Injection Of Endotoxin Research Articles

Usefulness of a protease inhibitor (urinastatin) in ARDS with infectious diseases

The usefulness of urinastatin (UST) for adult respiratory distress syndrome (ARDS) induced by gram-negative sepsis was evaluated in clinical and experimental studies. Twelve cases of clinical septic ARDS were treated with combination therapy of UST and methylprednisolone (M-PSL). Ten out of 12 responded favorably. This result was considered to some extent to be better than that of our previous experience with single administration of M-PSL for patients with septic ARDS. Pathophysiologic experiments on UST in endotoxic status were then performed. Immediately after the intravenous administration of endotoxin to rats, serum levels of beta-glucuronidase and elastase released from PMNs were increased and pulmonary edema was observed at 48-hours after the endotoxin injection. Various degrees of pulmonary edema were also observed by the intravenous administration of beta-glucuronidase and PMNs-elastase. These changes induced by the endotoxin were significantly inhibited by the intraperitoneal administration of UST, and they were inhibited more by the combination therapy of UST and M-PSL. In an in vitro study, significantly large amounts of beta-glucuronidase and elastase were released from PMNs by incubating human PMNs with endotoxin. By adding UST to this system, the release of these PMNs proteases was inhibited. These results suggested that UST neutralizes the PMNs-elastase release in the status endotoxemics, and consequently resulted in a better prognosis in cases of septic ARDS.

Effect of endotoxin administration in pregnant camels

Intravenous administration of Escherichia coli endotoxin at a dose of 0.05 μg/kg bodyweight to pregnant camels resulted in abortion. The injection of endotoxin caused significant increases in the plasma concentration of 13,14-dihydro-15-prostaglandin F 2α, the metabolite of prostaglandin F 2α (PG F 2α) and cortisol and a significant decrease in the concentration of progesterone. It is suggested that endotoxin caused abortion in camels was a consequence of endotoxin induced PG F 2α secretion resulting in luteal regression and decreased progesterone concentration.

Protein metabolism in leg muscle following an endotoxin injection in healthy volunteers

The human endotoxin model has been used to study the early phase of sepsis. The aim of the present study was to assess leg muscle protein kinetics after an endotoxin challenge given to healthy human volunteers. Six healthy male subjects were studied in the post-absorptive state before and during 4 h following an intravenous endotoxin bolus (4 ng/kg of body weight). Primed continuous infusion of [(2)H(5)]phenylalanine and [(2)H(3)]3-methylhistidine in combination with sampling from the radial artery, femoral vein and muscle tissue were used to assess leg muscle protein kinetics. Both two- and three-compartment models were used to calculate protein kinetics. In addition 26S proteasome activity and protein ubiquitination were assessed. An increase in the net release of phenylalanine from the leg following the endotoxin challenge was observed; however, this phenylalanine originates from the free intracellular pool and not from protein. Net protein balance was unchanged, whereas both protein synthesis and breakdown were decreased. Degradation rates of contractile proteins were not affected by endotoxin, as indicated by an unchanged rate of appearance of 3-methylhistidine from leg muscle. In addition, proteasome activity and protein ubiquitination were unaffected by endotoxaemia. In conclusion, intravenous endotoxin administration to healthy volunteers resulted in an increased release of free phenylalanine from skeletal muscle, whereas protein balance was unaffected. Both protein synthesis and breakdown were decreased to a similar extent.

