Equine Endotoxaemia Research Articles

Anti-inflammatory effects of four potential anti-endotoxaemic drugs assessed in vitro using equine whole blood assays.

Despite the severity and common occurrence of equine endotoxaemia, the available anti-endotoxic treatments do not effectively target key inflammatory mechanisms such as leucocyte activation and cytokine production. In this study, four compounds with potential anti-endotoxic effects, namely rolipram, azithromycin, ethyl pyruvate and metformin, were investigated in vitro using equine whole blood stimulated with bacterial lipopolysaccharide. TNF-α and IL-1β production were measured in plasma. Rolipram was the most potent inhibitor of cytokine production (IC50 0.84 and 4.68 μm for TNF-α and IL-1β, respectively) with almost complete inhibition of TNF-α, but inhibited IL-1β by only 39.46%. Azithromycin produced almost complete inhibition of both cytokines, but tended to be less potent than rolipram (IC50 10.66 and 17.4 μm for TNF-α and IL-1β, respectively). Metformin inhibited TNF-α production with similar potency to rolipram and azithromycin (IC50 3.35 μm) but showed significantly lower efficacy (45.93%; P < 0.05), and had no inhibitory effect on IL-1β. Ethyl pyruvate was the least potent (IC50 68.35 μm and >10 mm for TNF-α and IL-1β production, respectively). Further work is required to investigate whether these or related compounds may have potential use in the treatment of equine endotoxaemia in vivo.

Pattern recognition receptors in equine endotoxaemia and sepsis

SummaryPattern recognition receptors (PRRs) on host cells detect pathogens to activate innate immunity which, in turn, initiates inflammatory and adaptive immune responses. Successful activation of PRRs is, therefore, critical to controlling infections and driving pathogen‐specific adaptive immunity, but overactivity of PRRs causes systemic inflammation, which is detrimental to the host. Here we review the PRR literature as it relates to horses and speculate on the role PRRs may play in sepsis and endotoxaemia.

Regulation of platelet activating factor‐induced equine platelet activation by intracellular kinases

Lipopolysaccharide (LPS) can activate equine platelets directly or indirectly, via leukocyte-derived platelet activating factor (PAF). Thromboxane (Tx) production by LPS-stimulated equine platelets requires p38 MAPK and this kinase has been suggested as a therapeutic target in endotoxaemia. The present study has utilised selective inhibitors to investigate the role of p38 MAPK and two other kinases, phosphatidylinositol-3 kinase (PI3K) and protein kinase C (PKC), in regulating PAF-induced Tx production, aggregation and 5-HT release in equine platelets, and the modification of these responses by LPS. LPS enhanced PAF-induced 5-HT release, an effect that was reduced by the p38 MAPK inhibitor, SB203580 (60 +/- 8% reduction; n = 6). SB203580 did not affect responses to PAF alone; whereas inhibition of PKC reduced PAF-induced 5-HT release, Tx production and aggregation (maximal inhibition by the PKCdelta inhibitor, rottlerin: 69 +/- 13%, 63 +/- 14% and 97 +/- 1%, respectively; n = 6). Wortmannin and LY249002, which inhibit PI3K, also caused significant inhibition of PAF-induced aggregation (maximal inhibition 78 +/- 3% and 88 +/- 2%, respectively; n = 6). These data suggest that inhibition of platelet p38 MAPK may be of benefit in equine endotoxaemia by counteracting some of the effects of LPS. However, detrimental effects of platelet activation mediated by PAF and not enhanced by LPS are unlikely to be markedly affected.

Mesenteric Arterionecrosis in Natural and Experimental Equine Endotoxaemia

To test the hypothesis that mesenteric arterionecrosis (MA) occurs in horses with naturally occurring endotoxaemia (ET) and in those with experimentally induced ET, the mesentery and gastrointestinal tract of 21 Thoroughbred racehorses (15 with spontaneous colic suspected to be due to ET, and six with experimentally induced ET) were examined. MA, which occurred in 13 of the 15 horses with spontaneous colic and in all six of the cases of experimental ET, was morphologically similar in the two groups of animals. This suggested that the pathogenesis of the MA was fundamentally similar in the two groups, and that MA is a pathognomonic feature of equine ET. In addition to histolysis of the arterial walls associated with infiltration of blood components, changes were noted in the medial smooth muscle including formation of many intracellular vacuoles within single smooth muscle cells, cytoplasmolysis, necrosis with granules and vacuoles, and coagulation necrosis; similar changes have been observed in cases of prolonged angiospasm or vasoconstriction. It is suggested that the effects of sustained arterial contraction leading to intimal and medial damage influence the pathomorphogenesis of MA.

Ketamine inhibits LPS-induced tumour necrosis factor-alpha and interleukin-6 in an equine macrophage cell line

Ketamine is widely used in equine anaesthesia. Beside its anaesthetic and analgesic properties, ketamine possesses a cytokine-modulating activity. However, to date, no data are available regarding the inhibitory effect of ketamine on the cytokine response in horses. In horses, cytokines such as tumour necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) play a pivotal role in the pathogenesis of equine endotoxaemia following gastrointestinal disorders. Hence, the objective of this study was to assess the influence of ketamine on LPS-induced TNF-alpha and IL-6 formation in an equine macrophage cell line (eCAS cells). The results demonstrate a cytokine-modulating activity of ketamine in an equine cell line, suggesting a beneficial role for ketamine in the treatment of equine endotoxaemia.

