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Abstract Endotoxin administration is commonly used to study the inflammatory response, and though traditionally given as a bolus injection, it can be administered as a continuous infusion over multiple hours. Several studies hypothesize that the latter better represents the prolonged and pronounced inflammation observed in conditions like sepsis. Yet very few experimental studies have administered endotoxin using both strategies, leaving significant gaps in determining the underlying mechanisms responsible for their differing immune responses. We used mathematical modelling to analyse cytokine data from two studies administering a 2 ng kg−1 dose of endotoxin, one as a bolus and the other as a continuous infusion over 4 h. Using our model, we simulated the dynamics of mean and subject‐specific cytokine responses as well as the response to long‐term endotoxin administration. Cytokine measurements revealed that the bolus injection led to significantly higher peaks for interleukin (IL)‐8, while IL‐10 reaches higher peaks during continuous administration. Moreover, the peak timing of all measured cytokines occurred later with continuous infusion. We identified three model parameters that significantly differed between the two administration methods. Monocyte activation of IL‐10 was greater during the continuous infusion, while tumour necrosis factor α and IL‐8 recovery rates were faster for the bolus injection. This suggests that a continuous infusion elicits a stronger, longer‐lasting systemic reaction through increased stimulation of monocyte anti‐inflammatory mediator production and decreased recovery of pro‐inflammatory catalysts. Furthermore, the continuous infusion model exhibited prolonged inflammation with recurrent peaks resolving within 2 days during long‐term (20–32 h) endotoxin administration.

Endotoxin administration is commonly used to study the inflammatory response, and though traditionally given as a bolus injection, it can be administered as a continuous infusion over multiple hours. Several studies hypothesize that the latter better represents the prolonged and pronounced inflammation observed in conditions like sepsis. Yet very few experimental studies have administered endotoxin using both strategies, leaving significant gaps in determining the underlying mechanisms responsible for their differing immune responses. We used mathematical modelling to analyse cytokine data from two studies administering a 2 ng kg−1 dose of endotoxin, one as a bolus and the other as a continuous infusion over 4 h. Using our model, we simulated the dynamics of mean and subject‐specific cytokine responses as well as the response to long‐term endotoxin administration. Cytokine measurements revealed that the bolus injection led to significantly higher peaks for interleukin (IL)‐8, while IL‐10 reaches higher peaks during continuous administration. Moreover, the peak timing of all measured cytokines occurred later with continuous infusion. We identified three model parameters that significantly differed between the two administration methods. Monocyte activation of IL‐10 was greater during the continuous infusion, while tumour necrosis factor α$ {\alpha} $ and IL‐8 recovery rates were faster for the bolus injection. This suggests that a continuous infusion elicits a stronger, longer‐lasting systemic reaction through increased stimulation of monocyte anti‐inflammatory mediator production and decreased recovery of pro‐inflammatory catalysts. Furthermore, the continuous infusion model exhibited prolonged inflammation with recurrent peaks resolving within 2 days during long‐term (20–32 h) endotoxin administration. What is the central question of this study?How do dynamic immune responses to continuous and bolus endotoxin administration compare using mathematical models, and what physiological mechanisms do we hypothesize attribute to these observed differences? How do extended and enhanced continuous infusions impact immune responses, and do they express similar characteristics to infection dynamics?What is the main finding and its importance?This study proposes a mathematical model for a continuous infusion of endotoxin. We hypothesize that the dynamic switch from a bolus to a continuous infusion of endotoxin, which results in higher peak concentrations of IL‐10, lower peak concentrations of IL‐8, and later peak timing of all measured cytokines, is attributed to increased monocyte activation rate of anti‐inflammatory cytokine IL‐10 and decreased decay rates of pro‐inflammatory cytokines TNF‐a and IL‐8. Additionally, prolonged LPS administration significantly extends the cytokine response and results in recurrent cytokine spikes. Our results provide a better understanding of the mechanisms impacting the change in immune responses with long‐term endotoxin infusion. [ABSTRACT FROM AUTHOR]

