Adenosine A1 receptor antagonist, L-97-1, improves survival and protects the kidney in a rat model of cecal ligation and puncture induced sepsis.

Previously it was reported that combining antibiotics with L-97-1, an adenosine A1 receptor antagonist, significantly improves survival and blocks acute lung injury induced by Yersinia pestis CO 99 in a rat model of pneumonic plague. In the current studies using a conscious rat model of cecal ligation and puncture (CLP) sepsis, L-97-1 was administered in daily intravenous infusions in combination with antibiotics to simulate the use of L-97-1 as an anti-sepsis therapeutic in the clinical setting. In these studies, when administered at 12 h following CLP, in combination with broad spectrum antibiotics, ceftriaxone and clindamycin, L-97-1 improves 7 day (d) survival [25%, 35%, and 75% for L-97-1 (10 mg/kg/h, 12.5 mg/kg/h, and 15 mg/kg/h, respectively) versus (vs.) 25% for antibiotics alone] in a dose-dependent manner. The addition of L-97-1, 15 mg/kg/h to antibiotics significantly increased 7 d survival following CLP compared to therapy with either antibiotics alone (P=0.002) or L-97-1 at 15 mg/kg/h alone (P<0.001) and was not significantly different than survival in sham CLP animals (Log-rank (Mantel-Cox) test with Bonferroni׳s correction for multiple comparisons). Moreover, in these studies, in combination with antibiotics L-97-1 dose-dependently protects the kidney, significantly improving renal function at 24 h post CLP at 10 mg/kg/h (P<0.001), 12.5 mg/kg/h (P<0.0001), and 15 mg/kg/h (P<0.0001) vs. antibiotics alone (ANOVA followed by Tukey׳s post-hoc test for pair-wise comparisons). The results of these studies support efficacy for L-97-1 as an anti-sepsis therapeutic.

A novel post-exposure medical countermeasure L-97-1 improves survival and acute lung injury following intratracheal infection with Yersinia pestis.

Yersinia pestis, a Gram-negative bacillus causing plague and Centers for Disease Control and Prevention (CDC) classified Category A pathogen, has high potential as a bioweapon. Lipopolysaccharide, a virulence factor for Y. pestis, binds to and activates A(1) adenosine receptor (AR)s and, in animals, A(1)AR antagonists block induced acute lung injury (ALI) and increase survival following cecal ligation and perforation. In this study, rats were infected intratracheally with viable Y. pestis [CO99 (pCD1( + )/Δpgm) 1 × 10( 8 ) CFU/animal] and treated daily for 3 d with ciprofloxacin (cipro), the A(1)AR antagonist L-97-1, or cipro plus L-97-1 starting at 0, 6, 24, 48, or 72 h post-Y. pestis. At 72 h post-Y. pestis, cipro plus L-97-1 significantly improved 6-d survival to 60-70% vs 28% for cipro plus H(2)O and 33% for untreated Y. pestis controls (P = 0.02, logrank test). Lung edema, hemorrhage and leukocyte infiltration index (LII) were evaluated histologically to produce ALI scores. Cipro plus L-97-1 significantly reduced lung edema, as well as aggregate lung injury scores vs controls or cipro plus H(2)O, and LII vs controls (P < 0.05, Student's unpaired t test). These results support efficacy for L-97-1 as a post-exposure medical countermeasure, adjunctive therapy to antibiotics for Y. pestis.

Stratified assessment of the role of inhaled hypertonic saline in reducing cystic fibrosis pulmonary exacerbations: a retrospective analysis.

