Pretreatment of mice with clindamycin improves survival of endotoxic shock by modulating the release of inflammatory cytokines

Suppression of endotoxin release and subsequent production of inflammatory cytokines is crucial in the treatment of septic shock. We investigated the effect of clindamycin (CLI) on endotoxic shock induced in mice by Escherichia coli lipopolysaccharide (LPS). Mice were treated with CLI (160 to 600 mg/kg) or saline and then injected with E. coli LPS and D-(+)-galactosamine intraperitoneally 0.5 h after CLI administration. Pretreatment with CLI significantly improved survival in a dose-dependent manner (CLI, at 160, 300, and 440 mg/kg) and significantly lowered the peak concentrations of tumor necrosis factor alpha and interleukin-1beta (IL-1beta) in serum. However, the peak concentrations of IL-6 in the sera of CLI-treated mice were higher than in control mice. Our findings suggest that CLI alters LPS-induced inflammatory cytokine production and suppresses endotoxin-induced mortality in this murine model.     

Protection against lipopolysaccharide-induced death by fluoroquinolones

Because fluoroquinolones have an immunomodulatory effect on cytokine production by lipopolysaccharide (LPS)-treated human monocytes, we examined the effect of fluoroquinolones on the survival of mice injected with a lethal dose of LPS. Trovafloxacin (100 mg/kg), ciprofloxacin (250 mg/kg), and tosufloxacin (100 mg/kg) protected 75% (P = 0.0001), 25% (P = 0.002), and 50% (P = 0.002), respectively, of mice against death. The fluoroquinolones significantly reduced serum levels of interleukin-6 and tumor necrosis factor alpha in LPS-treated mice. The protective effects of fluoroquinolones in LPS-induced shock in mice may also occur in humans.     

Improved bacterial clearance and decreased mortality can be induced by LPS tolerance and is not dependent upon IFN-gamma

Endotoxin (LPS) tolerance is induced by exposure to sublethal doses of LPS, resulting in a suppressed proinflammatory response and an improved survival rate after challenge with a normally lethal dose of LPS. We studied the effects of tolerance induced by either Escherichia coli-derived LPS or Pseudomonas aeruginosa-derived LPS on the innate immune response to a subsequent P. aeruginosa bacterial challenge and determined if the induction of tolerance was dependent on interferon gamma (IFN-gamma) activity. LPS tolerance was induced in wild-type (WT) and IFN-gamma knockout mice by i.p. injection of 1 microg of LPS on 2 consecutive days. Mice were challenged with an i.p. injection of live P. aeruginosa (1 x 10(8) colony-forming units) 2 days after the second LPS dose. LPS tolerance in WT mice was associated with diminished serum IFN-gamma and IL-12 and increased serum IL-10 responses to the Pseudomonas challenge. Both clearance of the bacterial challenge and survival were improved in WT animals pretreated with either E. coli LPS or P. aeruginosa LPS compared with saline-pretreated control mice. Similarly, IFN-gamma knockout mice exposed to LPS before the Pseudomonas challenge also had improved bacterial clearance of the challenge and an improved survival rate. In separate experiments, priming with IFN-gamma at a dose that approximated the serum concentration induced by LPS priming did not alter cytokine production or bacterial clearance after a Pseudomonas challenge. Finally, administration of IFN-gamma at the time of Pseudomonas challenge amplified cytokine production in LPS-tolerant animals but did not affect bacterial clearance. These results suggest that IFN-gamma is not necessary for the induction of LPS tolerance. Furthermore, IFN-gamma seems to play a role in propagating the inflammatory cytokine response to Pseudomonas challenge, but it did not seem to have any role in bacterial clearance.     