EVOLUTION OF PORTAL-SYSTEMIC COLLATERAL VASOPRESSIN RESPONSE IN ENDOTOXEMIC PORTAL HYPERTENSIVE RATS

Cirrhotic patients with portal hypertension and variceal hemorrhage are vulnerable to endotoxemia. However, the direct influence of endotoxemia on portal-systemic collateral vasculature remains unexplored. In this study, portal hypertension was induced in Sprague-Dawley rats by partial portal vein ligation. On the 7th day after portal vein ligation, at 0.5, 1.5, and 5 h post endotoxin (LPS; Escherichia coli serotype O111:B4, 3 mg/kg, i.p., E0.5, E1.5 and E5, respectively) or saline (control, C0.5, C1.5, and C5, respectively) injection, hemodynamic measurements and concentration-response relationships to arginine vasopressin (AVP; 10(-10)-10(-7) mol/L) in collateral vascular bed were obtained. In another six parallel groups, reverse-transcriptase-polymerase chain reaction of iNOS, eNOS, and endothelin 1 (ET-1) mRNA expressions for splenorenal shunt, the most prominent intra-abdominal collateral vessel, was performed. The results showed that E0.5 had lower perfusion pressure changes to AVP and higher splenorenal shunt eNOS expression than C0.5 group (P < 0.05). Compared with C1.5, tachycardia, higher perfusion pressure changes and enhanced splenorenal shunt iNOS and ET-1 expression were observed in E1.5 group (P < 0.05). In E5, systemic and portal hypotension with markedly enhanced collateral AVP responsiveness and splenorenal shunt iNOS and ET-1 expressions were noted (P < 0.05). In conclusion, vasoactive substances counterregulation participates, at least in part, the time-dependent changes of collateral AVP responsiveness in endotoxemic portal hypertensive rats.

Immunological Response to (1,4)‐α‐d‐Glucan in the Lung and Spleen of Endotoxin‐Stimulated Juvenile Rats

We investigated the effects of (1,4)-alpha-D-glucan (alpha-DG), a novel immune stimulatory drug from Tinospora cordifolia, on the concentration of pro- and anti-inflammatory cytokines (interleukin [IL]-1beta, IL-6, tumour necrosis factor-alpha [TNF-alpha], gamma-interferon [IFN-gamma] and IL-10) in the lung and spleen of endotoxin-stimulated juvenile rats. Experimental groups (n = 16/group) included controls with an intraperitoneal injection of saline, endotoxaemic rats with a non-lethal dose of 10 mg/kg Escherichia coli endotoxin, and endotoxaemic rats treated with two doses of 10 mg/kg alpha-DG, intraperitoneally, 2 and 4 hr after endotoxin injection. At 24 hr of treatment, rats were euthanized and lungs and spleen were removed for cytokines determination and lung injury. Endotoxaemia increased IL-1beta concentration by fivefold in both organs, while creating a moderate pulmonary hypercellularity (demonstrated by about 11% increase in the alveolar-septal thickening and 11% decrease in the alveolar-interstitial space ratio). In the lung, alpha-DG treatment reduced concentrations of IL-1beta by 30% (p > 0.05), IL-6 by 43% (p < 0.01), IFN-gamma by 46% (p < 0.01) and the anti-inflammatory cytokine, IL-10, by 31% (p > 0.05) compared to endotoxaemia. In the spleen, alpha-DG treatment decreased the ratio of IL-1beta to IL-10 by 55% (p < 0.05), demonstrating an anti-inflammatory trend. These data suggest that alpha-DG differentially modulates cytokine response in the lung and spleen and modifies the pro- and anti-inflammatory balance during an early period of endotoxaemia in juvenile rats.