Objective Limited data exist concerning the role of inhaled hypertonic saline (HS) in decreasing pulmonary exacerbations in cystic fibrosis (CF), especially as more advanced stages of CF lung disease were excluded in prior studies. Herein, the authors retrospectively determined the efficacy of inhaled HS in reducing CF pulmonary exacerbations when stratified according to the severity of CF lung disease. Stratification was based on the framework of the Pulmonary Therapeutics Committee's published gradation of obstructive lung physiology in CF, that is, mild (FEV(1) >70%), moderate (FEV(1) 40-70%) and severe (FEV(1) <40%) lung disease, respectively. Design A retrospective review of the Port CF database over a 3-year period performed at an academic CF care centre. Results 340 pulmonary exacerbations were identified; inhaled HS was being used in 99 of these cases. Univariate analysis demonstrated a significant reduction in pulmonary exacerbations only in mild obstruction (OR=0.09, CI 0.01 to 0.81, p=0.012); however, multivariate logistic regression that adjusted for confounding variables showed a reduction in pulmonary exacerbations across the entire spectrum of obstructive lung disease when using inhaled HS, that is, mild obstructive CF lung disease (OR=0.17, CI 0.05 to 0.58, p=0.004), moderate obstructive CF lung disease (OR=0.39, CI 0.16 to 0.93, p=0.034), as well as severe obstructive CF lung disease (OR=0.02, CI 0.001 to 0.45, p=0.015). Moreover, inhaled HS appeared reasonably well tolerated across all stages of lung-disease severity, and was discontinued in only 7% of cases (n=4) with severe lung disease. Conclusion In this study, inhaled HS appeared to reduce pulmonary exacerbations in CF lung disease at all stages of obstruction. This underscores the importance of therapeutic inhaled HS in CF lung disease, regardless of the severity of lung obstruction.

High-frequency percussive ventilation for airway clearance in cystic fibrosis: a brief report.

Exacerbations of cystic fibrosis (CF) lung disease are characterized by increased inspissation of abnormally viscid pulmonary secretions with resultant plugging of small airways, worsened ventilation/perfusion mismatch, and increased physiological deadspace. In this circumstance, hypoxic respiratory failure necessitating mechanical ventilation can be life-threatening. We present such a case of CF lung disease poorly responsive to conventional mechanical ventilatory strategies, in which high-frequency percussive ventilation (HFPV) using volumetric diffusive respiration mobilized copious amounts of inspissated pulmonary secretions and improved refractory hypoxia. Subsequent transient hypercarbia necessitated titrating ventilator parameters to return the PaCO(2) to baseline; the voluminous clearance of secretions and improvement in oxygenation were sustained. HFPV appears unique in its ability to function as a methodological continuum from noninvasive percussion to invasive percussive ventilation for airway clearance, a fundamental tenet of the CF treatment paradigm.

Acute hypoxia decreases E. coli LPS-induced cytokine production and NF-kappaB activation in alveolar macrophages.

Reductions in alveolar oxygenation during lung hypoxia/reoxygenation (H/R) injury are common after gram-negative endotoxemia. However, the effects of H/R on endotoxin-stimulated cytokine production by alveolar macrophages are unclear and may depend upon thresholds for hypoxic oxyradical generation in situ. Here TNF-alpha and IL-1beta production were determined in rat alveolar macrophages stimulated with Escherichia coli lipopolysaccharide (LPS, serotype O55:B5) while exposed to either normoxia for up to 24h, to brief normocarbic hypoxia (1.5h at an atmospheric PO(2)=10+/-2mm Hg), or to combined H/R. LPS-induced TNF-alpha and IL-1beta were reduced at the peak of hypoxia and by reoxygenation in LPS+H/R cells (P<0.01) compared with normoxic controls despite no changes in reduced glutathione (GSH) or in PGE2 production. Both TNF-alpha mRNA and NF-kappaB activation were reduced by hypoxia that suppressed superoxide anion generation. Thus, dynamic reductions in the ambient PO(2) of alveolar macrophages that do not deplete GSH suppress LPS-induced TNF-alpha expression, IL-1beta production, and NF-kappaB activation even as oxyradical production is decreased.

Septic shock and nonpulmonary organ dysfunction in pneumonic plague: the role of Yersinia pestis pCD1- vs. pgm- virulence factors.