In vivo and in vitro effects of fluoroquinolones on lipopolysaccharide-induced pro-inflammatory cytokine production

Fluoroquinolones have been reported to affect cytokine production in vitro. We investigated the effects of fluoroquinolones on lipopolysaccharide (LPS)-induced inflammatory cytokine production in vivo and in vitro. LPS was administered to mice treated with ciprofloxacin, gatifloxacin, norfloxacin, and levofloxacin, and the serum levels of tumor necrosis factor α (TNF-α), interleukin 1β (IL-1β), and interleukin 6 (IL-6) were measured. In addition, peritoneal macrophages collected from mice were treated with the four fluoroquinolones for 1 h, followed by the addition of LPS, and the TNF-α, IL-1β, and IL-6 levels in culture fluid were measured. In LPS-treated mice, ciprofloxacin, gatifloxacin, and norfloxacin (100 mg/kg) significantly reduced the serum TNF-α level (6.8%–63.6% of control). Levofloxacin at 100 mg/kg did not affect the TNF-α level, whereas levofloxacin at a lower dose (10 mg/kg) significantly increased the level. All four fluoroquinolones (100 mg/kg) investigated in this study tended to decrease the serum IL-1β levels (65.5%–65.9% of control), but this was not a significant change. The serum IL-6 levels were increased in ciprofloxacin-administered mice, whereas the other fluoroquinolones did not affect the serum IL-6 levels. In mouse peritoneal macrophages, LPS induced TNF-α, IL-1β, and IL-6 production. Ciprofloxacin, gatifloxacin, and norfloxacin (100 μg/ml) inhibited both TNF-α (12.1%–69.0% of control) and IL-1β production (22.1%–68.8% of control). Levofloxacin (100 μg/ml) inhibited IL-1β production (65.0% of control), but not TNF-α production. LPSstimulated IL-6 production was inhibited only by norfloxacin (59.5 % of control). Our in vivo and in vitro results suggest that fluoroquinolones, especially ciprofloxacin, gatifloxacin, and norfloxacin, which have a cyclopropyl group at the N1 position and/or a piperazinyl group at the C7 position, modify inflammatory responses.     

Lung effects during a generalized Shwartzman reaction and therapeutic intervention with dexamethasone or vitamin E

We investigated if a two-hit shock model, commonly referred to as generalized Shwartzman reaction (GSR), can prime for indirect acute respiratory distress syndrome (ARDS) in mice. The GSR was provoked in C57BL/6 mice by two consecutive i.p. injections of 100 microg lipopolysaccharide (LPS) at t = 0 and t = 20 h. These mice demonstrated a dramatic decrease in respiratory capacity and 80% mortality after the second injection. No such effect was observed when LPS was given as a single 200 microg dose at t = 0. Increased expression of proinflammatory cytokines in serum (interleukin-1beta, interleukin-6 and interferon-gamma), lung neutrophilia, and edema formation were observed in mice injected with one dose of LPS, but notably, mice exposed twice did not further increase their inflammatory response. Early treatment 1 h after the first LPS injection (t = 1 h) with either dexamethasone (10 mg/kg) or vitamin E (50 mg/kg) improved respiratory function and down-modulated the induction of proinflammatory cytokines in serum. In conclusion, mice with a generalized Shwartzman reaction exhibited features resembling some aspects of the pathophysiology in septic ARDS, i.e., neutrophilic inflammation, edema formation, impaired respiratory capacity, and mortality. Our data indicate that a systemic cytokine response and lung neutrophilia may prime for the GSR but that other mechanisms account for the rapid decline in lung function after the second challenge. We suggest that this model can be used for studies of pathogenesis and therapeutic prevention of acute respiratory failure.     

Protective effect of tumor necrosis factor-alpha against subsequent endotoxemia in mice is mediated, in part, by interleukin-10