Abstract Background Surfactant phospholipid (PL) composition plays an important role in lung diseases. We compared the PL composition of non-invasively collected exhaled breath particles (PEx) with bronchoalveolar lavage (BAL) and induced sputum (ISP) at baseline and following endotoxin (LPS) challenges. Methods PEx and BAL were collected from ten healthy nonsmoking participants before and after segmental LPS challenge. Four weeks later, PEx and ISP were sampled in the week before and after a whole lung LPS inhalation challenge. PL composition was analysed using mass spectrometry. Results The overall PL composition of BAL, ISP and PEx was similar, with PC(32:0) and PC(34:1) representing the largest fractions in all three sample types (baseline PC(32:0) geometric mean mol%: 52.1, 56.9, and 51.7, PC(34:1) mol%: 11.7, 11.9 and 11.4, respectively). Despite this similarity, PEx PL composition was more closely related to BAL than to ISP. For most lipids comparable inter-individual differences in BAL, ISP, and PEx were found. PL composition of PEx was repeatable. The most pronounced increase following segmental LPS challenge was detected for SM(d34:1) in BAL (0.24 to 0.52 mol%) and following inhalation LPS challenge in ISP (0.45 to 0.68 mol%). An increase of SM(d34:1) following segmental LPS challenge was also detectable in PEx (0.099 to 0.103 mol%). The inhalation challenge did not change PL composition of PEx. Conclusion Our data supports the peripheral origin of PEx. The lack of PL changes in PEx after inhalation challenge might to be due to the overall weaker response of inhaled LPS which primarily affects the larger airways. Compared with BAL, which always contains lining fluid from both peripheral lung and central airways, PEx analysis might add value as a selective and non-invasive method to investigate peripheral airway PL composition. Trial registration NCT03044327, first posted 07/02/2017.

Background: Surfactant phospholipid (PL) composition plays an important role in lung diseases. We compared the PL composition of non-invasively collected exhaled breath particles (PEx) with bronchoalveolar lavage (BAL) and induced sputum (ISP) at baseline and following endotoxin (LPS) challenges. Methods: PEx and BAL were collected from ten healthy nonsmoking participants before and after segmental LPS challenge. Four weeks later, PEx and ISP were sampled in the week before and after a whole lung LPS inhalation challenge. PL composition was analysed using mass spectrometry. Results: The overall PL composition of BAL, ISP and PEx was similar, with PC(32:0) and PC(34:1) representing the largest fractions in all three sample types (baseline PC(32:0) geometric mean mol%: 52.1, 56.9, and 51.7, PC(34:1) mol%: 11.7, 11.9 and 11.4, respectively). Despite this similarity, PEx PL composition was more closely related to BAL than to ISP. For most lipids comparable inter-individual differences in BAL, ISP, and PEx were found. PL composition of PEx was repeatable. The most pronounced increase following segmental LPS challenge was detected for SM(d34:1) in BAL (0.24 to 0.52 mol%) and following inhalation LPS challenge in ISP (0.45 to 0.68 mol%). An increase of SM(d34:1) following segmental LPS challenge was also detectable in PEx (0.099 to 0.103 mol%). The inhalation challenge did not change PL composition of PEx. Conclusion: Our data supports the peripheral origin of PEx. The lack of PL changes in PEx after inhalation challenge might to be due to the overall weaker response of inhaled LPS which primarily affects the larger airways. Compared with BAL, which always contains lining fluid from both peripheral lung and central airways, PEx analysis might add value as a selective and non-invasive method to investigate peripheral airway PL composition. Trial registration: NCT03044327, first posted 07/02/2017. [ABSTRACT FROM AUTHOR]

The objective was to evaluate the influence of dams grazing endophyte-infected tall, toxic, fescue (E+) during gestation on innate immune response of male offspring to an endotoxin challenge. Crossbred pregnant multiparous cows (n = 36) bred to Red Angus sires were housed in replicated E+ or novel endophyte-infected, non-toxic, fescue (NE+) pastures before breeding and through the length of gestation. After parturition, cows and calves were moved to a common bermudagrass pasture until calves were weaned. From this calf-crop, a subset of post-weaned steers (E+ n = 8; NE+ n = 8) were selected for a lipopolysaccharide (LPS) challenge. Steers (302 ± 36 d of age; 261 ± 62 kg body weight) were fitted with jugular vein catheters >7 h before sampling. Blood was collected every 30 min starting 2 h before (Pre-LPS) and 9 h following (Post-LPS) LPS administration (0.5 µg/kg body weight). Sickness behavior score (SBS), cortisol, and glucose were analyzed every 30 min. Complete blood count (CBC) and rectal temperature (RT) were analyzed hourly. Peak concentration, time at peak concentration, baseline, return to baseline, and area under the curve (AUC) were evaluated for cortisol and glucose. Cortisol and glucose were determined by radioimmunoassay and hexokinase colorimetric assay, respectively. The CBC data were analyzed using a Hemavet hemocytology analyzer. Pre-LPS and Post-LPS cortisol, glucose, SBS, RT, and CBC were analyzed using the MIXED procedure of SAS specific for repeated measures with treatment, time, and the interaction as fixed effects and dam sire breed as a random effect. Peak concentration, time at peak concentration, baseline, return to baseline, and AUC for cortisol and glucose were analyzed using the MIXED procedure of SAS with treatment as a fixed effect and dam sire breed as a random effect. There was a Pre-LPS treatment by time interaction for RT (P < 0.05). Pre-LPS monocyte count was lower in offspring of dams grazing E+ relative to NE+ during gestation (P < 0.05). Pre-LPS hematocrit was increased in offspring of dams grazing E+ relative to NE+ during gestation (P < 0.05). There was a Pre-LPS effect of time on SBS (P < 0.05). As expected, Post-LPS cortisol, glucose, RT, WBC, neutrophil count, lymphocyte count, lymphocyte percentage, eosinophils count, eosinophils percentage, hematocrit percentage, RBC, and platelets changed over time in response to LPS (P < 0.05). Post-LPS lymphocyte count and monocyte percentage were decreased in offspring of dams grazing E+ relative to NE+ during gestation (P < 0.05). Post-LPS eosinophils count, eosinophil percentage, and hematocrit were increased in offspring of dams grazing E+ relative to NE+ during gestation (P < 0.05). There was a Post-LPS treatment by time interaction for monocyte count and neutrophil percentage (P < 0.05). Preliminary results suggest an altered innate immune response of male offspring of dams grazing toxic relative to novel fescue during gestation. [ABSTRACT FROM AUTHOR]