OBJECTIVE: Pneumonic plague resulting from Yersinia pestis induces swiftly lethal sepsis and is a major concern as a weapon of bioterrorism. However, the role of specific plasmid-encoded vs. chromosomal Y. pestis virulence factors in the pathogenesis of acute lung injury, shock, and nonpulmonary dysfunction is unclear. We hypothesized that the pathophysiology of pneumonic plague resulting from expression of proteins encoded by the thermally regulated pCD1 plasmid differs from cardiopulmonary and inflammatory changes attributable to the chromosomal pgm gene locus. DESIGN: Prospective, experimental study. SETTING: Research laboratory at a university medical center. SUBJECTS: Conscious, chronically catheterized male Sprague-Dawley rats (total n=104). INTERVENTIONS: Rats were intratracheally infected with 109 colony-forming units of Y. pestis attenuated strains CO99 (pCD1+/DeltaApgm) or KIM6+ (pCD1-/pgm+) and evaluated over 6 days. Serial evaluations of vital signs, cardiorespiratory parameters, blood cultures, inflammatory biomarkers, and organ function were obtained, as well as organ histopathology and cytokine production. MEASUREMENTS AND MAIN RESULTS: Despite equivalent endotoxin contents between the inocula, CO99-infected animals had a median survival of 3 days with greater nonpulmonary organ injury, microbial growth, serum alanine aminotransferase, and liver microvascular permeability vs. KIM6+-infected animals (p<.05). Parallel differences occurred in serum tumor necrosis factor-alpha levels. Notably, 75% of CO99 rats developed fatal hypotension after developing nonpulmonary organ damage. CONCLUSION: These results suggest that progression to lethal sepsis with augmented liver injury during plague pneumonia requires factors encoded by the pCD1 plasmid but not chromosomal proteins present within the pgm gene locus.

Endobronchial leiomyoma: case report and literature review.

Endobronchial leiomyomas are rare benign tumors of the lung, arising from the smooth muscle of the bronchial tree. Symptomatology is based on the degree of endoluminal bronchial obstruction, and surgical resection has generally been the mainstay of treatment. We describe a mechanically ventilated patient with recurrent atelectasis and a postobstructive pneumonia caused by an occlusive endobronchial leiomyoma who was successfully weaned off the ventilator after treatment with argon plasma coagulation delivered via flexible bronchoscopy. We also briefly review the literature.

Idiopathic Progressive Tracheobronchial Stenosis of 20 Years' Duration: Response to Anti-inflammatory Treatment.

We report a unique case of progressive tracheobronchial stenosis in a 52-year-old woman who presented to us with stridor and dyspnea at rest. Her initial symptoms began 20 years earlier, at which time subglottic stenosis of ill-defined etiology necessitated tracheal resection with end-to-end anastomosis. Tracheal biopsy at the time revealed nonspecific inflammation without granulomas, vasculitis, infection, amyloidosis, or malignancy. Over subsequent years, she underwent multiple endobronchial laser resections of the trachea for recurrent disease. On presentation to us, flexible bronchoscopy showed inflammatory stenoses of the left mainstem bronchus and bronchus intermedius. Bronchial biopsy showed acute and chronic stromal inflammation with scattered plasma cells and myofibroblasts against a background of dense fibrosis. Review of the initial tracheal resection specimens and subsequent bronchial specimens revealed areas of high collagenous content with a relatively scant overall myofibroblastic cellular infiltrate; stains for S-100 and anaplastic lymphoma kinase were negative. A diagnosis of idiopathic tracheal stenosis was made with unusual accompanying bronchial involvement, that is, idiopathic tracheobronchial stenosis. Inflammatory airway bronchostenoses were stabilized by high-dose steroids followed by weekly methotrexate therapy, as evidenced by serial flexible bronchoscopies and sequential chest computed tomography with 3-dimensional reconstruction imaging. To our knowledge, this is the first reported case of combined idiopathic tracheal and bronchial stenosis stabilized with anti-inflammatory treatment.

Spinal ceramide modulates the development of morphine antinociceptive tolerance via peroxynitrite-mediated nitroxidative stress and neuroimmune activation.

The effective treatment of pain is typically limited by a decrease in the pain-relieving action of morphine that follows its chronic administration (tolerance). Therefore, restoring opioid efficacy is of great clinical importance. In a murine model of opioid antinociceptive tolerance, repeated administration of morphine significantly stimulated the enzymatic activities of spinal cord serine palmitoyltransferase, ceramide synthase, and acid sphingomyelinase (enzymes involved in the de novo and sphingomyelinase pathways of ceramide biosynthesis, respectively) and led to peroxynitrite-derive nitroxidative stress and neuroimmune activation [activation of spinal glial cells and increase formation of tumor necrosis factor-alpha, interleukin (IL)-1beta, and IL-6]. Inhibition of ceramide biosynthesis with various pharmacological inhibitors significantly attenuated the increase in spinal ceramide production, nitroxidative stress, and neuroimmune activation. These events culminated in a significant inhibition of the development of morphine antinociceptive tolerance at doses devoid of behavioral side effects. Our findings implicate ceramide as a key upstream signaling molecule in the development of morphine antinociceptive tolerance and provide the rationale for development of inhibitors of ceramide biosynthesis as adjuncts to opiates for the management of chronic pain.