OBJECTIVE: Tumor necrosis factor (TNF)-alpha administration in large amounts can induce a state of shock similar to that observed in patients suffering from septic shock. Small doses of TNF-alpha induce only mild, transient hemodynamic alterations and can confer protection against subsequent inflammatory stimuli. The objective of this study was to determine whether this protective mechanism could be attributed to activity of the anti-inflammatory cytokine interleukin (IL)-10. DESIGN: Prospective, randomized, controlled study. SETTING: Investigative intensive care unit at a medical university. SUBJECTS: Female BALB-c mice, 10-12 wks of age (approximately 20 g). INTERVENTIONS: All mice were subjected to intraperitoneal (ip) injection of lipopolysaccharide (LPS; Escherichia coli 0111:B4, 125 microg). Mice were randomly assigned to the following groups: TNF-alpha pretreated (100 microg ip 24 hrs before LPS); control (TNF vehicle alone 24 hrs before LPS); TNF/anti-IL-10 pretreated (TNF pretreatment as above and a neutralizing anti-IL-10 antibody); TNF/anti-IL-10 control (TNF pretreatment as above and an isotype-matched control antibody with no IL-10 activity); IL-10 (100 microg ip 1 hr before LPS); and IL-10 control (IL-10 vehicle 1 hr before LPS). MEASUREMENTS AND MAIN RESULTS: Mice were observed for a 48-hr period after endotoxin administration. Mortality in each group was recorded. Separate groups of mice were pretreated with TNF (or vehicle) and killed at 0, 2, or 4 hrs after LPS injection for collection of serum and peritoneal lavage samples that were used to assay IL-10 concentrations. A small dose of TNF-alpha attenuated mortality in mice that were subsequently injected with a highly lethal dose of endotoxin and observed for 48 hrs. Peritoneal lavage fluid concentrations of IL-10 were consistently higher in TNF-pretreated mice after endotoxin administration. The TNF-alpha protective effect was reversed by administration of a neutralizing antibody directed against murine IL-10. CONCLUSIONS: These findings indicate that administration of a low dose of TNF-alpha can induce cross-tolerance to endotoxin by induction of endogenous anti-inflammatory mechanisms.     

类固醇受体辅活化子-3缺失对脂多糖诱导炎症反应的影响

目的 观察类固醇受体辅活化子(SRC)-3蛋白缺失对脂多糖(LPS)诱导炎症反应的影响.方法 健康清洁野生型(SRC-3~(+/+))小鼠、SRC-3基因敲除(SRC-3~(-/-))小鼠各20只,雌性,体质量约20 g,分为SRC-3~(+/+)组和SRC-3~(-/-)组,每组设正常(N)、1h、4h、12h四个时相点,每个时相点各5只小鼠.采用腹腔注射5 mg/kg体质量LPS构建炎症反应动物模型,观察各时相点重要脏器的病理变化并测定血清TNF-α、IL-1β、IL-6和IL-10浓度.结果 LPS腹腔注射后,SRC-3~(-/-)组小鼠全身情况好于SRC-3~(+/+)组小鼠,但两组的肝、脾、肾、心肌和胸腺病理均未见明显差别.LPS腹腔注射后两组血清TNF-α、IL-1β、IL-6和IL-10的水平均显著升高,但SRC-3~(-/-)组小鼠血清TNF-α、IL-1β、IL-6水平显著低于SRC-3~(+/+)组小鼠,IL-10水平却显著高于SRC-3~(+/+)组.结论 SRC-3蛋白与致炎细胞因子的分泌和释放有关,SRC-3蛋白缺失可减轻炎症反应程度,抑制LPS诱导的TNF-α、IL-1β和IL-6分泌和释放.     

Pharmacokinetics of tulathromycin in healthy and neutropenic mice challenged intranasally with lipopolysaccharide from Escherichia coli

Tulathromycin represents the first member of a novel subclass of macrolides, known as triamilides, approved to treat bovine and swine respiratory disease. The objectives of the present study were to assess the concentration-versus-time profile of tulathromycin in the plasma and lung tissue of healthy and neutropenic mice challenged intranasally with lipopolysaccharide (LPS) from Escherichia coli O111:B4. BALB/c mice were randomly allocated into four groups of 40 mice each: groups T-28 (tulathromycin at 28 mg/kg of body weight), T-7, T7-LPS, and T7-LPS-CP (cyclophosphamide). Mice in group T-28 were treated with tulathromycin at 28 mg/kg subcutaneously (s.c.) (time 0 h). The rest of the mice were treated with tulathromycin at 7 mg/kg s.c. (time 0 h). Animals in dose groups T-7-LPS and T7-LPS-CP received a single dose of E. coli LPS intranasally at -7 h. Mice in group T7-LPS-CP were also rendered neutropenic with cyclophosphamide (150 mg/kg intraperitoneally) prior to the administration of tulathromycin. Blood and lung tissue samples were obtained from 5 mice from each dose group at each sampling time over 144 h after the administration of tulathromycin. There were not statistical differences in lung tissue concentrations among groups T-7, T-7-LPS, and T7-LPS-CP. For all dose groups, the distribution of tulathromycin in the lungs was rapid and persisted at relatively high levels during 6 days postadministration. The concentration-versus-time profile of tulathromycin in lung tissue was not influenced by the intranasal administration of E. coli LPS. The results suggest that in mice, neutrophils may not have a positive influence on tulathromycin accumulation in lung tissue when the drug is administered during either a neutrophilic or a neutropenic state.     

