Lipopolysaccharide and cytokines modulate leukotriene (LT)B4 and LTC4 production by porcine endometrial endothelial cells

Uterine inflammatory response is mediated by inflammatory mediators including eicosanoids and cytokines produced by immune and endometrial cells. Interactions between lipopolysaccharide (LPS) and cytokines, and leukotrienes (LTs) in endothelium, important for the host defence during the inflammation, are unknown. We studied the effect of LPS, tumour necrosis factor (TNF)‐α, interleukin (IL)‐1β, IL‐4 and IL‐10 on 5‐lipooxygenase (5‐LO), LTA₄ hydrolase (LTAH) and LTC₄ synthase (LTCS) mRNA and protein expression, LTB₄ and LTC₄ release from porcine endometrial endothelial cells, and cell viability. For 24 hr, cells were exposed to LPS (10 or 100 ng/ml of medium) and cytokines (each 1 or 10 ng/ml). 5‐LO mRNA/protein expression augmented after incubation with larger doses of LPS, TNF‐α, IL‐4 and IL‐10 and smaller dose of IL‐1β. Larger dose of TNF‐α, smaller doses of LPS and IL‐1β and both doses of IL‐10 increased LTAH mRNA/protein expression. LTAH protein content was up‐regulated by larger dose of LPS, but it was reduced in response to both doses of IL‐4. LTCS mRNA expression was elevated by larger doses of LPS, IL‐4 and IL‐10 or both doses of TNF‐α and IL‐1β. LTCS protein level increased after treatment with both doses of IL‐1β, IL‐4 and IL‐10, smaller dose of LPS and larger dose of TNF‐α. Both doses of LPS and larger doses of TNF‐α and IL‐10 increased LTB₄ release. LPS, IL‐1β and IL‐10 at smaller doses, or TNF‐α and IL‐4 at larger doses stimulated LTC₄ release. Smaller doses of TNF‐α and IL‐1β or both doses of IL‐4 enhanced the cell viability. This work provides new insight on the participation of LPS, TNF‐α, IL‐1β, IL‐4 and IL‐10 in LTB₄ and LTC₄ production/release from porcine endometrial endothelial cells, and the effect of above factors on these cells viability. The used cellular model gives the possibility to further establish the interactions between inflammatory mediators.     

Regulation of uterine function by cytokines in cows: Possible actions of tumor necrosis factor-α, interleukin-1α and interferon-τ

When animals do not become pregnant, regression of the corpus luteum (CL) is essential for normal cyclicity because it allows the development of a new ovulatory follicle. Luteal regression is caused by a pulsatile release of prostaglandin (PG) F_2α from the uterus in the late luteal phase in most mammals including cattle. Although it has been proposed in ruminants that pulsatile PGF_2α secretion is generated by a positive feedback loop between luteal and/or hypophyseal oxytocin and uterine PGF_2α, the bovine endometrium may possess other mechanisms for initiation of luteolytic PGF_2α secretion. There is increasing evidence that several cytokines mainly produced by immune cells modulate CL and uterine function in many species. Tumor necrosis factor‐α (TNF‐α) stimulates PGF_2α output from bovine endometrium not only at the follicular phase but also at the late luteal phase. Administration of TNF‐α at a high concentration prolongs luteal lifespan, whereas administration of a low concentration of TNF‐α accelerates luteal regression in cows. The data obtained from the authors’ previous in vitro and in vivo studies strongly suggest that TNF‐α is a crucial factor in regulating luteolysis in cows. The authors’ recent study has shown that interleukin‐1α mediates PG secretion from bovine endometrium as a local regulator. Furthermore, interferon‐τ (IFN‐τ) suppresses the action of TNF‐α on PGF_2α synthesis by the bovine endometrium in vitro, suggesting that IFN‐τ plays a luteoprotective role by inhibiting TNF‐α‐induced PGF_2α production in early pregnancy. The purpose of the present review is to summarize current understanding of the endocrine mechanisms that regulate uterine function by cytokines during the estrous cycle and early pregnancy in cows.     