The objectives of this study were to (1) evaluate the use of a pressure algometer and an automated rumination monitoring system to assess changes in pain sensitivity and rumination time in response to endotoxin-induced clinical mastitis and (2) evaluate the effect of the nonsteroidal antiinflammatory drug meloxicam on pain sensitivity and rumination time, as well as other clinical signs, in dairy cattle with endotoxin-induced clinical mastitis. Clinical mastitis was induced in 12 primiparous and 12 multiparous lactating dairy cows by intramammary infusion of 25 µg of Escherichia coli lipopolysaccharide (LPS) into 1 uninfected quarter. Immediately after, half the cows were injected subcutaneously with meloxicam (treated group) and half with the same volume of a placebo solution (control group). Pain sensitivity was assessed by measuring the difference in pressure required to elicit a response on the control and challenged quarter using an algometer 3 d before, immediately before, and at 3, 6, 12, and 24 h after LPS infusion and either meloxicam or placebo injection. Rumination was continuously monitored from 2 d before to 3 d after LPS infusion using rumination loggers. Udder edema, body temperature, somatic cell score, and dry matter intake were also monitored to evaluate the occurrence and the duration of the inflammation after LPS infusion. In control animals, the difference in the pressure applied to the control and challenged quarters (control -- challenged quarter) increased by 1.1 ± 0.4 kg of force 6 h after LPS infusion compared with the baseline, suggesting an increase in pain sensitivity in the challenged quarter. Neither the LPS infusion nor the meloxicam treatment had an effect on daily rumination time. However, the rumination diurnal pattern on the day of LPS infusion showed an overall deviation from the baseline pattern. Cows spent less time ruminating in the hours following LPS infusion and more time ruminating later in the day. Meloxicam did not alter somatic cell score or dry matter intake. However, meloxicam-treated animals had less udder edema and a lower body temperature in the hours following LPS infusion compared with control animals. In conclusion, pressure algometers and rumination loggers show promise as tools to detect mastitis and monitor recovery on farm. Further, meloxicam has a beneficial effect in relieving pain and decreasing udder edema and body temperature in LPS-induced clinical mastitis. [ABSTRACT FROM AUTHOR]