Circulating and muscle glutathione turnover in human endotoxaemia

Patients with septic shock have high plasma glutathione concentrations, whereas intracellular concentrations in erythrocytes and muscle are low. In the present study, we investigated the temporal pattern of glutathione status and glutathione kinetics in healthy volunteers during the initial phase of sepsis using a human endotoxin model. The present study was a descriptive pilot study in healthy male volunteers (n=8) before and after an endotoxin challenge. The glutathione status was determined in plasma and whole blood at baseline and hourly for 4 h after intravenous endotoxin injection and in skeletal muscle at baseline and at 2 and 4 h after endotoxin injection. In plasma, the concentration of total glutathione decreased 24% (P<0.05) at 3 h after endotoxin injection and 32% (P<0.001) at 4 h. In whole blood and skeletal muscle, the concentrations of both GSH and total glutathione as well as the redox status remained unaltered during the initial 4 h after the endotoxin challenge. The FSR (fractional synthesis rate) of glutathione in whole blood was 38+/-20%/day before and 59+/-22%/day 4 h after the endotoxin challenge (P=0.088) and in skeletal muscle this was 41+/-25 and 46+/-18%/day (P=0.68) respectively. During the initial phase of sepsis, as represented by an intravenous endotoxin challenge to healthy volunteers, plasma concentrations of total glutathione decreased, whereas glutathione status and synthesis rate in skeletal muscle and whole blood remained unaltered. However, due to the variation in the synthesis measurements, larger studies are needed to confirm these findings.

Systemic Inflammatory Response Reactivates Immune-Mediated Lesions in Rat Brain

The potential association between microbial infection and reactivation of a multiple sclerosis (MS) lesion is an important issue that remains unresolved, primarily because of the absence of suitable animal models and imaging techniques. Here, we have evaluated this question in an empirical manner using immunohistochemistry and magnetic resonance imaging (MRI), before and after the induction of a systemic inflammatory response in two distinct models of MS. In a pattern-II-type focal myelin oligodendrocyte glycoprotein-experimental autoimmune encephalomyelitis model, systemic endotoxin injection caused an increase in regional cerebral blood volume (rCBV) around the lesion site after 6 h, together with a reduction in the magnetization transfer ratio of the lesioned corpus callosum. These changes were followed by an increase in the diffusion of tissue water within the lesion 24 h after endotoxin challenge and new leukocyte recruitment as revealed both immunohistochemically and by MRI tracking of ultrasmall superparamagnetic iron oxide-labeled macrophages. Importantly, we detected in vivo expression of E- and P-selectin in quiescent lesions by MRI-detectable glyconanoparticles conjugated to sialyl Lewis(X). This finding may explain, at least in part, the ability of quiescent MS lesions to rapidly reinitiate the cell recruitment processes. In a pattern-I-type delayed-type hypersensitivity response model, a similar effect of endotoxin challenge on rCBV was observed, together with delayed breakdown of the blood-brain barrier, showing that systemic infection can alter the pathogenesis of MS-like lesions regardless of lesion etiology. These findings will have important implications for the management and monitoring of individuals with MS.

Immunologic Response to (1,4)‐α‐D‐Glucan in the Lung and Spleen of Endotoxin‐Stimulated Rats

We investigated the effects of (1,4)‐α‐D‐glucan (α‐DG), a novel immune stimulator drug from Tinospora cordifolia, on pro‐ and anti‐inflammatory cytokines balance in the lung and spleen of endotoxin‐stimulated rats. Experimental groups (n=16/group) included, controls with an intraperitoneal injection of saline, endotoxemic rats with a non‐lethal dose of 10 mg/kg E‐coli endotoxin, and rats treated with two doses of 10mg/kg α‐DG, intraperitoneally, at 2 and 4‐h after endotoxin injection. A 24‐h after saline/endotoxin injection rats were euthanized and the lung and spleen were removed for cytokine profiling. Endotoxemia increased IL‐1β concentration by about 5‐fold in both organs. The lung and spleen showed different responses to endotoxemia. In the lung, α‐DG treatment created approximately 30% (p&gt;0.05), 43% and 46% (p&lt;0.01) reductions in IL‐1β, IL‐6, and INF‐γ concentration, respectively, as compared to endotoxemia (ANOVA followed by Tukey‐Kramer test). In the spleen, the ratio of IL‐1βto IL‐10 concentration was reduced from 89 to 49 (55%; p&lt;0.05), demonstrating an anti‐inflammatory trend at the 24‐h of the study (p&lt;0.05). These data suggest that α‐DG influences the pro‐ and anti‐inflammatory cytokine balance in the lung and spleen during early endotoxemia.