Diffuse alveolar hemorrhage following alemtuzumab.

This study describes an unusual patient with X-linked Alport syndrome (XLAS) in whom diffuse alveolar hemorrhage (DAH) developed as a complication of alemtuzumab therapy following renal transplantation. A 26-year-old man with XLAS underwent retransplantation with a cadaveric renal allograft. He received alemtuzumab therapy as a part of an immunosuppressive induction protocol, and dyspnea and hemoptysis developed. A chest CT scan showed diffuse alveolar opacities. Bronchoscopy was performed to determine the cause of hemoptysis and hypoxia. BAL showed a characteristic increasingly bloody return in the sequential aliquots. There was no growth of pathogenic bacteria or evidence of opportunistic infection. Clinical improvement occurred with the initiation of steroids, and the patient required short-term mechanical ventilation for acute respiratory failure. To our knowledge, this is the first reported case of DAH associated with use of alemtuzumab therapy, although other pulmonary toxicities have been described. The prevalence of this form of pulmonary toxicity is unclear and requires further systematic study.

Therapeutic manipulation of peroxynitrite attenuates the development of opiate-induced antinociceptive tolerance in mice.

Severe pain syndromes reduce quality of life in patients with inflammatory and neoplastic diseases, often because chronic opiate therapy results in reduced analgesic effectiveness, or tolerance, leading to escalating doses and distressing side effects. The mechanisms leading to tolerance are poorly understood. Our studies revealed that development of antinociceptive tolerance to repeated doses of morphine in mice was consistently associated with the appearance of several tyrosine-nitrated proteins in the dorsal horn of the spinal cord, including the mitochondrial isoform of superoxide (O2-) dismutase, the glutamate transporter GLT-1, and the enzyme glutamine synthase. Furthermore, antinociceptive tolerance was associated with increased formation of several proinflammatory cytokines, oxidative DNA damage, and activation of the nuclear factor poly(ADP-ribose) polymerase. Inhibition of NO synthesis or removal of O2- blocked these biochemical changes and inhibited the development of tolerance, pointing to peroxynitrite (ONOO-), the product of the interaction between O2- and NO, as a signaling mediator in this setting. Indeed, coadministration of morphine with the ONOO- decomposition catalyst, Fe(III) 5,10,15,20-tetrakis(N-methylpyridinium-4-yl)porphyrin, blocked protein nitration, attenuated the observed biochemical changes, and prevented the development of tolerance in a dose-dependent manner. Collectively, these data suggest a causal role for ONOO- in pathways culminating in antinociceptive tolerance to opiates. Peroxynitrite (ONOO-) decomposition catalysts may have therapeutic potential as adjuncts to opiates in relieving suffering from chronic pain.

Hypoxia-induced mitogenic factor enhances angiogenesis by promoting proliferation and migration of endothelial cells.

Our previous studies have indicated that hypoxia-induced mitogenic factor (HIMF) has angiogenic properties in an in vivo matrigel plug model and HIMF upregulates expression of vascular endothelial growth factor (VEGF) in mouse lungs and cultured lung epithelial cells. However, whether HIMF exerts angiogenic effects through modulating endothelial cell function remains unknown. In this study, mouse aortic rings cultured with recombinant HIMF protein resulted in enhanced vascular sprouting and increased endothelial cell spreading as confirmed by Dil-Ac-LDL uptake, von Willebrand factor and CD31 staining. In cultured mouse endothelial cell line SVEC 4-10, HIMF dose-dependently enhanced cell proliferation, in vitro migration and tubulogenesis, which was not attenuated by SU1498, a VEGFR2/Flk-1 receptor tyrosine kinase inhibitor. Moreover, HIMF stimulation resulted in phosphorylation of Akt, p38 and ERK1/2 kinases in SVEC 4-10 cells. Treatment of mouse aortic rings and SVEC 4-10 cells with LY294002, but not SB203580, PD098059 or U0126, abolished HIMF-induced vascular sprouting and angiogenic responses. In addition, transfection of a dominant-negative mutant of phosphatidylinositol 3-kinase (PI-3K), Deltap85, blocked HIMF-induced phosphorylation of Akt, endothelial activation and tubulogenesis. These results indicate that HIMF enhances angiogenesis by promoting proliferation and migration of endothelial cells via activation of the PI-3K/Akt pathways.

