Lipopolysaccharide Challenge In Rats Research Articles

MTOR inhibition as a possible pharmacological target in the management of systemic inflammatory response and associated neuroinflammation by lipopolysaccharide challenge in rats.

Neuroinflammation plays a critical role during sepsis triggered by microglial activation. Mammalian target of rapamycin (mTOR) has gained attraction in neuroinflammation, however, the mechanism remains unclear. Our goal was to assess the effects of mTOR inhibition by rapamycin on inflammation, microglial activation, oxidative stress, and apoptosis associated with the changes in the inhibitor-κB (IκB)-α/nuclear factor-κB (NF-κB)/hypoxia-inducible factor-1α (HIF-1α) pathway activity following a systemic challenge with lipopolysaccharide (LPS). Rats received saline (10 mL/kg), LPS (10mg/kg), and (or) rapamycin (1mg/kg) intraperitoneally. Inhibition of mTOR by rapamycin blocked phosphorylated form of ribosomal protein S6, NF-κB p65 activity by increasing degradation of IκB-α in parallel with HIF-1α expression increased by LPS in the kidney, heart, lung, and brain tissues. Rapamycin attenuated the increment in the expression of tumor necrosis factor-α and interleukin-1β, the inducible nitric oxide synthase, gp91phox, and p47phox in addition to nitrite levels elicited by LPS in tissues or sera. Concomitantly, rapamycin treatment reduced microglial activation, brain expression of caspase-3, and Bcl-2-associated X protein while it increased expression of B cell lymphoma 2 induced by LPS. Overall, this study supports the hypothesis that mTOR contributes to the detrimental effect of LPS-induced systemic inflammatory response associated with neuroinflammation via IκB-α/NF-κB/HIF-1α signaling pathway.

Time and frequency dependent changes in resting state EEG functional connectivity following lipopolysaccharide challenge in rats.

Research has shown that inflammatory processes affect brain function and behavior through several neuroimmune pathways. However, high order brain functions affected by inflammation largely remain to be defined. Resting state functional connectivity of synchronized oscillatory activity is a valid approach to understand network processing and high order brain function under different experimental conditions. In the present study multi-electrode EEG recording in awake, freely moving rats was used to study resting state connectivity after administration of lipopolysaccharides (LPS). Male Wistar rats were implanted with 10 cortical surface electrodes and administered with LPS (2 mg/kg) and monitored for symptoms of sickness at 3, 6 and 24 h. Resting state connectivity and power were computed at baseline, 6 and 24 h. Three prominent connectivity bands were identified using a method resistant to spurious correlation: alpha (5–15 Hz), beta-gamma (20–80 Hz), and high frequency oscillation (150–200 Hz). The most prominent connectivity band, alpha, was strongly reduced 6 h after LPS administration, and returned to baseline at 24 h. Beta-gamma connectivity was also reduced at 6 h and remained reduced at 24 h. Interestingly, high frequency oscillation connectivity remained unchanged at 6 h and was impaired 24 h after LPS challenge. Expected elevations in delta and theta power were observed at 6 h after LPS administration, when behavioral symptoms of sickness were maximal. Notably, gamma and high frequency power were reduced 6 h after LPS and returned to baseline by 24 h, when the effects on connectivity were more evident. Finally, increases in cross-frequency coupling elicited by LPS were detected at 6 h for theta-gamma and at 24 h for theta-high frequency oscillations. These studies show that LPS challenge profoundly affects EEG connectivity across all identified bands in a time-dependent manner indicating that inflammatory processes disrupt both bottom-up and top-down communication across the cortex during the peak and resolution of inflammation.

Guanylyl Cyclase A in Both Renal Proximal Tubular and Vascular Endothelial Cells Protects the Kidney against Acute Injury in Rodent Experimental Endotoxemia Models.