Transient increase in cytokines and nerve growth factor in the rat dorsal root ganglia after nerve lesion and peripheral inflammation

Inflammatory response occurs, in general, at the peripheral site of injury or irritation and in the corresponding spinal dorsal horn segments. In this study, we show transient increases in the protein concentrations of interleukins 1β, 6, 8 and NGF in lumbar DRGs within hours after sciatic mononeuropathy in the rat. Comparable increases, with variation in temporal patterns, were observed after skin inflammation induced by intraplantar injection of endotoxin or complete Freund's adjuvant. The non-matching temporal profiles of cytokines and NGF increases and those of behavioral hypersensitivity, particularly peak values, suggest that the roles of these proteins are not necessarily pro-nociceptive in the DRGs at the chronic phase of neuropathy.

Human adult stem cells derived from adipose tissue protect against experimental colitis and sepsis

Background and aims:Inflammatory bowel diseases (IBDs) are associated with uncontrolled innate and adaptive immunity against normal constituents, including commensal bacteria and microbial products. Mesenchymal stem cells (MSCs) suppress effector T...

The effect of glutamine infusion on the inflammatory response and HSP70 during human experimental endotoxaemia

IntroductionGlutamine supplementation has beneficial effects on morbidity and mortality in critically ill patients, possibly in part through an attenuation of the proinflammatory cytokine response and a stimulation of heat shock protein (HSP)70. We infused either alanine-glutamine or saline during endotoxin challenge and measured plasma cytokines and HSP70 protein expression.MethodsThis crossover study, conducted in eight healthy young men, was double-blind, randomized and placebo-controlled. It was performed on 2 trial days, separated by a 4-week washout period. The volunteers received an infusion of alanine-glutamine at a rate of 0.025 g/(kg body weight × hour) or saline for 10 hours. After 2 hours, an intravenous bolus of Escherichia coli endotoxin (0.3 ng/kg) was administered. Blood samples were collected hourly for the following 8 hours. HSP70 protein content in isolated blood mononuclear cells (BMNCs) was measured by Western blotting.ResultsPlasma glutamine increased during alanine-glutamine infusion. Endotoxin reduced plasma glutamine during both trials, but plasma glutamine levels remained above baseline with alanine-glutamine supplementation. Endotoxin injection was associated with alterations in white blood cell and differential counts, tumour necrosis factor-α, IL-6, temperature and heart rate, but glutamine affected neither the endotoxin-induced change in these variables nor the expression of HSP70 in BMNCs.ConclusionsEndotoxin reduced plasma glutamine independently of alanine-glutamine infusion, but supplementation allows plasma levels to be maintained above baseline. Glutamine alters neither endotoxin-induced systemic inflammation nor early expression of HSP70 in BMNCs.Trial RegistrationClinicalTrials.gov ID: NCT 00780520.

Comparison of a cytokine adsorbing column and an endotoxin absorbing column for the treatment of experimental endotoxemia

The present study compared the effects of direct hemoperfusion (DHP) using a cytokine absorbing (CTR) column with those using an endotoxin absorbing (PMX) column with endotoxin-induced shock in rats. Thirty-six rats were injected intravenously with endotoxin (15 mg/kg), and then allocated into one of the following three groups: control group, treated without CTR; PMX group, treated with a PMX column for 120 min; CTR group, treated with a CTR column for 120 min. The present study showed that CTR treatment decreased the mortality rate 8 h after endotoxin injection and inhibited inflammatory responses (cytokine, hemodynamics and acidosis) similar to PMX treatment.