Effect of Intramammary Infusion of Tumour Necrosis Factor‐α on Milk Protein Composition and Induction of Acute‐Phase Protein in the Lactating Cow

The present study was designed to evaluate the effects of tumour necrosis factor‐α (TNF‐α) on lactating bovine mammary functions such as milk protein secretion and the integrity of the milk‐blood barrier. The effect on the induction of the systemic inflammatory response was also examined using concentrations of serum haptoglobin (Hp), a major inflammatory acute‐phase protein, as an index. One hundred micrograms per mammary gland of recombinant bovine (rBo) TNF‐α or placebo saline was individually infused into a rear mammary gland of each of four lactating cows, and milk and blood samples were collected before and 4, 8, 24, 32, 48, 96 and 168 h after infusion. In the rBoTNF‐α‐infused gland, increases of somatic cell counts were observed at 4–48 h. Although concentrations of total milk protein were not changed, compositions of milk proteins varied following rBoTNF‐α infusion. Concentrations of caseins, α‐lactalbumin and β‐lactoglobulin were significantly decreased at 4 and 8 h. Lactoferrin concentrations were significantly increased at 4 h. Significant infiltrations of serum albumin, immunoglobulin G1 (IgG1) and IgG2 were observed at 4 and 8 h. Elevations of the serum concentration of Hp were detected at 8‐32 h, but were very small in comparison with those reported in inflammatory diseases. Changes in rectal temperature and white blood cell counts were not significant. These results show that single rBoTNF‐α infusion into the lactating mammary gland suppresses the lactogenic function of the gland and influences the function of the milk‐blood barrier, with little effect on the generalized inflammatory response.     

Effects of tumour necrosis factor‐alpha and interleukin‐1 beta on in vitro development of bovine secondary follicles

The objective of this study was to determine the effects of TNF‐α and IL‐1β on development and survival of bovine secondary follicle culture in vitro for 18 days. Secondary follicles (~0.2 mm) were isolated from ovarian cortex and individually cultured at 38.5°C, with 5% CO₂ in air, for 18 days, in TCM‐199⁺ alone (cultured control) or supplemented with 10 ng/ml IL‐1β, 10 ng/ml TNF‐α or both TNF‐α and IL‐1β. The effects of these treatments on growth, follicular survival, antrum formation, viability, ultrastructure and mRNA levels for GDF‐9, c‐MOS, H1foo and Cyclin B1 were evaluated. The results showed that addition of TNF‐α to culture medium increased follicular diameter and rate of antrum formation, whereas that of IL‐1β and a mixture of IL‐1β and TNF‐α did not do so. Ultrastructural analysis showed that, among the tested cytokine treatments, follicles cultured in the presence of TNF‐α had the best‐preserved oocytes and granulosa cells. The presence of TNF‐α, IL‐1β or both did not influence the expression of mRNAs analysed. In conclusion, in contrast to IL‐1β, TNF‐α promotes growth of and antrum formation in in vitro cultured bovine secondary follicles, while their ultrastructure and viability were maintained.     

Eleutheroside B increase tight junction proteins and anti‐inflammatory cytokines expression in intestinal porcine jejunum epithelial cells (IPEC‐J2)

Eleutheroside B (EB) is a phenylpropanoid glycoside with anti‐inflammatory properties, neuroprotective abilities, immunomodulatory effects, antinociceptive effects, and regulation of blood glucose. The aim of this study was to investigate the effects of EB on the barrier function in the intestinal porcine epithelial cells J2 (IPEC‐J2). The IPEC‐J2 cells were inoculated into 96‐well plates at a density of 5 × 10³ cells per well for 100% confluence. The cells were cultured in the presence of EB at concentrations of 0, 0.05, 0.10, and 0.20 mg/ml for 48 hr. Then, 0.10 mg/ml was selected as the suitable concentration for the estimation of transepithelial electric resistance (TEER) value, alkaline phosphatase activity, proinflammatory cytokines mRNA expression, tight junction mRNA and protein expression. The results of this study indicated that the supplementation of EB in IPEC‐J2 cells decreased cellular membrane permeability and mRNA expression of proinflammatory cytokines, including interleukin‐6 (IL‐6), interferon‐γ (INF‐γ), and tumour necrosis factor‐α (TNF‐α). The supplementation of EB in IPEC‐J2 cells increased tight junction protein expression and anti‐inflammatory cytokines, interleukin 10 (IL‐10) and transforming growth factor beta (TGF‐β). In addition, the western blotting and real‐time quantitative polymerase chain reaction (RT‐qPCR) results indicated that EB significantly (p < 0.05) increased the mRNA and protein expression of intestinal tight junction proteins, Claudin‐3, Occludin, and Zonula Occludins protein‐1 (ZO‐1). Therefore, dietary supplementation of EB may increase intestinal barrier function, tight junction protein expression, anti‐inflammatory cytokines, and decrease proinflammatory cytokines synthesis in IPEC‐J2 cells.     