Participation of the PI-3K/Akt-NF-kappa B signaling pathways in hypoxia-induced mitogenic factor-stimulated Flk-1 expression in endothelial cells.

BACKGROUND: Hypoxia-induced mitogenic factor (HIMF), a lung-specific growth factor, promotes vascular tubule formation in a matrigel plug model. We initially found that HIMF enhances vascular endothelial growth factor (VEGF) expression in lung epithelial cells. In present work, we tested whether HIMF modulates expression of fetal liver kinase-1 (Flk-1) in endothelial cells, and dissected the possible signaling pathways that link HIMF to Flk-1 upregulation. METHODS: Recombinant HIMF protein was intratracheally instilled into adult mouse lungs, Flk-1 expression was examined by immunohistochemistry and Western blot. The promoter-luciferase reporter assay and real-time RT-PCR were performed to examine the effects of HIMF on Flk-1 expression in mouse endothelial cell line SVEC 4-10. The activation of NF-kappa B (NF-kappaB) and phosphorylation of Akt, IKK, and IkappaBalpha were examined by luciferase assay and Western blot, respectively. RESULTS: Intratracheal instillation of HIMF protein resulted in a significant increase of Flk-1 production in lung tissues. Stimulation of SVEC 4-10 cells by HIMF resulted in increased phosphorylation of IKK and IkappaBalpha, leading to activation of NF-kappaB. Blocking NF-kappaB signaling pathway by dominant-negative mutants of IKK and IkappaBalpha suppressed HIMF-induced Flk-1 upregulation. Mutation or deletion of NF-kappaB binding site within Flk-1 promoter also abolished HIMF-induced Flk-1 expression in SVEC 4-10 cells. Furthermore, HIMF strongly induced phosphorylation of Akt. A dominant-negative mutant of PI-3K, Deltap85, as well as PI-3K inhibitor LY294002, blocked HIMF-induced NF-kappaB activation and attenuated Flk-1 production. CONCLUSION: These results suggest that HIMF upregulates Flk-1 expression in endothelial cells in a PI-3K/Akt-NF-kappaB signaling pathway-dependent manner, and may play critical roles in pulmonary angiogenesis.

Superoxide potentiates NF-kappaB activation and modulates endotoxin-induced cytokine production in alveolar macrophages.

Gram-negative bacterial infection predisposes to the development of shock and acute lung injury with multiple organ dysfunction in the critically ill. Although overexpression of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha) and interleukin (IL)-1beta, IL-6, IL-8, and other mediators is causally implicated in the pathogenesis of shock and lung injury, the underlying mechanisms following cellular exposure to gram-negative endotoxin remain unclear. De novo generation of reactive oxygen species (ROS) by monocytes/macrophages in particular has been proposed as a pivotal regulatory mechanism by which enhanced transactivation of redox-sensitive genes culminates in augmented cytokine expression within the lower respiratory tract. Here we sought to characterize the mechanism of action of a synthetic, nonpeptide, low-molecular-weight, Mn-containing superoxide dismutase mimetic (SODm), M40403, in modulating E. coli lipopolysaccharide serotype 0111:B4 (LPS)-induced cytokine production by cultured rat alveolar macrophages. Intracellular superoxide (O2) ion generation was measured using hydroethidine (HE) dye, and the dose-dependent effects of M40403 on TNF-alpha and IL-6 biosynthesis by ELISAs. Upstream redox-sensitive signaling events involving the pleiotropic transcription factor NF-kappaB were determined in nuclear extracts by electrophoretic mobility shift assays (EMSAs) and p65 subunit Western blot. The levels of the cytosolic inhibitory protein IkappaB-alpha were also assessed by Western analysis. We found that M40403 potently suppressed the production of superoxide, TNF-alpha, and IL-6 in LPS-stimulated alveolar macrophages, suggesting a key role for superoxide in endotoxin-induced cytokine production in the distal air spaces. In addition, M40403 decreased E. coli LPS-induced activation of NF-kappaB, and this effect was associated with modest suppression of cytoplasmic IkappaB-alpha degradation. Together, these results suggest that removal of superoxide by M40403 inhibits endotoxin-induced production of TNF-alpha and IL-6 in alveolar macrophages by a mechanism involving suppression of redox-sensitive NF-kappaB transactivation or signaling.