Metabolic demand for sulfur-containing AA increases during inflammation in nonruminants. Therefore, Met supplementation may alleviate the negative effects of infection on N balance. Effects of gram-negative bacterial lipopolysaccharide (LPS) and supplemental dietary Met on N balance, serum hormones and haptoglobin, and plasma urea-N and AA were evaluated in 20 Angus-cross steers (BW - 262 ± 6.3 kg). Treatments (2 x 2 factorial) were infusion of no LPS (-LPS) or a prolonged low dose of LPS (+LPS) and dietary supplementation of no (-MET) or 14 g/d (+MET) of rumen-protected Met (providing 7.9 g/d of DL-Met). Steers were adapted to a roughage-based diet (DMI = 1.4% of BW daily) and supplemental Met for 14 d, and were then infused (1 mL/min via intravenous catheter) with LPS on d 1 (2 µg/kg of BW) and 3 (1 µg/kg of BW) of a 5-d collection period. Blood was collected on d 1, before LPS infusion, and at 2, 4, 6, 8, 10, 12, and 24 h after LPS challenge. Diet samples, feed refusals, feces, and urine were collected daily for 5 d. Rectal temperature and serum concentrations of cortisol, prolactin, tumor necrosis factor-a, and haptoglobin increased, whereas thyroxine and triiodothyronine decreased for +LPS vs. -LPS steers (LPS x h; P < 0.01). Plasma urea-N was greater for +LPS than -LPS steers (LPS; P = 0.0:3), and serum IGF-1 was not affected (P ≥ 0.26) by LPS or Met. Plasma concentrations of Thr, Lys, Leu, Tie, Phe, Trp, Asn, Glu, and Orn decreased, plasma Ala increased, and Gly and Ser initially increased, then declined in +LPS vs. -LPS steers (LPS x h; P ≤ 0.04). Plasma Met was greater for +MET than -MET steers before LPS infusion, but declined in +MET steers after LPS infusion (LPS x Met x h; P < 0.01). By design, DMI, was not different, but DM digested was less (P = 0.04) for +LPS than -LPS steers. Infusion of LPS did not affect (P ≥ 0.24) N intake, fecal N excretion, or N digested, but resulted in greater (P < 0.01) urinary N excretion and less (P < 0.01) N retention. The absence of an LPS x Met interaction (P = 0.26) for N retention indicates that supplemental Met does not improve the N utilization of growing beef steers exposed to a gram-negative bacterial endotoxin. Decreases in plasma concentrations of several essential AA in +LPS steers suggest that metabolic demand for these AA likely increased in steers exposed to endotoxin. [ABSTRACT FROM AUTHOR]

Five studies were conducted to evaluate the effects of lipid source on performance and health of stressed feeder cattle. A total of 332 heifers (195 ± 2.37 kg initial BW) in trial 1 and 336 heifers (206 ± 1.70 kg initial BW) in trial 2 were fed diets containing ground fiaxseed (FLAX), rolled full-fat soybeans (SOY), or tallow (TAL) at 13, 20, or 4%, respectively (DM basis). All diets were formulated to be isonitrogenous and isocaloric. The ADG and G:F for the first 7 d and for the entire feeding period were greater (P < 0.05) for TAL and FLAX than for SOY. Percentage of animals treated and retreated for bovine respiratory disease did not differ among dietary treatments. The FLAX treatment increased (P < 0.05) total n-3 PUFA concentrations in the plasma, whereas SOY increased (P < 0.05) plasma concentrations of total n-6 PUFA. In trial 3, 18 steers were individually fed diets containing TAL and 18 steers were fed a diet containing SOY (20% of DM). In trials 4 and 5, 18 steers were individually fed diets containing TAL and 18 steers were fed diets containing FLAX (12.9% of DM). On d 14 and 17 of study 3, 4, and 5, 16 steers from each dietary treatment were injected i.v. with Escherichia coli O55:B5 lipopolysaccharide (LPS), and 2 steers from each diet were injected with saline. Rectal temperatures after LPS challenge were lower (P < 0.05) for SOY and FLAX than for TAL, and plasma TNF was greater (P < 0.05) for SOY than for TAL. Serum haptoglobin and blood fibrinogen increased and white blood cell count decreased in response to LPS, but none of these variables was affected by treatment. Although this research failed to measure an effect of lipid source on feedlot morbidity or mortality, these studies indicate that altering the source and type of dietary fatty acids may modify the immune response in stressed feeder cattle and that performance may be hindered by feeding full-fat soybeans to receiving cattle. [ABSTRACT FROM AUTHOR]

Objective: To assess the effects of intravenous fish oil fat emulsion on the metabolic alterations induced by lipopolysaccharide (LPS) challenge in healthy volunteers.Design: Two groups of eight healthy subjects were randomized to receive either two pharmacological doses of intravenous FO fat emulsion or no treatment. The FO group received twice 0.5 g/kg 10% emulsion (Omegaven) 48 and 24h before investigation. LPS (2 ng/kg) was injected as a bolus on the investigation day. Systemic parameters, indirect calorimetry, heart rate variability, and platelet membrane phospholipid composition were measured.Results: Basal EPA and DHA content in platelet phospholipids was low (0.28% and 2.54%, respectively) and increased significantly after FO to 1.68% and 3.32%. LPS induced reproducible effects in all subjects. Fever was higher in the control [corrected] group than in FO group [corrected] the difference was significant from t (120) until t (360). FO blunted the neuroendocrine response: the rise in plasma norepinephrine was sevenfold lower at t (120) while the ACTH peak was fourfold lower. Tumor necrosis factor alpha was significantly lower between t (360) and t (180) in the FO group.Conclusions: Two doses of intravenous FO fat emulsion modified the phospholipid composition of platelets in healthy subjects. FO blunted fever and increased the neuroendocrine and the inflammatory responses to LPS. [ABSTRACT FROM AUTHOR]