WHAT THIS ARTICLE TELLS US THAT IS NEW: BACKGROUND:: Natriuretic peptides are used, based on empirical observations, in intensive care units as antioliguric treatments. We hypothesized that natriuretic peptides prevent lipopolysaccharide-induced oliguria by activating guanylyl cyclase A, a receptor for natriuretic peptides, in proximal tubules and endothelial cells. Normal Sprague-Dawley rats and mice lacking guanylyl cyclase A in either endothelial cells or proximal tubular cells were challenged with lipopolysaccharide and assessed for oliguria and intratubular flow rate by intravital imaging with multiphoton microscopy. Recombinant atrial natriuretic peptide efficiently improved urine volume without changing blood pressure after lipopolysaccharide challenge in rats (urine volume at 4 h, lipopolysaccharide: 0.6 ± 0.3 ml · kg · h; lipopolysaccharide + fluid resuscitation: 4.6 ± 2.0 ml · kg · h; lipopolysaccharide + fluid resuscitation + atrial natriuretic peptide: 9.0 ± 4.8 ml · kg · h; mean ± SD; n = 5 per group). Lipopolysaccharide decreased glomerular filtration rate and slowed intraproximal tubular flow rate, as measured by in vivo imaging. Fluid resuscitation restored glomerular filtration rate but not tubular flow rate. Adding atrial natriuretic peptide to fluid resuscitation improved both glomerular filtration rate and tubular flow rate. Mice lacking guanylyl cyclase A in either proximal tubules or endothelium demonstrated less improvement of tubular flow rate when treated with atrial natriuretic peptide, compared with control mice. Deletion of endothelial, but not proximal tubular, guanylyl cyclase A augmented the reduction of glomerular filtration rate by lipopolysaccharide. Both endogenous and exogenous natriuretic peptides prevent lipopolysaccharide-induced oliguria by activating guanylyl cyclase A in proximal tubules and endothelial cells.

Endotoxin Mediated-iNOS Induction Causes Insulin Resistance via ONOO− Induced Tyrosine Nitration of IRS-1 in Skeletal Muscle

BackgroundIt is believed that the endotoxin lipopolysaccharide (LPS) is implicated in the metabolic perturbations associated with both sepsis and obesity (metabolic endotoxemia). Here we examined the role of inducible nitric oxide synthase (iNOS) in skeletal muscle insulin resistance using LPS challenge in rats and mice as in vivo models of endotoxemia.Methodology/Principal FindingsPharmacological (aminoguanidine) and genetic strategies (iNOS−/− mice) were used to counter iNOS induction in vivo. In vitro studies using peroxynitrite (ONOO−) or inhibitors of the iNOS pathway, 1400 W and EGCG were conducted in L6 myocytes to determine the mechanism by which iNOS mediates LPS-dependent insulin resistance. In vivo, both pharmacological and genetic invalidation of iNOS prevented LPS-induced muscle insulin resistance. Inhibition of iNOS also prevented insulin resistance in myocytes exposed to cytokine/LPS while exposure of myocytes to ONOO− fully reproduced the inhibitory effect of cytokine/LPS on both insulin-stimulated glucose uptake and PI3K activity. Importantly, LPS treatment in vivo and iNOS induction and ONOO− treatment in vitro promoted tyrosine nitration of IRS-1 and reduced insulin-dependent tyrosine phosphorylation.Conclusions/SignificanceOur work demonstrates that iNOS-mediated tyrosine nitration of IRS-1 is a key mechanism of skeletal muscle insulin resistance in endotoxemia, and presents nitrosative modification of insulin signaling proteins as a novel therapeutic target for combating muscle insulin resistance in inflammatory settings.

Lipoprotein lipase is nitrated in vivo after lipopolysaccharide challenge

Lipopolysaccharide (LPS) administration down-regulates lipoprotein lipase (LPL) activity at the posttranscriptional level. Hypertriglyceridemia is the main metabolic consequence of this fall in LPL activity and is presumably involved in the innate immune response to infection. Nitric oxide (NO) has been implicated in LPS-induced down-regulation of LPL activity, but whether its effects are direct or indirect remains unclear. Here we examined the potential nitration of LPL in vivo in response to LPS challenge in rats. We found hypertriglyceridemia, iNOS expression, NO overproduction, and a generalized decrease in LPL activity in tissues 6 h after LPS administration. LPL sensitivity to nitration was first explored by in vitro exposure of bovine LPL to peroxynitrite, a reactive nitrogen species (RNS). Nitration was confirmed by anti-nitrotyrosine Western blot and subsequent identification of specific nitrotyrosine-containing LPL sequences by tandem mass spectrometry. Further analysis by targeted mass spectrometry revealed three in vivo-nitrated tyrosine residues in heart LPL from LPS-challenged rats. This is the first study to identify nitrated tyrosine residues in LPL, both in vitro and in vivo, and it demonstrates that LPL is a target for RNS in endotoxemia. These results indicate that LPL nitration may be a new mechanism of LPL activity regulation in vivo.