Hypoxic suppression of E. coli-induced NF-kappa B and AP-1 transactivation by oxyradical signaling.

Transactivation of the DNA-binding proteins nuclear factor-kappa B (NF-kappa B) and activator protein (AP)-1 by de novo oxyradical generation is a stereotypic redox-sensitive process during hypoxic stress of the liver. Systemic trauma is associated with splanchnic hypoxia-reoxygenation (H/R) followed by intraportal gram-negative bacteremia, which collectively have been implicated in posttraumatic liver dysfunction and multiple organ damage. We hypothesized that hypoxic stress of the liver before stimulation by Escherichia coli serotype O55:B5 (EC) amplifies oxyradical-mediated transactivation of NF-kappa B and AP-1 as well as cytokine production compared with noninfectious H/R or gram-negative sepsis without prior hypoxia. Livers from Sprague-Dawley rats underwent perfusion for 180 min with or without 0.5 h of hypoxia (perfusate PO(2), 40 +/- 5 mmHg) followed by reoxygenation and infection with 10(9) EC or 0.9% NaCl infusion. In H/R + EC livers, nuclear translocation of NF-kappa B and AP-1 was unexpectedly reduced in gel shift assays vs. normoxic EC controls, as were perfusate TNF-alpha and IL-1 beta levels. Preceding hypoxic stress paradoxically increased postbacteremic reduced-to-oxidized glutathione ratios plus nuclear localization of I kappa B alpha and phospho-I kappa B alpha, but not JunB/FosB profiles. Notably, xanthine oxidase inhibition increased transactivation as well as cytokine production in H/R + EC livers. Thus brief hypoxic stress of the liver before intraportal gram-negative bacteremia potently suppresses activation of canonical redox-sensitive transcription factors and production of inflammatory cytokines by mechanisms including xanthine oxidase-induced oxyradicals functioning in an anti-inflammatory signaling role. These results suggest a novel multifunctionality of oxyradicals in decoupling hepatic transcriptional activity and cytokine biosynthesis early in the posttraumatic milieu.

Cocaine enhances susceptibility to endotoxemic shock in a subset of rats.

OBJECTIVE: We hypothesized that the sympathomimetic cocaine may alter cardiovascular and inflammatory responses and enhance susceptibility to endotoxemia due to innate differences in patterns of sympathetic and cardiovascular responsiveness. DESIGN: Prospective study. SETTING: Experimental animal laboratory. SUBJECTS: Fifty-six conscious, instrumented albino rats. INTERVENTIONS: Rats were instrumented for determination of arterial pressure and intravenous drug administration and, in some rats, for cardiac output. After recovery, rats were given cocaine (5 mg/kg i.v., twice daily with 4-6 trials) to identify one of two hemodynamic response patterns: a) an increase in systemic vascular resistance with cardiac depression (vascular responders) or b) smaller increases in systemic vascular resistance and no change or an increase in cardiac output (mixed responders). At least 1 month after characterizing response patterns to cocaine, animals were pretreated with cocaine (5 mg/kg i.v.) or an equivalent bolus of vehicle (0.9% saline) while recording hemodynamics. Five minutes later, Escherichia coli lipopolysaccharide (serotype O55:B5, 20 mg/kg i.v.) was administered for 15 mins. MEASUREMENTS AND MAIN RESULTS: Hemodynamic responses, pupillary diameter, and serum cytokines were determined at several time points. Lipopolysaccharide administration (5-40 mg/kg) without cocaine produced dose-dependent depressor responses with recovery typically within 2 hrs. Although 87% of rats survived a single 20 mg/kg dose of lipopolysaccharide when given alone, pretreatment of vascular responders with cocaine before lipopolysaccharide resulted in greater increases in systemic vascular resistance and pupillary mydriasis and lethality in five of six vascular responders, whereas only one of six mixed responders died. Pretreatment with the alpha1-adrenoceptor antagonist prazosin (0.1 mg/kg i.v.) before cocaine and lipopolysaccharide attenuated hemodynamic responses and improved survival among vascular responders. Serum interleukin-6 and interleukin-10 were elevated in rats treated with cocaine and lipopolysaccharide compared with rats treated with lipopolysaccharide alone, whereas serum tumor necrosis factor-alpha was reduced by cocaine pretreatment. Moreover, serum interleukin-1beta, tumor necrosis factor-alpha, and interleukin-6 were elevated in nonsurvivors compared with survivors after cocaine and lipopolysaccharide administration. CONCLUSIONS: We conclude that cocaine enhances susceptibility and worsens outcome from endotoxic shock by augmenting sympathetic activity, particularly in vascular responders, and that alpha-adrenoceptors mediate the altered inflammatory responses.