Immune dysregulation and osteosarcoma: Staphylococcus aureus downregulates TGF‐β and heightens the inflammatory signature in human and canine macrophages suppressed by osteosarcoma

Since William Coley utilized bacterial immunotherapy to treat sarcomas in the late 19th century, an association between infection and improved survival has been reported for human and canine osteosarcoma patients. One of the reasons for this improved survival is likely a reactivation of the host immune system towards an inflammatory anti‐tumour response, and one of the key players is the macrophage. Yet, despite their importance, the response of macrophages to infectious agents in the context of osteosarcoma has not been thoroughly evaluated. The aim of this study was to evaluate how in vitro exposure to a bacterial agent (Staphylococcus aureus) influenced canine and human macrophage differentiation in the presence of osteosarcoma. Our hypothesis was that S. aureus would, in the presence of osteosarcoma, induce a macrophage phenotype with significantly increased inflammatory signatures. Consistent with our hypothesis, human macrophages co‐cultured with osteosarcoma and S. aureus exhibited increased IFN‐γ, TNF‐α and IL‐12p70 cytokine secretion, decreased TGF‐β cytokine secretion and increased mRNA expression of TNF‐α when compared with macrophages co‐cultured with osteosarcoma and to macrophages cultured alone. Canine macrophages similarly exhibited increased IFN‐γ and TNF‐α cytokine secretion, decreased TGF‐β cytokine secretion, increased mRNA expression of TNF‐α and increased surface receptor expression of CD80 when co‐cultured with osteosarcoma and S. aureus. Collectively, the findings of this study suggest that infection upregulates the inflammatory immune response to counteract osteosarcoma‐induced immune suppression. This work informs a potential therapeutic strategy to optimize inflammatory stimuli for triggering an anti‐osteosarcoma macrophage response.     

Effects of diet and weight gain on circulating tumour necrosis factor‐α concentrations in Thoroughbred geldings

Low‐grade inflammation precedes the development of obesity‐related metabolic disorders in humans, but whether the same is true in the horse is not known. The objective of this study was to examine the effects of weight gain and diet on the inflammatory state of horses as determined by serum concentrations of tumour necrosis factor‐α (TNF), an inflammatory cytokine. Fifteen mature Thoroughbred geldings with an initial body weight (BW) of 519 ± 12 kg and body condition score (BCS) of 4.3 ± 0.1 were fed a diet of hay plus a concentrate that was either high in non‐structural carbohydrates (NSC) (i.e. starch and sugar), similar to those commercially available (CON) or one that had the energy source replaced with fat and fibre (FAT) for 32 weeks. Weight gain was achieved by feeding an additional 20 Mcal/day in excess of digestible energy maintenance requirements and resulted in a final BW of 608 ± 12 kg and BCS of 6.9 ± 0.1. Horses were exercised twice daily at a walk during the weight gain period. Horses were assessed bi‐weekly for BW and BCS. Serum TNF was analysed from blood samples collected at 4‐week intervals. Although treatment groups began the study with similar mean serum TNF concentrations, 12 weeks of FAT feeding promoted a decrease in circulating TNF that was maintained throughout the study with the exception of weeks 20 and 32. For either diet, there were no linear correlations between serum TNF concentration and BCS when horses increased in BCS from four to seven. The higher level of TNF observed in horses fed the CON diet indicates an increase in some level of systemic inflammation that was independent of their weight gain from a moderately thin to fleshy condition. The influence of diet on serum TNF concentrations should be investigated in horses fed to maintain body condition.     