Differential behavioral, physiological, and hormonal sensitivity to LPS challenge in rats

Differential behavioral, physiological, and hormonal sensitivity to LPS challenge in rats Silvia Bison1, Lucia Carboni1, Roberto Arban1, Simon Bate2, Philip A Gerrard1, Maria Razzoli11Department of Biology, Neurosciences Centre of Excellence for Drug Discovery, GlaxoSmithKline, Medicines Research Centre, Verona, Italy; 2Statistical Sciences, GlaxoSmithKline, New Frontiers Science Park, Harlow, Essex CM195AW, UKAbstract: In animals, systemic administration of lipopolysaccharide (LPS) induces a characteristic set of responses that has been termed sickness behavior. In the present study, the effects of a wide dose range of LPS injections on behavior and physiology were examined in rats. Male rats were injected IP with either saline or LPS (1, 5, 15, 50, 125, or 250 μg/kg). Body temperature, body weight, and home-cage activity were monitored over 24 h after injection. Behaviorally the social interaction and the saccharin preference tests were assessed at 2 and 24 h post-injection, respectively. Further animals were treated with LPS to assess hypothalamic–pituitary adrenal (HPA) axis and immune system responses 2 h post-injection. LPS significantly reduced body weight (1 μg/kg and above), increased ACTH (15 μg/kg and above) and serum corticosterone levels (5 μg/kg and above). In addition LPS raised serum interleukins (5 μg/kg and above) and tumor necrosis factor-alpha (1 μg/kg and above). Social behavior and preference for saccharin were significantly decreased at all doses of LPS tested (1–250 μg/kg) while a robust reduction in home-cage activity was observed starting at 15 μg/kg. The present finding suggests that LPS caused profound behavioral alterations and stimulation of the immune system even at very low doses.Keywords: sickness behavior, lipopolysaccharide, cytokines, ACTH, corticosterone, social interaction

Kinetics of tumour necrosis factor α and interleukin 10 in rats with endotoxaemia and obstructive jaundice-induced anergy

Abstract Background Obstructive jaundice leads to a significant increase in septic complications. In addition, endotoxaemia may cause a secondary insult. The effects of endotoxaemia and obstructive jaundice-induced anergy on the kinetics of two early mediators of the response to sepsis, tumour necrosis factor (TNF) α and interleukin (IL) 10, were investigated. Methods Wistar rats had either bile duct ligation (BDL) or sham operation. After 2 weeks, anergy was confirmed by a delayed-type hypersensitivity response in rats that had undergone BDL. All rats were challenged with Escherichia coli lipopolysaccharide (LPS) 20 mg kg−1 or saline. After 90 and 180 min, rats were killed and blood samples were obtained for TNF-α and IL-10 measurement. Results There was no intragroup difference after saline challenge. LPS injection significantly increased both TNF-α and IL-10 levels in all groups compared with all saline injection groups. LPS injection following BDL led to significantly higher TNF-α levels and lower IL-10 levels at 90 min compared with those after sham operation (P < 0·05). There were no differences between TNF-α and IL-10 levels at 180 min following LPS challenge in rats that had undergone BDL. Conclusion BDL primes for an increase in TNF-α levels to a subsequent LPS challenge. However, the IL-10 increase following an LPS challenge is suppressed by BDL. These results suggest that BDL primes for a more vigorous inflammatory response to LPS challenge.

Colonic production and expression of IL-4, IL-6, and IL-10 in neonatal suckling rats after LPS challenge.