ABSTINENCE PROMOTES REGRESSION WHILE CONTINUED ALCOHOL INTAKE PROMOTES PERSISTENCE OF LPS-INDUCED PANCREATIC INJURY IN ALCOHOL-FED RATS

Anecdotal evidence suggests that abstinence from alcohol is an important factor in inhibiting progression in chronic pancreatitis. However, experimental evidence assessing this issue is lacking. Aims: 1) To compare the effects of alcohol withdrawal with those of alcohol continuation on the extent and pattern of pancreatic injury in a novel rat model of alcoholic chronic pancreatitis induced by repeated endotoxin (LPS) injections in alcohol-fed rats. 2) To assess the effect of alcohol ± LPS on pancreatic stellate cell (PSC) apoptosis in vitro. Methods: 1) SD rats fed isocaloric Lieber-DeCarli diets ± alcohol for 10 weeks were challenged with LPS (3 mg/kg; 1 IV injection/week x 3 wks). Alcohol-fed rats were then either switched to a non-alcohol diet or continued on alcohol for 3 wks. Thus, the experimental groups (n = 5 rats/group) included: control diet + LPS (CL); alcohol diet + LPS (AL), alcohol diet + LPS continued on alcohol (AL+A), alcohol diet + LPS switched to control diet (AL+C). H&E sections of the head, body and tail of each pancreas (20 HPF/section) were scored for oedema, acinar vacuolization and necrosis, haemorrhage and inflammation. Pancreatic fibrosis (collagen deposition) was scored (30 HPF/section) using Sirius Red stained sections. 2) Rat PSCs were exposed to ethanol 10mM (E10) ± LPS 1μg/ml (L1) for 48h and apoptosis assessed by Annexin V and Caspase-3 staining. Results: Pancreatic injury and fibrosis were significantly greater in AL rats than in CL rats. Withdrawal of alcohol led to complete normalisation of the pancreas with disappearance of fibrosis, while continuation of alcohol resulted in persistent acinar injury and fibrosis [Histological score (mean ± SEM): CL 1.29 ± 0.51; AL 6.36 ± 0.38*; AL+A 5.6 ± 0.22*; AL+C 0.67 ± 0.15#; *p < 0.001 vs CL; #p < 0.001 vs AL and AL+A. Fibrosis score (% total area, mean ± SEM): CL 0.56 ± 0.15; AL 2.19 ± 0.36ˆ; AL+A 1.62 ± 0.18ˆ; AL+C 0.53 ± 0.12¥; ˆp < 0.05 vs CL; ¥p < 0.01 vs AL and AL+A; n = 5 animals / group]. Both alcohol and LPS significantly reduced PSC apoptosis [% of control (mean ± SEM); E10: 70 ± 10.2; L1: 45.01 ± 11.8; E10+L1: 38.43 ± 9.38; p < 0.03 vs Control and E10+L1 vs E10; n = 3 separate PSC preparations]. Similar results were obtained for Caspase-3. Conclusions: 1) Continuation of alcohol after LPS-induced pancreatic injury in alcohol-fed rats perpetuates the disease. In contrast, withdrawal of alcohol reverses pancreatic damage. 2) Both alcohol and LPS inhibit PSC apoptosis. Our findings are the first to support the concept that abstinence improves patient outcome in chronic pancreatitis. Perpetuation of fibrosis when alcohol is continued may be due to decreased PSC apoptosis.

Integrated Regulation of Toll-like Receptor Responses by Notch and Interferon-γ Pathways

Toll-like receptor (TLR) responses are regulated to avoid toxicity and achieve coordinated responses appropriate for the cell environment. We found that Notch and TLR pathways cooperated to activate canonical Notch target genes, including transcriptional repressors Hes1 and Hey1, and to increase production of canonical TLR-induced cytokines TNF, IL-6, and IL-12. Cooperation by these pathways to increase target gene expression was mediated by the Notch-pathway component and transcription factor RBP-J, which also contributed to lethality after endotoxin injection. TLR- and Notch-induced Hes1 and Hey1 attenuated IL-6 and IL-12 production. This Hes1- and Hey1-mediated feedback inhibitory loop was abrogated by interferon-gamma (IFN-gamma), which blocked TLR-induced activation of canonical Notch target genes by inhibiting Notch2 signaling and downstream transcription. These findings identify new immune functions for RBP-J, Hes, and Hey proteins and provide insights into mechanisms by which Notch, TLR, and IFN-gamma signals are integrated to modulate specific effector functions in macrophages.