TGF-β and TNF-α stimulate MMP-9 production in murine epidermal keratinocytes

Matrix metalloproteinases 2 and 9 (MMP‐2 and ‐9) are zinc‐dependent metalloenzymes and have gelatin‐degrading activity. Both MMP are known to be secreted by many types of cells and play important roles in several biological changes including tissue remodeling and wound healing. In the present study, a primary culture of murine epidermal keratinocytes was prepared and effects of transforming growth factor‐β (TGF‐β), tumor necrosis factor‐α (TNF‐α) and interferon‐γ (IFN‐γ) on expression of MMP‐2 and MMP‐9 by the keratinocytes was examined. Gelatin zymography revealed that murine epidermal keratinocytes secreted proenzyme forms of MMP‐2 and MMP‐9, but the active forms of both MMP were hardly detectable, indicating that in vitro autoactivation of these proenzymes did not occur. Both TGF‐β and TNF‐α stimulated MMP‐9 production in a dose‐dependent manner, but the MMP‐2 level was not changed. Interferon‐γ hardly affected production of MMP‐2 or MMP‐9. Ribonuclease protection assay demonstrated that TNF‐α increased the level of MMP‐9 mRNA 6‐fold compared to the control, whereas TGF‐β slightly up‐regulated it. These results suggest that expression of MMP‐9 could be regulated by several cytokines in murine epidermal keratinocytes.     

Effect of lipopolysaccharide infusion on gene expression of inflammatory cytokines in normal horses in vivo

Horses are exquisitely sensitive to bacterial endotoxin and endotoxaemia is common in colic cases. In this study, gene expression of inflammatory cytokines was characterised in the blood of healthy horses following i.v. administration of lipopolysaccharide (LPS). Six horses received an LPS infusion and 6 controls received an equivalent volume of saline. Gene expression of genes encoding interleukin (IL)‐1α, IL‐1β, IL‐6, IL‐8, and tumour necrosis factor‐α (TNF‐α) was quantified by real‐time PCR. Gene expression of all inflammatory cytokines was upregulated following administration of LPS. Interleukin‐1α, IL‐1β, IL‐8 and TNF‐α gene expression peaked at 60 min, while IL‐6 expression peaked at 90 min post LPS infusion. Interleukin‐1β and IL‐6 messenger RNA expression levels were above the baseline values 3 h post LPS infusion, whereas IL‐1α, IL‐8 and TNF‐α expression levels returned to baseline values by 3 h after LPS infusion. It was concluded that LPS infusion upregulated gene expression of inflammatory cytokines in the blood of healthy horses.     

MicroRNA‐145 regulates immune cytokines via targeting FSCN1 in Staphylococcus aureus‐induced mastitis in dairy cows

Dairy cow mastitis is a detrimental factor in milk quality and food safety. Mastitis generally refers to inflammation caused by infection by pathogenic microorganisms. Our studies in recent years have revealed the role of miRNA regulation in Staphylococcus aureus‐induced mastitis. In the present study, we overexpressed and suppressed miR‐145 to investigate the function of miR‐145 in Mac‐T cells. Flow cytometry, ELISA and EdU staining were used to detect changes in the secretion of several Mac‐T cytokines and in cell proliferation. We found that overexpression of miR‐145 in Mac‐T cells significantly reduced the secretion of IL‐12 and TNF‐α, but increased the secretion of IFN‐γ; the proliferation of bovine mammary epithelial cells was also inhibited. Using quantitative real‐time PCR (qRT‐PCR), Western blotting and luciferase multiplex verification techniques, we found that miR‐145 targeted and regulated FSCN1. Knock‐down of FSCN1 significantly increased the secretion of IL‐12, while the secretion of TNF‐α was significantly downregulated in Mac‐T cells. Upon S. aureus infection of mammary gland tissue, the body initiated inflammatory responses; Bta‐miR‐145 expression was downregulated, which reduced the inhibitory effect on the FSCN1 gene; and upregulation of FSCN1 expression promoted mammary epithelial cell proliferation to allow the recovery of damaged tissue. The results of the present study will aid in understanding the immune mechanism opposing S. aureus infection in dairy cows and will provide a laboratory research basis for the prevention and treatment of mastitis.     

Proinflammatory Cytokine and Chemokine Gene Expression Profiles in Subcutaneous and Visceral Adipose Tissue Depots of Insulin‐Resistant and Insulin‐Sensitive Light Breed Horses