Low-density lipoprotein receptor-deficient mice are protected against lethal endotoxemia and severe gram-negative infections.

Lipoproteins can bind lipopolysaccharide (LPS) and decrease the LPS-stimulated production of pro-inflammatory cytokines. We investigated the effect of increased plasma concentrations of low-density-lipoproteins (LDL) on survival and cytokine production after a lethal challenge with either LPS or live Gram-negative bacteria in LDL receptor deficient mice (LDLR-/-). The LDLR-/- mice challenged with LPS had an eightfold increased LD50 when compared with the wild type controls (C57Bl/6J), while tumor necrosis factor alpha (TNFalpha) and interleukin-1 alpha (IL-1 alpha) plasma concentrations were decreased twofold. LDLR-/- mice had significantly lower and delayed mortality than control mice after infection with Klebsiella pneumoniae. No differences in the outgrowth of bacteria in the organs were present between the two groups, while circulating cytokine concentrations were decreased twofold in LDLR-/- mice. In contrast, the LPS-stimulated in vitro production of cytokines by peritoneal macrophages of LDLR-/- mice was significantly increased compared with controls. This increase was associated with enhanced specific binding of LPS to the macrophages of LDLR-/- mice. In conclusion, endogenous LDL can protect against the lethal effects of endotoxin and Gram-negative infection. At least part of this protection is achieved through decreased in vivo production of pro-inflammatory cytokines, in spite of increased cytokine production capacity.     

Impact of ciprofloxacin on theophylline clearance and steady-state concentrations in serum.

The effect of a multiple-dose regimen of oral ciprofloxacin (750 mg every 12 h for 11 doses) on the clearance and steady-state concentrations of theophylline in trough (predose) serum was evaluated in nine healthy male subjects, each serving as his own control. Theophylline was taken as a sustained release tablet per os in a dose of 200 mg every 12 h for 19 doses. Theophylline concentrations in serum were measured immediately before each theophylline dose. Ciprofloxacin was administered on study day 4 through the first dose of study day 8. Theophylline concentrations in serum were also measured on study days 3, 6, 8, and 10 at the following times after the first dose of each day: 0, 0.25, 0.50, 1, 2, 4, 6, 8, 10, and 12 h. Steady-state theophylline concentrations in trough serum were significantly higher during ciprofloxacin treatment (day 8) than before (day 3) or after (day 10) ciprofloxacin administration (P less than 0.01). Likewise, theophylline clearance was significantly slower (P less than 0.01) during ciprofloxacin treatment (day 8) than before it (day 3) or after it (day 10). The magnitude of ciprofloxacin-induced changes was approximately 30%. These results suggest that a multidose regimen of ciprofloxacin significantly slows the clearance of theophylline and elevates theophylline concentrations in serum.     

Multiple-dose pharmacokinetics and safety of ciprofloxacin in normal volunteers

The multiple-dose pharmacokinetics and safety of ciprofloxacin, a new quinoline carboxylic acid derivative, were evaluated in normal volunteers. The drug was administered orally every 12 h during successive 7-day periods at doses of 250, 500, and 750 mg. Samples of serum, urine, and saliva obtained after the first dose on days 1, 4, and 7 of each dosing period were assayed by microbiological methods. Peak concentrations of ciprofloxacin in serum were achieved generally from 1 to 1.5 h after administration. Mean peak serum levels were 1.35 to 1.42 micrograms/ml after the 250-mg dose, 2.60 to 2.89 micrograms/ml after the 500-mg dose, and 3.41 to 4.21 micrograms/ml after the 750-mg dose. Terminal serum half-lives ranged from 3.8 to 4.3, 4.5 to 4.9, and 3.9 to 6.6 h after the 250-, 500-, and 750-mg doses, respectively. Mean concentrations of ciprofloxacin in urine samples collected 0 to 2 h after dosing were 205 to 261, 255 to 518, and 243 to 846 micrograms/ml after the 250-, 500-, and 750-mg doses, respectively. Between 30 and 45% of the dose was recovered in urine 0 to 12 h after drug administration. Mean concentrations of ciprofloxacin in saliva at 2 h after dosing were 0.43, 1.23, and 1.45 micrograms/ml after the 250-, 500-, and 750-mg doses, respectively. These levels were 30 to 45% of the peak levels in serum and between 40 and 65% of the levels in serum measured 2 h after dosing. Ciprofloxacin was well tolerated.     