Changes of Bcl-2 and Bax expression in white matter of fetal rats after maternal endotoxin administration

Objective To investigate the changes of Bcl-2 and Bax expression in white matter of fetal rats after maternal endotoxin administration. Methods Pregnant rats were randomly divided into two groups: infection group and control group. The infection group was established by intraperitoneal injection of endotoxin in pregnant rats when gestation was 70% complete (15 days). The control group was treated with normal saline. The fetal rats were killed at 2, 4, 12, 24 and 72 hours after maternal endotoxin administration. The pathological changes in placenta and in fetal rat brain were determined by HE staining. Immunohistochemical method was used to detect the expression of Bcl-2 and Bax proteins in fetal rat brains. Results The major pathological changes in fetal rats niter maternal endotoxin administration included neutrophil infiltration in the placenta, weak staining of white matter and focal infarction. After maternal endotoxin administration, the expression of Bcl-2 gradually decreased from 2 h and arrived at the valley at 12 h, while that of Bax gradually increased 2 h and reached a peak at 12 h. Between the endotoxin-gronp and the control group, the number of positive cells of Bcl-2 and Bax in brain of the fetal rat had significant difference at 4, 12, 24 and 72 hours (P 0.05). The ratio of Bax to Bcl-2 in the endotoxingroup was significantly higher than that in the control group at each time point (P<0.01).Conclusion Endotoxin can be used to eatablish intrauterine irfection models and the infection may cause damage to the white matter. Overexpression of Bcl-2 protects cell from apoptosis, but Bax may function as a cell death effector pro-tein. The ratio of Bax to Bcl-2 may play an important role for apoptosis in the lesion of the white matter. Key words: White matter damage; Intrauterine infection; Rat; Apoptosis; Bcl-2; Bax

Differential dose-dependent effects of prednisolone on shedding of endothelial adhesion molecules during human endotoxemia

Low dose prednisolone was shown to be beneficial in the treatment of the Acute respiratory distress syndrome (ARDS) and septic shock. One corticosteroid-induced effect, postulated to mediate corticosteroid-induced anti-inflammatory effects, is decreased expression of adhesion molecules on endothelial cells, thereby preventing leukocyte recruitment at inflammatory sites. The current study aimed to investigate the effect of increasing doses of prednisolone on the release of soluble adhesion molecules in healthy volunteers challenged with endotoxin. Therefore, 32 healthy, male volunteers received prednisolone orally at doses of 0 mg, 3 mg, 10 mg or 30 mg at 2 h before injection of endotoxin prepared from Escherichia coli (4 ng/kg) and levels of soluble E-selectin (sE-selectin), soluble VCAM-1 (sVCAM-1) and soluble ICAM-1 (sICAM-1) were measured. Levels of all markers were increased after induction of endotoxemia. Levels of sE-selectin were inhibited by a dose of 3 mg prednisolone and levels of sVCAM-1 were decreased after a dose of 10 mg. Maximal inhibition of both sE-selectin and sVCAM-1 levels was achieved by the highest dose of prednisolone 30 mg. Remarkably, prednisolone 3 mg potentiated endotoxin-induced sVCAM-1 release. Levels of sICAM-1 were not affected by prednisolone. Together, the data suggest that prednisolone differentially and dose-dependently influences the release of soluble endothelial adhesion molecules during human endotoxemia.