Background: Insulin resistance has been associated with risk of laminitis in horses. Genes coding for proinflammatory cytokines and chemokines are expressed more in visceral adipose tissue than in subcutaneous adipose tissue of insulin‐resistant (IR) humans and rodents. Hypothesis/Objectives: To investigate adipose depot‐specific cytokine and chemokine gene expression in horses and its relationship to insulin sensitivity (SI). Animals: Eleven light breed mares. Methods: Animals were classified as IR (SI = 0.58 ± 0.31 × 10^?4 L/min/mU; n = 5) or insulin sensitive (IS; SI = 2.59 ± 1.21 × 10^?4 L/min/mU; n = 6) based on results of a frequently sampled intravenous glucose tolerance test. Omental, retroperitoneal, and mesocolonic fat was collected by ventral midline celiotomy; incisional nuchal ligament and tail head adipose tissue biopsy specimens were collected concurrently. The expression of tumor necrosis factor‐α (TNF‐α), interleukin (IL)‐1β, IL‐6, plasminogen activator inhibitor‐1 (PAI‐1), and monocyte chemoattractant protein‐1 (MCP‐1) in each depot was measured by real‐time quantitative polymerase chain reaction. Data were analyzed by 2‐way analysis of variance for repeated measures (P < .05). Results: No differences in TNF‐α, IL‐1β, IL‐6, PAI‐1, or MCP‐1 mRNA concentrations were noted between IR and IS groups for each depot. Concentrations of mRNA coding for IL‐1β (P= .0005) and IL‐6 (P= .004) were significantly higher in nuchal ligament adipose tissue than in other depots. Conclusions and Clinical Importance: These data suggest that the nuchal ligament depot has unique biological behavior in the horse and is more likely to adopt an inflammatory phenotype than other depots examined. Visceral fat may not contribute to the pathogenesis of obesity‐related disorders in the horse as in other species.     

Polysaccharides from Acanthopanax senticosus enhances intestinal integrity through inhibiting TLR4/NF‐κB signaling pathways in lipopolysaccharide‐challenged mice

To investigate the role of polysaccharide from Acanthopanax senticosus (ASPS) on lipopolysaccharide (LPS)‐induced intestinal injury, mice in three treatments were administrated orally with or without ASPS (300 mg/kg body weight) for 14 days, followed by challenge with LPS or saline. At 4 h post‐injection, blood and intestinal samples of six mice / treatment were collected. The results showed ASPS ameliorated LPS‐induced intestinal morphological deterioration, proven by improved villus height (P < 0.05) and villus height : crypt depth ratio (P < 0.05). ASPS also elevated the mucosal barrier of LPS‐challenged mice, supported by reduced plasma diamine oxidase (DAO) activity (P < 0.05) and L‐lactate (P < 0.05), increased mucosal DAO activity (P < 0.05) as well as enhanced intestinal tight junction proteins expression involving occludin‐1 (P < 0.05) and zonula occludens‐1 (P < 0.05). In addition, ASPS decreased LPS‐induced secretion of inflammatory mediators, including tumor necrosis factor (TNF)‐α (P < 0.05) and prostaglandin E₂ (P < 0.05). Also, ASPS down‐regulated messenger RNA expression of toll‐like receptor 4 (TLR4) and its downstream signals, including myeloid differentiation factor 88 (P < 0.05), TNF‐α receptor‐associated factor 6 (P < 0.05), as well as nuclear factor (NF)‐κB p65 (P < 0.05) and its protein expression. These findings suggest that ASPS improves intestinal integrity under inflammation conditions connected with inhibiting TLR4/NF‐κB signaling pathways.     

β‐carotene attenuates lipopolysaccharide‐induced inflammation via inhibition of the NF‐κB,JAK2/STAT3 and JNK/p38 MAPK signaling pathways in macrophages

β‐carotene is one of the most abundant carotenoids, has potential anti‐inflammatory effect, it has been reported that β‐carotene could suppress LPS‐induced inflammatory responses by inhibiting nuclear factor kappa B (NF‐κB) translocation, but the more detailed molecular mechanisms underlying the anti‐inflammatory action of β‐carotene remain to be fully understood. In this study, we investigated the influence of β‐carotene on the activation of JAK2/STAT3, MAPK, and NF‐κB signaling pathway induced by LPS in RAW264.7 cells and peritoneal macrophages. Cells were treated with different concentrations of β‐carotene for 3 hr after LPS treatment for 24 hr. The mRNA expression and the release of IL‐1β, IL‐6, and TNF‐α were evaluated by RT‐PCR and ELISA, and the level of signaling proteins of JAK2/STAT3, MAPK, and NF‐κB signaling pathway were detected by Western blot. The results showed that β‐carotene significantly suppressed (p < 0.05) LPS‐induced release of IL‐1β, IL‐6, and TNF‐α and their mRNA expression. LPS‐induced JAK2/STAT3, IκB/NF‐κB p65, JNK/p38 MAPK signal activation were significantly attenuated (p < 0.05) by β‐carotene in a dose‐dependent manner. In conclusion, β‐carotene could attenuate LPS‐induced inflammation via inhibition of the NF‐κB, JAK2/STAT3, and JNK/p38 MAPK signaling pathways in macrophages.     