It has been demonstrated that the neonatal suckling rat is more susceptible to endotoxin [lipopolysaccharide (LPS)]-induced colonic damage compared with weaned littermates. There is evidence to suggest that differences in the production of certain cytokines, including interleukin (IL)-4, IL-6, and IL-10, are associated with intestinal inflammation in children. We have examined the production, localization, and mRNA detection of these cytokines in suckling and weaned rat colons after bacterial LPS challenge. Suckling (10 day old) and weaned (25 day old) rats were injected with LPS (3 mg/kg ip). Colon samples were taken up to 4 h after treatment, and cytokines were measured by ELISA. LPS-induced cytokine levels were significantly different in suckling rats compared with weaned rats. Cytokine localization to the colonic mucosa was evident in suckling rats up to 4 h after LPS administration but was not consistently seen in weaned rats. The mRNA for cytokines examined were detected by RT-PCR in suckling but not in weaned rat colons after LPS treatment. Induction of neutropenia via anti-neutrophil serum (ANS) administration did not affect cytokine mRNA detection in neonates after LPS treatment. Weaned animals displayed positive detection of all cytokines examined after ANS. Therefore, we have shown that the suckling rat displays a different production and expression of colonic IL-4, IL-6, and IL-10 compared with weaned littermates after LPS challenge. Furthermore, neutrophils may be implicated in colonic cytokine expression after LPS challenge in rats.

Acid instillation enhances the inflammatory response to subsequent lipopolysaccharide challenge in rats.

Aspiration of gastric contents is one of leading causes of the acute respiratory distress syndrome (ARDS). The pathogenesis of acid aspiration-induced acute lung injury is well understood. Less clear is why patients who have suffered acid aspiration are susceptible to ARDS. We studied the effects of acid instillation on the inflammatory response to subsequent lipopolysaccharide (LPS) challenge in rats. Instillation of acid into the right lung worsened the pathology induced by LPS that was administered 24 h after acid instillation. This included worsened oxygenation, increased pulmonary edema, increased production of tumor necrosis factor-alpha (TNF-alpha) and cytokine-induced neutrophil chemoattractant, neutrophil accumulation and mobilization to the alveolar spaces, and nitric oxide (NO) production. Of interest, neutrophil mobilization, NO production, and protein permeability were also magnified in the left lung. These effects were attenuated by administration of the protein tyrosine kinase (PTK) inhibitors genistein and tyrphostin AG556. These data suggest that acid instillation primes the rat to enhance the inflammatory response to subsequent endotoxin challenge and that at least part of the augmented inflammatory response depends on PTK.

Differential effect of a single high dose of the tricyclic antidepressant imipramine on interleukin-1 β and tumor necrosis factor- α secretion following an in vivo lipopolysaccharide challenge in rats

Tricyclic antidepressants (TCAs) of which imipramine is one, are commonly used in the treatment of depressive disorders and other forms of psychiatric illness. There have been many reports regarding the suppressive effects of TCAs on immune function. However, information is still limited regarding the effects of TCAs on the immune system, as many of the studies conducted to date have concentrated on in vitro exposure to such drugs, or ex vivo measures of immunity following drug administration. Thus in the present investigation, an in vivo challenge with bacterial lipopolysaccharide (LPS) (100 μg/kg; i.p.) was used to assess immunocompetence following administration of a single high dose of the TCA, imipramine (100 mg/kg, p.o.). The results demonstrated that imipramine pretreatment inhibits LPS-induced increases in serum concentrations of the proinflammatory cytokine, tumor necrosis factor (TNF)- α both 3 and 6 h, following administration. However, LPS-induced interleukin (IL)-1 β secretion was not significantly altered following imipramine treatment at either of the timepoints examined. In addition, serum concentrations of corticosterone and the antiinflammatory cytokine IL-10 were measured, and imipramine treatment failed to alter either basal, or LPS-induced increases in these immunosuppressive agents. In conclusion, although IL-1 β and TNF- α are both macrophage-derived proinflammatory cytokines, the present study demonstrates a differential sensitivity of these cytokines to the suppressive effects of the TCA imipramine. Furthermore, the suppressive effects of imipramine on LPS-induced TNF- α secretion could not be attributed to either increased glucocorticoid levels, or increased secretion of the antiinflammatory cytokine IL-10. The relevance of these findings to antidepressant-induced immunotoxicity are discussed.