Edaravone,a novel free radical scavenger,prevents liver injury and mortality in rats administered endotoxin

We postulated that a novel free radical scavenger, 3-methyl-1-phenyl-2-pyrazolin-5-one (edaravone; EDA), would attenuate inflammatory cytokine and chemokine expression in the liver after lipopolysaccharide (LPS) challenge through its antioxidant effect. Rats were administered EDA (0.3, 1.5, 3.0, 6.0, and 12.0 mg/kg) or the same volume of saline intravenously just after LPS (10 mg/kg) injection and then was continued intermittently every 2 h (five administrations in total). Survival was assessed for the next 24 h. In separate experiments, rats were sacrificed at 60 min, 90 min, 6 h, and 9 h after LPS injection. Serum and liver sections were collected for further analysis. Survival was improved by EDA in a dose-dependent manner up to 3 mg/kg, and maximum effects were observed at a dose of 3 mg/kg. After LPS injection, alanine aminotransferase levels increased significantly to about 1,250 IU/l in the vehicle-treated group, whereas values were blunted by about 80% by EDA. Furthermore, increases in 4-hydroxynonenal-modified proteins were also blunted in the liver by EDA. Moreover, mRNA expressions of macrophage infiltrating protein-2, monocyte chemoattractant protein (MCP)-1 and MCP-5 were attenuated by EDA. As a result, increases in the number of infiltrating inflammatory cells and mRNA expression of inflammatory cytokines such as tumor necrosis factor-alpha and interleukin-6 were significantly blunted in the liver by EDA. This reduction was accompanied by a significant reduction of their serum levels. In conclusion, EDA prevented liver injury by both inhibition of recruitments of inflammatory cells and expression of inflammatory cytokine levels in the liver.     

Pharmacokinetics of intraperitoneal ciprofloxacin in patients on CAPD

The concentration of ciprofloxacin was assayed in plasma and peritoneal dialysate following intraperitoneal administration of the drug in the absence of bacterial peritonitis. After administration of a single dose of 5 mg/kg, ciprofloxacin was rapidly absorbed, producing a peak plasma concentration of 1.9 +/- 0.6 mg/l after 3-4 h with an apparent bioavailability of 0.84. In a separate study, following the administration of 25 mg/l for eight consecutive CAPD cycles the intraperitoneal concentration of ciprofloxacin fell to a mean of 8.4 +/- 4.6 mg/l after 4 h cycles and to a mean of 3.0 +/- 3.2 mg/l after 12 h cycles. During the period of administration the mean plasma concentration was 0.5 +/- 0.2 mg/l. Analysis of dialysate for 48 h after cessation of drug administration demonstrated ciprofloxacin to be present in effluent from only two of the six patients, confirming its poor peritoneal elimination.     

Oral administration of geranylgeranylacetone improves survival rate in a rat endotoxin shock model: administration timing and heat shock protein 70 induction

The present study was performed to determine whether oral pretreatment with geranylgeranylacetone (GGA) inhibits proinflammatory cytokine liberation and nitric oxide (NO) production in lipopolysaccharide (LPS)-treated rats and protects rats against death from LPS-induced endotoxin shock, and whether such protection by GGA is related to heat shock protein (HSP) 70 induction in multiple organs of rats. GGA (200 mg/kg) was given orally to rats. LPS (20 mg/kg) was administered intraperitoneally 4, 8, 16, or 24 h after GGA administration. The survival of rats was monitored over 24 h after LPS administration. GGA treatment at 8 or 16 h before LPS dramatically improved the survival rate of LPS-treated rats. Plasma levels of proinflammatory cytokines (tumor necrosis factor-alpha and interleukin-6) and NO 6 h after LPS administration in these GGA-pretreated rats were less than one-half of those in rats treated with LPS alone. A GGA challenge 8 or 16 h before LPS administration enhanced HSP70 expression in rat organs after LPS. Treatment with GGA 8 h before LPS minimized hepatic and renal damage. Furthermore, the protective effect of GGA on mortality in LPS-treated rats was inhibited with quercetin, known as an HSP70 inhibitor. These results suggest that oral administration of GGA at an optimal time before LPS injection induces and enhances HSP70 expression in several organs, inhibits proinflammatory cytokine and NO production, and prevents organ damage, resulting in an improved survival rate.     