Summary: Background & aims: Fish oil (FO) has been shown to modulate the acute and chronic inflammatory responses. Endotoxin (LPS) has been shown to mimic several aspects of sepsis. The study aimed at testing the effects of oral FO supplements in healthy subjects submitted to intravenous LPS on systemic and endocrine response. Subjects and methods: Fifteen healthy men (aged 26.0±3.1 years, BMI 23.8±1.9kg/m2), were enrolled. Subjects were randomised to 3–4 weeks of oral FO supplementation (7.2g/day, providing 1.1 g/day of 20:5(n-3) and 0.7 g/day of 22:6 (n-3) fatty acids) or no supplementation and then submitted to endotoxin challenge: 2ng/kg of LPS. All subjects were studied twice (placebo and LPS). Measurements: vital signs, energy expenditure (EE), glucose and lipid metabolism (2H2-glucose), plasma cytokines and stress hormones for 6h after LPS or placebo. Results: LPS caused cytokine release, fever, increases in heart rate, resting EE and substrate oxidation, plasma glucagon and glucose concentrations; the neuro-endocrine response was characterised by increased plasma stress hormones. FO significantly blunted fever, ACTH and cortisol plasma levels (no effect on cytokine release). FO blunted the peak norepinephrine after LPS. Conclusion: FO supplements blunted the endocrine stress response and the increase in body temperature, but had no impact on cytokine production after LPS. These findings conflict with the postulated anti-inflammatory effects of FO on arachidonic acid metabolism and cytokine release. These results suggest that FO may exert beneficial effects in sepsis though non-inflammatory which require further investigations [Copyright &y& Elsevier]

Abstract: Embryonic stem (ES) cells and ES cell-derived differentiated cells can be used in tissue regeneration approaches. However, inflammation may pose a major hurdle. To define the inflammatory response of ES and ES cell-derived vascular cells, we exposed these cells to LPS. With the exception of MIF no significant cytokine mRNA levels were observed either at baseline or after stimulation. Further experiments revealed that these cells do not express TLR4. Analysis of the DNA methylation status of the TLR4 upstream region showed increased methylation. Moreover, in vitro methylation suppressed TLR4 promoter activity in reporter gene assays. ChIP assays showed that in this region histones H3 and H4 are hypoacetylated in ES cells. Interestingly, 5-aza-dC or TSA partially relieves this gene repression. Finally, the increased levels of TLR4 observed in ES cells after treatment with 5-aza-dC or TSA confer responsiveness to LPS, as induction of IL-6 and TNFα mRNA was detected in endotoxin stimulated ES cells. [Copyright &y& Elsevier]

The inflammatory response in swine challenged with lipopolysaccharide (LPS) has only been partially characterized. As swine are increasingly used in biomedical research, it is important to determine if they respond to endotoxin challenge in a manner similar to other model systems. Accordingly, 24 Poland China×Landrace barrows were treated with saline, LPS, dexamethasone, or LPS and dexamethasone, with six animals in each treatment group.The kinetics of TNFα, IL-1β, IL-6, IL-8, IL-10, nitric oxide (nitrate/nitrite), and neopterin production in swine plasma were examined at 1, 3, 6, 9, and 24 h after acute LPS challenge. Lipopolysaccharide increased plasma TNFα levels, which peaked 1 h post-challenge. Dexamethasone decreased LPS-induced TNFα by approximately 60%. Plasma IL-6 levels peaked 3 h post-LPS challenge, returning to basal levels by 9 h. Swine given both LPS and dexamethasone had minimal IL-6 levels. Control and dexamethasone-only treated animals never exhibited systemic TNFα or IL-6 levels. Lipopolysaccharide increased plasma IL-10 1 h after challenge. Dexamethasone did not alter plasma IL-10 levels in LPS-challenged swine. Interleukin-1β was constitutively present in plasma and was not altered by any combination of treatments. Plasma IL-8 was not observed in any treatment group. Plasma nitrate/nitrite levels were maximal 24 h post-challenge. Dexamethasone treatment prevented increases in plasma nitrate/nitrite levels in LPS-treated animals. Lipopolysaccharide induced levels of neopterin; dexamethasone served to further increase plasma neopterin levels in LPS-challenged animals. The discordant regulation of inflammatory mediators suggests that the immunological responses by swine to LPS are distinct from the responses seen in rodent and human studies. [Copyright &y& Elsevier]