr;B activity. Our data demonstrate that cAMP modulates MØ TNF production and upregulates COX‐2 gene and PGE2production.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

ABSTRACT—In our study the pathomechanism of sepsis‐induced early myocardial depression was investigated. We determined the effects of the inducible nitric oxide synthase inhibitor and free radical scavenger mercaptoethylguanidine (MEG) on the myocardial contractility, the endothelial and inducible nitric oxide synthase (eNOS and iNOS) activities, and the activation and tissue accumulation of polymorphonuclear leukocytes in hyperdynamic endotoxemia in dogs. Group 1 served as endotoxemic control. Mean arterial pressure and cardiac output were measured, myocardial contractility was estimated from the end‐systolic pressure‐diameter relationship. The eNOS, iNOS and myeloperoxidase activities were determined on myocardial biopsy samples, and the free radical‐producing capacity of granulocytes was measured from separated cells. The effect of MEG on thein vitrofree radical production of isolated granulocytes was measured by chemiluminometry. Endotoxin induced a hyperdynamic circulatory reaction and significant myocardial depression. The myocardial eNOS activity was significantly increased 4 h after induction of endotoxemia and remained elevated, the iNOS activity was increased only 8 h after endotoxemia induction. The free radicalproducing capacity and the myocardial accumulation of the granulocytes were significantly increased. In group 2, MEG treatment selectively inhibited the iNOS activity, prolonged the hyperdynamic circulatory reaction, prevented myocardial depression and decreased the activation and tissue accumulation of granulocytes. The compound dose‐dependently decreased thein vitroactivation of previously resting granulocytes. Our study demonstrates that iNOS do not contribute to the early cardiac failure in endotoxemia. MEG selectively inhibits iNOSin vivo,but its beneficial effects are rather related to the decreases in leukocyte and free radical‐mediated myocardial dysfunction during early endotoxemia.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID

摘要:

ABSTRACT—Sustained whole‐body exposure of anesthetized rats to 35‐GHz radio frequency radiation produces localized hyperthermia and hypotension, leading to circulatory failure and death. The physiological mechanism underlying the induction of circulatory failure by 35‐GHz microwave (MW) heating is currently unknown. We hypothesized that oxidative stress may play a role in the pathophysiology of MW‐induced circulatory failure and examined this question by probing organs for 3‐nitrotyrosine (3‐NT), a marker of oxidative stress. Animals exposed to low durations of MW that increased colonic temperature but were insufficient to produce hypotension showed a 5‐ to 12‐fold increase in 3‐NT accumulation in lung, liver, and plasma proteins relative to the levels observed in control rats that were not exposed to MW. 3‐NT accumulation in rats exposed to MW of sufficient duration to induce circulatory shock returned to low, baseline levels. Leukocytes obtained from peripheral blood showed significant accumulation of 3‐NT only at exposure levels associated with circulatory shock. 3‐NT was also found in the villus tips and vasculature of intestine and within the distal tubule of the kidney but not in the irradiated skin of rats with MW‐induced circulatory failure. The relationship between accumulation in liver, lung, and plasma proteins and exposure duration suggests either that nitro adducts are formed in the first 20 min of exposure and are then cleared or that synthesis of nitro adducts decreases after the first 20 min of exposure. Taken together, these findings suggest that oxidative stress occurs in many organs during MW heating. Because nitration occurs after microwave exposures that are not associated with circulatory collapse, systemic oxidative stress, as evidenced by tissue accumulation of 3‐NT, is not correlated with circulatory failure in this model of shock.

ISSN:1073-2322    

出版商:OVID    

年代:2000

数据来源: OVID