Effects of cannabinoid receptor agonist and antagonist ligands on production of inflammatory cytokines and anti-inflammatory interleukin-10 in endotoxemic mice

Previous studies have shown that mice primed with Corynebacterium parvum produce higher levels of inflammatory cytokines than unprimed mice upon challenge with lipopolysaccharide (LPS). Herein, we describe experiments in which two cannabinoid (CB) agonists, WIN 55212-2 [(R)-(+)-[2, 3-dihydro-5-methyl-3-[(4-morpholinyl)methyl]pyrrolo[1,2,3-de]1, 4-benzoxazin-6-yl](1-naphthyl)methanone) and HU-210 [(-)-11-hydroxy-delta(8) tetrahydrocannabinol-dimethylheptyl], were examined for their effects on LPS-induced cytokines in C. parvum-primed and unprimed mice. These agonists have been reported to bind selectively to the CB2 and CB1 receptor subtypes, respectively. WIN 55212-2 (3.1-50 mg/kg i.p.) and HU-210 (0.05-0.4 mg/kg i.p.) decreased serum tumor necrosis factor-alpha and interleukin-12 (IL-12) and increased IL-10 when administered to mice before LPS. The drugs also protected C. parvum mice (but not unprimed mice) against the lethal effects of LPS. The protection afforded to C. parvum mice could not be attributed to the higher levels of IL-10 present in these mice after agonist treatment. The WIN 55212-2- and HU-210-mediated changes in the responsiveness of mice to LPS were antagonized by SR141716A [N-(piperdin-1-yl)-5-(4-chloropheny)-1-(2, 4-dichloropheny)-4-methyl-1H-pyrazole-3-carboxamide hydrochloride], a selective CB1 receptor antagonist, but not by SR144528 [N-[(1S)-endo-1,3,3-trimethylbicyclo[2.2. 1]heptan-2-yl]5-(4-choro-3-methylphenyl)-1-(4-methylbenzyl)p yrazole-3 -carboxamide], a selective antagonist at the CB2 receptor. Therefore, both CB agonists modulated LPS responses through the CB1 receptor. Surprisingly, SR141716A itself modulated cytokine responses in a manner identical with that of WIN 55212-2 and HU-210 when administered alone to mice. The agonist-like effects of SR141716A, which were more striking in unprimed than in primed mice, suggested that the antagonist also could function as a partial agonist at the CB1 receptor. Our findings indicate a role for the CB1 receptor subtype in cytokine modulation by CB ligands.     

Stimulation of Murine Interferon by a Substituted Pyrimidine

2-Amino-5-bromo-6-methyl-4-pyrimidinol (U-25,166) induced high levels of circulating interferon in mice when administered either parenterally or orally. Peak titers of interferon were found in the serum between 6 and 12 h after inoculation of the drug. Lower but significant levels of interferon were found in rat serum after administration of U-25,166 by either the intraperitoneal or oral route, and good levels of circulating interferon were observed in cats after oral treatment. Repeated intraperitoneal doses (50 mg/kg) of U-25,166 protected mice against intranasal encephalomyocarditis virus challenge. The minimal effective acute oral dose for antiviral activity was approximately 250 mg/kg. This was also the minimal dose that produced detectable levels of interferon. Maximum tolerated doses in mice were four to six times the minimal effective doses. A single oral treatment was protective in mice against challenge virus inoculated 24 h later. The compound protected mice from challenge with high levels of encephalomyocarditis virus, up to 20,000 mean lethal doses. Antiviral activity in mice was retained when certain minor substitutions were made in the U-25,166 structure.     