Effects of polymyxin‐B on TNF‐α production in equine whole blood stimulated with three different bacterial toxins

Polymyxin‐B is used to treat equine systemic inflammation. Bacterial toxins other than lipopolysaccharide (LPS) contribute to systemic inflammation but the effects of polymyxin‐B on these are poorly defined. Whole blood aliquots from six healthy horses diluted 1:1 with RPMI were incubated for 21 hr with 1 μg/ml of LPS, lipoteichoic acid (LTA) or peptidoglycan (PGN) in the presence of increasing concentrations of polymyxin‐B (10–3000 μg/ml). A murine L929 fibroblast bioassay was used to measure TNF‐α activity. Polymyxin‐B significantly inhibited the effects of all three bacterial toxins. Analysis of variance showed the IC₅₀ value for polymyxin‐B for TNF‐α inhibition caused by LTA (11.19 ± 2.89 μg/ml polymyxin‐B) was significantly lower (p = .009) than the values for LPS (46.48 ± 9.93 μg/ml) and PGN (54.44 ± 8.97 μg/ml). There was no significant difference in IC₅₀ values between LPS and PGN (p > .05). Maximum inhibition of TNF‐α was 77.4%, 73.0% and 82.7% for LPS, PGN and LTA, respectively and was not significantly different between toxins. At the two highest concentrations of polymyxin‐B, TNF‐α began to increase. These data suggest that polymyxin‐B may inhibit the effects of bacterial toxins other than LPS and might be a more potent inhibitor of LTA than LPS or PGN.     

Autologous conditioned serum: the comparative cytokine profiles of two commercial methods (IRAP and IRAP II) using equine blood

Reasons for performing study: Osteoarthritis (OA) is one of the most prevalent and debilitating conditions affecting the horse. Autologous conditioned serum (ACS), commercially available as IRAP and IRAP II, is a recently developed treatment for OA in which plasma is prepared from venous blood by incubation with glass beads for 24 h. This product has been shown to increase anti‐inflammatory cytokines and growth factors in human blood. However, data for equine ACS preparations are lacking. Objectives: To characterise the protein profiles produced by commercially available ACS systems in equine blood. Methods: Blood was drawn from 5 horses into 6 groups: red top vacutainer (control), IRAP and IRAP II, with and without heparin. Samples were collected 1 or 24 h post draw and analysed for IL‐1ra, IL‐10, IGF‐1, TGF‐β, TNF‐α and IL‐1β using ELISAs. Results: Twenty‐four hour IRAP and IRAP II samples contained significantly higher levels of all cytokines relative to 1 h serum controls. At 24 h, IRAP II contained significantly higher levels of IL‐1ra and IRAP contained significantly higher levels of TNF‐α, compared to 24 h controls. In addition, TGF‐β, IL‐10 and IL‐1β in IRAP and IRAP II sera were similar to 24 h serum controls. The addition of heparin significantly reduced levels of IGF‐1, TNF‐α and TGF‐β, and significantly elevated levels of IL‐1ra. Conclusions: The cytokine profile that IRAP II produced is modestly better than IRAP. Incubation of whole blood in glass tubes stimulated cytokine synthesis, although not as efficiently as IRAP II. Potential relevance: Although high levels of IL‐1ra were found in ACS, elevation of other factors suggests these cytokines play a previously understated role in clinical improvements. Because ACS has been shown to alleviate clinical symptoms of OA, the present study suggests that factors other than IL‐1ra alone might be involved in its clinical efficacy. Species‐dependent elevations of cytokines warrant further investigation and optimisation of the systems appears to be necessary based on the differences between human and equine blood.     