A 20-HYDROXYEICOSATETRAENOIC ACID AGONIST, N-[20-HYDROXYEICOSA-5(Z),14(Z)-DIENOYL]GLYCINE, OPPOSES THE FALL IN BLOOD PRESSURE AND VASCULAR REACTIVITY IN ENDOTOXIN-TREATED RATS

Endotoxic shock is a systemic inflammatory response that is associated with an increase in nitric oxide production and a decrease in the formation of 20-hydroxyeicosatetraenoic acid (20-HETE), which may contribute to the fall in blood pressure and vascular reactivity. The present study examined the effects of a synthetic analogue of 20-HETE, N-[20-hydroxyeicosa-5(Z),14(Z)-dienoyl]glycine (5,14-HEDGE), on the fall in blood pressure and vascular responsiveness to vasoscontrictors and acetylcholine in rats treated with endotoxin. The MAP fell by 31 mmHg, and the heart rate rose by 90 beats/min in male Wistar rats treated with endotoxin (10 mg/kg, intraperitoneally). The fall in MAP was associated with a decrease in the vasoconstrictor response to norepinephrine in isolated aorta and superior mesenteric artery and increased levels of nitrite in the serum, kidney, heart, and vascular tissues. The effects of endotoxin were prevented by 5,14-HEDGE (30 mg/kg, s.c.) given 1 h after injection of endotoxin. Furthermore, a competitive antagonist of vasoconstrictor effects of 20-HETE, 20-hydroxyeicosa-6(Z),15(Z)-dienoic acid (30 mg/kg, s.c.), prevented the beneficial effects of 5,14-HEDGE on MAP and vascular tone in rats treated with endotoxin. These data are consistent with the view that a fall in the production of 20-HETE contributes to the fall in MAP and vascular reactivity in rats treated with endotoxin, and that 5,14-HEDGE has a beneficial effect.

All‐Trans Retinoic Acid and Intra‐Amniotic Endotoxin‐Mediated Effects on Fetal Sheep Lung

All-trans retinoic acid (RA) is a potent modulator of lung development. Chorioamnionitis, which is frequently associated with preterm birth, causes fetal lung inflammation and improves lung function but also results in alveolar simplification and microvascular injury. Endotoxin-mediated chorioamnionitis reduces RA concentration in the fetal lung to 16% of control values. We hypothesized that administration of RA to the fetus before induction of chorioamnionitis would preserve septation of the distal airspaces. Time-mated ewes with singletons were assigned to receive a fetal intramuscular treatment with 20,000 IU of RA in olive oil (or olive oil only) 3 hr prior to intra-amniotic injection of endotoxin (20 mg, E. coli 055:B5) or saline, at 124-day gestational age and 7 days after the fetal treatment. The right cranial lung lobe was processed for morphometric analysis. RA treatment did not affect chorioamnionitis-induced fetal and systemic inflammation or interleukin-8 concentrations in lung tissue. RA administration alone did not alter lung structure. Relative to control lungs (5 +/- 3 mL/kg), lung volume increased similarly with endotoxin (22 +/- 4 mL/kg) or RA plus endotoxin (20 +/- 3 mL/kg; P < 0.05). Alveolar wall thickness was 4.2 +/- 0.3 mum after endotoxin-induced chorioamnionitis, 6.0 +/- 0.4 mum in controls (P < 0.05 versus endotoxin) and 5.5 +/- 0.2 mum after RA and endotoxin (P < 0.05 versus control, n.s. versus endotoxin). The ratio of airspace versus tissue was 4.6 +/- 0.3 in endotoxin-induced chorioamnionitis, 2.1 +/- 0.3 in controls and 4.1 +/- 0.5 after RA and endotoxin. We conclude that fetal treatment with RA did not prevent inflammation-induced alveolar simplification.