Paeoniflorin attenuates cardiac dysfunction in endotoxemic mice via the inhibition of nuclear factor-κB

The impaired cardiac function caused by reduced myocardial contractility is a typical manifestation of sepsis/septic shock. Paeoniflorin (Pae) has reportedly exhibited anti-inflammatory effect and protection against LPS-induced cardiac dysfunction in mice, but the molecular mechanism is still not fully understood. This study was designed to investigate the protective effects of Pae on lipopolysaccharide (LPS)-induced septic cardiac dysfunction and inflammation response in mice. Mice were intraperitoneal injection with Pae (15 mg/kg) for 3d before the LPS challenge (10 mg/kg, i.p.). Pae significantly protected against LPS-induced cardiac dysfunction and damage. Pae decreased production of inflammatory cytokines, e.g., TNF-α, IL-1β, IL-6, IL-12, MCP-1, IFN-γ, and inducible nitric oxide synthase (iNOS), in the heart of LPS-treated mice. Furthermore, Pae prevented NF-κB activation in endotoxemic mice. Pae pretreatment preserved the level of phospho-Akt. Pae effectively improved cardiac function during endotoxemia in mice. This action is attributed to Pae-induced reduction of inflammatory cytokine release and NF-κB activation, which possibly occurred via the activation PI3K/Akt signaling.     

Upregulation of mouse CD14 expression in Kupffer cells by lipopolysaccharide

Western blot analysis showed that a monoclonal antibody against recombinant mouse CD14 (mCD14), designated rmC5-3, specifically reacted with mouse macrophage cell line J774, but not myeloma cell line NS1. Fluorographic and immunocytochemical analysis demonstrated specific binding of rmC5-3 with mouse resident macrophages, inflammatory monocytes and neutrophils, and macrophage cell lines. Immunohistochemical staining using rmC5-3 showed that CD14-positive Kupffer cells (KC) were small in number in the liver in nonstimulated mice. The number of stained KC, which were rich in the midzonal and periportal regions, gradually increased with time after intraperitoneal injection of lipopolysaccharide (LPS), peaked 6 h after injection, and returned to normal by 20 h after injection. Staining intensity over time was proportional to the number of KC. A slight increase in mCD14 expression was observed in peritoneal macrophages 2 h after LPS administration in vivo using flow cytometric analysis. mCD14 mRNA became detectable at 1 h after the intraperitoneal injection of LPS (20 micrograms/mice), and the level dramatically increased with time, peaking at 3 h, and sharply dropped at 6 h. The resident peritoneal macrophages demonstrated a constitutively high mCD14 mRNA expression, which slightly increased 2 h after LPS (100 ng/ml) stimulation in vitro. The level of mCD14 expression in macrophages did not increase after intraperitoneal injection of LPS (20 micrograms/mice).     

Comparative effects of ciprofloxacin and ceftazidime on cytokine production in patients with severe sepsis caused by gram-negative bacteria

In the present study the effect of ciprofloxacin versus ceftazidime on concentrations of pro- and anti-inflammatory cytokines in the sera of patients with severe sepsis was evaluated. The study included 58 previously healthy patients suffering from severe sepsis caused by gram-negative bacteria, treated with either ciprofloxacin or ceftazidime after thorough clinical and microbiological evaluation and followed up for clinical outcome. Levels of the proinflammatory cytokines tumor necrosis factor alpha (TNF-alpha), interleukin-1b (IL-1b), IL-6, and IL-8 and of the anti-inflammatory cytokine IL-10, as well as of IL-1 receptor antagonist and soluble TNF receptors I and II, in serum were measured at baseline and 24 and 48 h after the first antimicrobial dose. Mean SAPS-II scores, development of septic shock, and mortality rates were similar in the two groups (43.2 +/- 9.2, 21.4%, and 14.3% in the ceftazidime group versus 49.8 +/- 11.3, 20%, and 13.3% in the ciprofloxacin group). Serum TNF-alpha and IL-6 levels at 24 and 48 h were significantly lower in the ciprofloxacin group, while the IL-10/TNF-alpha ratio was significantly higher, than those for the ceftazidime group. Among patients with high baseline TNF-alpha levels, there were significant increases in the IL-10/TNF-alpha ratio at both 24 and 48 h over that at admission for the ciprofloxacin group, while no differences were noted in the ceftazidime group. These results indicate that ciprofloxacin may have an immunomodulatory effect on septic patients by attenuating the proinflammatory response, while there is no evidence that differences in the cytokines measured have any impact on the final outcome.