Modulation of serum cytokine levels by a novel superoxide dismutase mimetic, M40401, in an Escherichia coli model of septic shock: correlation with preserved circulating catecholamines.

OBJECTIVES: We have shown previously that inactivation of catecholamines by superoxide anions contributes to the loss of vascular reactivity to norepinephrine and the subsequent hypotension that develops in Gram-negative endotoxic shock. In addition to their vasopressor actions, catecholamines, via beta-adrenoceptor activation, are important regulators of cytokine production. Here we examined if maintenance of serum catecholamine levels by the superoxide dismutase mimetic, M40401, modulates serum cytokine levels and arterial hypotension in an Escherichia coli-infected conscious rat model of septic shock. DESIGN: Controlled laboratory animal study. SETTING: University animal research laboratory. SUBJECTS: Pathogen-free male Sprague-Dawley rats (n = 51). INTERVENTIONS: Conscious, antibiotic-treated animals with chronic in-dwelling carotid arterial and jugular venous catheters were intravenously infected with 10(10) live E. coli bacteria (O55:B5, n = 51) over 30 mins, ending at time = 0 hrs. At 0.5 or 3 hrs, infected rats were administered an intravenous infusion of either M40401 (n = 33) or 0.9% saline (n = 18) for 6 hrs at a rate of 1 mL/h. In additional experiments, anesthetized animals with catheterized left femoral arteries and veins were administered a dose-range of norepinephrine (0.1-1 microg/kg) as bolus intravenous injections. Thereafter, E. coli lipopolysaccharide (4 mg/kg, n = 6) was administered as a 0.3-mL slow bolus intravenous injection. One hour later, the norepinephrine protocol was repeated, after which the rats were administered an intravenous infusion of either M40401 or 0.9% saline for 15 mins. At 2 hrs, the dose response to norepinephrine was repeated. MEASUREMENTS AND MAIN RESULTS: Rats infected with live E. coli exhibited a biphasic fall in mean arterial pressure, with mortality reaching 83% by 24 hrs. Rats treated with M40401 (0.25, 2.5, or 25 microg x kg-1 x hr-1 ) 3 hrs after bacteremic sepsis maintained a normal mean arterial pressure, and mortality was dose-dependently reduced to 44, 33, and 22%, respectively, at 24 hrs. Furthermore, serum catecholamine levels were diminished in E. coli-infected rats treated with saline compared with rats treated with M40401. In separate experiments, E. coli-infected rats were administered M40401 (25 microg x kg-1 x hr-1 ) 0.5 hr after bacterial challenge. Blood samples taken at 0, 1.5, 3.5, and 6 hrs were analyzed for tumor necrosis factor-alpha, interleukin (IL)-1 beta, IL-6, and IL-10 and for norepinephrine and epinephrine. Serum levels of tumor necrosis factor-alpha and IL-1 beta were significantly depressed in M40401-treated septic rats, whereas IL-10 was elevated. Moreover, serum catecholamine levels were greater in M40401-treated septic rats at the same time points. IL-6 levels were unaffected by M40401 treatment. Finally we examined whether treatment with M40401 could reverse the hyporeactivity to norepinephrine typifying early septic shock. Using the E. coli lipopolysaccharide (4 mg/kg) challenged anesthetized rat model of shock, we demonstrated that the vasoconstrictor ability of norepinephrine was indeed restored after M40401 treatment (25 microg/kg). CONCLUSION: Postinfection treatment with the superoxide dismutase mimetic M40401 protects against hypotension, vascular hyporeactivity to catecholamines, and mortality associated with septic shock. Such beneficial effects correlate with both reduced oxidative inactivation of serum catecholamines and a reduction in canonical cytokine mediators of inflammation.