Dose- and time-related effects of dexmedetomidine on mortality and inflammatory responses to endotoxin-induced shock in rats

Our previous study demonstrated that dexmedetomidine drastically reduced mortality and inhibited the inflammatory response during endotoxemia in rats. The aim of this study was to clarify the dose- and time-related effects of dexmedetomidine on mortality and inflammatory responses to endotoxemia in rats. Male Wistar rats (n = 96) were anesthetized intraperitoneally with pentobarbital sodium and assigned to one of two protocols: one representing the dose-related effects of dexmedetomidine, and the other, the time-related effects of dexmedetomidine. To evaluate the dose-related effects, the animals were randomly assigned to one of four groups (n = 15 each): endotoxemic group (group E), receiving intravenous Escherichia coli endotoxin (15 mg x kg(-1) over 2 min); small-dose group (group S), treated with a small dose of dexmedetomidine (2.5 microg x kg(-1) x h(-1), IV); medium-dose group (group M), treated with a medium dose (5 microg x kg(-1) x h(-1), IV); and large-dose group (group L), treated with a large dose (10 microg x kg(-1) x h(-1), IV). To evaluate the time-related effects, the animals were randomly assigned to one of three groups (n = 12 per group): endotoxemic group; early posttreatment group, treated with 10 microg x kg(-1) x h(-1) dexmedetomidine at 1 h after endotoxin injection; and late posttreatment group, treated with 10 microg x kg(-1) x h(-1) at 2 h after endotoxin injection. Hemodynamics and arterial blood gases were recorded and plasma cytokine concentrations were measured throughout the observation period. The mortality rate was assessed up to 8 h after endotoxin injection. In the dose-related study, the mortality rates at 8 h after endotoxin injection were 81%, 26%, 32%, and 20% for groups E, S, M, and L, respectively. Plasma tumor necrosis factor-alpha (TNF) concentrations were lower in groups M and L than in group E at 2 h after endotoxin injection. Plasma interleukin-6 (IL-6) concentrations were lower in groups M and L than in group E at 4 and 5 h after endotoxin injection. In the time-related study, the mortality rates at 8 h after the endotoxin injection were 83%, 33%, and 58% for the endotoxemic, early posttreatment, and late posttreatment groups, respectively. The TNF concentration was lower in the early posttreatment group than in the endotoxemic group at 2 h after endotoxin injection, and the IL-6 concentration was lower in the early posttreatment group than in the endotoxemic group at 5 h after endotoxin injection. Dexmedetomidine dose-dependently attenuated extremely high mortality rates and increases in plasma cytokine concentrations after endotoxin injection. Moreover, the early administration of dexmedetomidine drastically reduced the high mortality rate and inhibited cytokine responses in endotoxin-exposed rats. These findings suggest that dexmedetomidine administration may be effective during sepsis.

Primary and secondary capture of platelets onto inflamed femoral artery endothelium is dependent on P-selectin and PSGL-1

Platelets constitute a key role in vascular injuries, however, the detailed mechanisms behind platelet–endothelial cell and platelet–leukocyte interactions in the femoral artery are not yet fully elucidated. We used intravital fluorescence microscopy of the femoral artery in C57BL/6 mice to study primary and secondary capture of platelets onto endothelial cells as well as onto adherent platelets and leukocytes in vivo. By use of monoclonal antibodies, the role of P-selectin and P-selectin glycoprotein ligand 1 (PSGL-1) in these adhesive interactions in mice exposed to endotoxin was determined. Intravenous injection of endotoxin significantly increased gene expression of P-selectin as well as platelet tethering, rolling and adhesion in the femoral artery. Pretreatment with the anti-PSGL-1 antibody decreased platelet tethering by 85%, platelet rolling by 88% and platelet adhesion by 96%. Immunoneutralization of P-selectin reduced platelet tethering by 91%, platelet rolling by 98%, and platelet adhesion by 97%. In addition, inhibition of P-selectin and PSGL-1 completely abolished secondary capture of platelets onto adherent platelets and leukocytes. Our data show that P-selectin and PSGL-1 mediate early interactions between platelets and other cells, including endothelial cells and leukocytes, in inflamed arteries. These novel results suggest that interference with P-selectin and PSGL-1 may be a useful target in strategies aiming to protect the vascular wall during arterial inflammation.