Effects of Lidocaine Infusion during Experimental Endotoxemia in Horses

Background: The clinical efficacy of IV infusion of lidocaine for treatment of equine endotoxemia has not been studied. Hypothesis: Lidocaine infusion after exposure to lipopolysaccharide (LPS) will inhibit the inflammatory response and have inhibitory effects on the hemodynamic and cytokine responses to endotoxemia. Animals: Twelve horses. Methods: Two equal groups (n = 6): saline (GI) and lidocaine (GII). In all animals, endotoxin (500 ng/kg body weight [BW]) was injected intraperitoneally over 5 minutes. Twenty minutes later, animals received a bolus of GI or GII (1.3 mg/kg BW) over 5 minutes, followed by a 6‐hour continuous rate infusion of GI or GII (0.05 mg/kg BW/min). Treatment efficacy was judged from change in arterial blood pressure, peripheral blood and peritoneal fluid (PF) variables (total and differential cell counts, enzyme activities, and cytokine concentrations), and clinical scores (CS) for behavioral evidence of abdominal pain or discomfort during the study. Results: Compared with the control group, horses treated with lidocaine had significantly lower CS and serum and PF tumor necrosis factor‐α (TNF‐α) activity. At several time points in both groups, total and differential cell counts, glucose, total protein and fibrinogen concentrations, and alkaline phosphatase, creatine kinase, and TNF‐α activities were significantly different from baseline values both in peripheral blood and in PF. Conclusions and Clinical Importance: Lidocaine significantly decreased severity of CS and inhibited TNF‐α activity in PF.     

Effect of recombinant bovine tumor necrosis factor-α on hormone release in lactating cows

Tumor necrosis factor (TNF)‐α is a powerful macrophage cytokine released during infection, circulating in the blood to produce diverse effects in the organism. We examined the effect of recombinant bovine TNF‐α (rbTNF‐α) administration on hormone release in dairy cows during early lactation. Twelve non‐pregnant Holstein cows were treated subcutaneously with rbTNF‐α (2.5 µg/kg) or saline twice (at 11.00 and 23.00 hours). At 11.00 hours the next day, the cows were given growth hormone‐releasing hormone (GHRH, 0.25 µg/kg), thyrotrophin‐releasing hormone (TRH, 1.0 µg/kg), thyroid‐stimulating hormone (TSH, 10 µg/kg) or adrenocorticotropic hormone (500 µg/head) via the jugular vein. In the growth hormone‐releasing hormone challenge, the plasma growth hormone concentration was lower in the rbTNF‐α group than in the control (saline) group. The growth hormone and TSH responses to TRH were also smaller in the rbTNF‐α group than in the control. The plasma prolactin response to TRH was not affected by the rbTNF‐α treatment. In the TSH challenge, the rbTNF‐α‐treated cows had lower responses, as measured by plasma triiodothyronine and thyroxine, than the control cows. The rbTNF‐α treatment produced an increase in the basal plasma cortisol level, but the cortisol response to adrenocorticotropic hormone was the same level in both groups. The plasma concentrations of TNF‐α and interleukin‐1β in the cows were elevated by the rbTNF‐α treatment. The milk yield was reduced by the rbTNF‐α administration during 4 days. These data demonstrate that TNF‐α alters the secretion of pituitary and thyroid hormones in lactating cows. This effect may contribute to the suppression of the lactogenic function of the mammary gland observed in cases of coliform mastitis with high circulating TNF‐α levels.     

Maternal obesity stimulates lipotoxicity and up‐regulates inflammatory signaling pathways in the full‐term swine placenta

This study aimed to investigate the effects of back‐fat thickness (BF), at mating of sows, on placental lipotoxicity, oxidative stress, and inflammation. We performed iTRAQ labeling‐based proteomic analysis on term placentas obtained by vaginal delivery from BFI (15–20 mm, control) and BFII (21–27 mm, obese) sows formed according to BF at mating. Proteomic analysis revealed 413 proteins to be significantly different in placenta from BFII sows by ≥1.2‐fold. Gene ontology (GO) analysis identified proteins related to lipid metabolism and inflammatory response to be altered in placenta from obese sows. Indicative of a lipotoxic placental environment, increased placental lipid, and up‐regulated mRNA expression of lipogenic genes, including ADRP (p = .06), PPARD, FASN, ACACA, DGAT1, and LIPIN3, were associated with decreased AMPK and increased activation of WNT signaling in placenta from BFII group (p < .05). Furthermore, we observed a 18% decrease in total antioxidant capacity (TAC), increased mRNA content of pro‐inflammatory cytokines IL‐6, IL‐18, and TNF‐α, and increased activation of inflammatory NF‐κB and JNK signaling in placenta from BFII sows that was significantly associated with macrophage accumulation (p < .05). These findings suggest that maternal obesity aggravates a lipotoxic environment in pig term placenta that may be associated with placental dysfunction and impaired fetal growth.