Equine endotoxaemia ‐ A state‐of‐the‐art review of therapy

The pathophysiology of endotoxaemia, a leading cause of death in the horse, is beginning to be understood in greater detail. Endotoxin may be absorbed into the systemic circulation in a number of different ways: most commonly the body's normal defense mechanisms are disrupted or bypassed, or the normal clearance mechanisms overwhelmed. Following this wide-spread effects are observed, although the most significant are seen in the cardiovascular system. Fever, arterial hypoxaemia and signs of abdominal pain are also common. With increased understanding of the disease new therapeutic agents have become available, however, while the newer agents offer some advantages it is important to recognise that supportive care is the mainstay of treatment for endotoxaemia. Supportive care consists of aggressive fluid therapy (crystalloid, colloid and hypertonic), the administration of non-steroidal antiinflammatory drugs and, where appropriate, antimicrobials. The principles of supportive care are discussed in detail. Other therapies such as hyperimmune plasma, polymyxin B, pentoxifylline, dimethyl sulfoxide and heparin are commonly used in the treatment of equine endotoxaemia and their use is reviewed here. Furthermore, newer agents such as anti-tumour necrosis factor antibodies, detergent, activated protein C and insulin, which have yet to gain widespread acceptance but may have an important role in the treatment of endotoxaemia in the future, are examined.

Arterionecrosis of the Equine Mesentery in Naturally Occurring Endotoxaemia

This report describes the mesenteric arteriolar lesions in a Thoroughbred racehorse with endotoxaemia due to colic. The vascular lesions consisted of a striking loss of medial smooth muscle cells, associated with granular cell debris derived from necrosed muscle cells, plasma insudation, erythrocyte infiltration and the deposition of a fibrinoid substance (fibrinoid degeneration) in the entire arterial wall, possibly produced by the infiltration of blood components through endothelial cell junctions into the arterial wall. The morphology of the mesenteric arteriolar necrosis closely resembled that seen in experimental equine endotoxaemia and in horses that died from colic; it also resembled that of Shiga toxin-induced arteriolar lesions in oedema disease of swine and of the arterionecrosis in human cerebral arteries that may lead to hypertensive intracerebral haemorrhage.

Lipopolysaccharide and interferon gamma activate nuclear factor kappa B and induce cyclo-oxygenase-2 in equine vascular smooth muscle cells

Equine endotoxaemia is an important cause of morbidity and mortality in horses caused by the interaction of bacterial lipopolysaccharide (LPS) with cells such as macrophages and vascular smooth muscle. In this study we isolated equine vascular smooth muscle from a variety of vessels and stimulated it with LPS and human interferon (hIFN)-γ. Using reverse transcriptase polymerase chain reaction (rt-PCR) and Western blot analysis we show that cyclooxygenase-2 (COX-2) is readily expressed in equine vascular smooth muscle. Vascular smooth muscle cells produced prostaglandin E2 in response to LPS and hIFNγ. Using similar approaches we saw very limited expression of inducible nitric oxide synthase (iNOS) in only one vascular smooth muscle preparation. LPS and IFNγ caused translocation of the transcription factor nuclear factor kappa B (NfκB) to the nucleus in equine cells suggesting the limited iNOS production seen in our cells is not due to deficits in this signal transduction pathway. These data suggest that in equine vascular smooth muscle COX-2 and NfκB are likely to play important roles in the pathogenesis of equine endotoxaemia.

Administration of a receptor antagonist for platelet‐activating factor during equine endotoxaemia

Platelet-activating factor (PAF) is an important mediator of endotoxaemia and various PAF receptor antagonists prevent many of the adverse effects of experimental endotoxaemia in laboratory animals. In this study a specific PAF receptor antagonist was used to investigate the role of PAF in equine endotoxaemia. At an interval of not greater than 10 days, 6 horses were each challenged with endotoxin and endotoxin with concurrent administration of SRI 63-441, a PAF receptor antagonist. The order of the treatments was randomised. Clinical signs, serum biochemical and coagulation profiles, and platelet aggregation in vitro were monitored in all horses for 24 h after treatment. Challenge with endotoxin increased maximal platelet aggregation induced by PAF. This response was blocked by administration of SRI 63-441 concurrently with endotoxin. No changes in percentage maximal platelet aggregation to ADP or collagen were noted after administration of endotoxin. The PAF receptor antagonist delayed the onset of fever, tachycardia, leucopenia and lactic acidaemia. Lack of more profound beneficial alterations of the horses' responses to endotoxin may have been due to the low dose of endotoxin administered in this model or to only partial effectiveness of SRI 63-441 in blocking the effects of endotoxin-induced PAF.

Effects of a specific thromboxane synthetase inhibitor in equine endotoxaemia

Thromboxane A2 may play a major role in circulatory shock. In some species, thromboxane synthetase inhibitors have a beneficial effect on shock induced by endotoxin, trauma, sepsis and administration of arachidonate. In some shock models, however, results with thromboxane synthetase inhibitors have been conflicting. The effect of UK-38,485, a selective thromboxane inhibitor, was evaluated in ponies injected with endotoxin intraperitoneally. Four groups of ponies were used to compare the effects of endotoxin alone, UK-38,485 alone, treatment with UK-38,485 before endotoxin challenge and treatment with UK-38,485 after endotoxin challenge. Haematological, metabolic, eicosanoid and clinical responses in each group were evaluated. The results indicated that UK-38,485 is an effective inhibitor of thromboxane A2 generation following endotoxin challenge. Prostacyclin values were elevated compared with baseline in ponies administered UK-38,485 and endotoxin. However, prostacyclin values were not significantly different from those of ponies receiving endotoxin alone. Furthermore, UK-38,485 failed to attenuate the haematological, metabolic and clinical manifestations commonly seen in the pony after endotoxin challenge.

Equine gastroenteritis and endotoxaemia – prohpylaxis and therapy with an endotoxic hyperimmunserum

Equine gastroenteritis and endotoxaemia – prohpylaxis and therapy with an endotoxic hyperimmunserum