Analysis of fecal microbiome and metabolome changes in goats with pregnant toxemia.

BACKGROUND: Pregnancy toxemia is a common disease, which occurs in older does that are pregnant with multiple lambs in the third trimester. Most of the sick goats die within a few days, which can seriously impact the economic benefits of goat breeding enterprises. The disease is believed to be caused by malnutrition, stress, and other factors, that lead to the disorder of lipid metabolism, resulting in increased ketone content, ketosis, ketonuria, and neurological symptoms. However, the changes in gut microbes and their metabolism in this disease are still unclear. The objective of this experiment was to evaluate the effect of toxemia of pregnancy on the fecal microbiome and metabolomics of does. RESULTS: Eight pregnant does suspected of having toxemia of pregnancy (PT group) and eight healthy does during the same pregnancy (NC group) were selected. Clinical symptoms and pathological changes at necropsy were observed, and liver tissue samples were collected for pathological sections. Jugular venous blood was collected before morning feeding to detect biochemical indexes. Autopsy revealed that the liver of the pregnancy toxemia goat was enlarged and earthy yellow, and the biochemical results showed that the serum levels of aspartate aminotransferase (AST) and ß-hydroxybutyric acid (B-HB) in the PT group were significantly increased, while calcium (Ca) levels were significantly reduced. Sections showed extensive vacuoles in liver tissue sections. The microbiome analysis found that the richness and diversity of the PT microbiota were significantly reduced. Metabolomic analysis showed that 125 differential metabolites were screened in positive ion mode and enriched in 12 metabolic pathways. In negative ion mode, 100 differential metabolites were screened and enriched in 7 metabolic pathways. CONCLUSIONS: Evidence has shown that the occurrence of pregnancy toxemia is related to gut microbiota, and further studies are needed to investigate its pathogenesis and provide research basis for future preventive measures of this disease.

Hepatic Perisinusoidal Myofibroblast Proliferation and Systemic Inflammatory Response Precedes Sep/Tox Hepatitis in Broilers.

Septicemia-toxemia (sep/tox) falls under U.S. Department of Agriculture (USDA) food safety Category 1 and is the most common and economically significant cause of broiler carcass condemnations. Hepatic lesions are considered a possible consequence of septicemia and associated bacterial contamination of the carcass. Thus, these lesions are considered an indicator of sep/tox (sep/tox hepatitis). This study was undertaken to analyze the histologic lesions preceding grossly visible liver lesions leading to condemnation because of sep/tox at the processing plant. Livers from carcasses of broilers condemned by USDA inspectors for sep/tox were used to establish microscopic and gross criteria of end-stage sep/tox hepatitis. Following the characterization of sep/tox hepatitis, broilers from a farm with a history of sep/tox condemnations were submitted for postmortem examination and bacteriologic investigation at four intervals during the final 20 days of production. Five healthy and five clinically ill chickens were submitted from four houses at 18, 25, 32, and 38 days of production (160 total). Microscopic lesions representing hepatic perisinusoidal myofibroblast proliferation (HPMP), periportal extramedullary granulopoiesis (PEMG), splenic follicular histiocytosis, and bone marrow cellularity (BMC) were graded subjectively for each bird, and subjective grading was evaluated with digital quantitative techniques. Perisinusoidal hepatic stellate cell morphology and progressive transformation of these cells into myofibroblasts was confirmed by immunohistochemistry for smooth muscle actin and desmin. Aerobic cultures of livers and gall bladders from sep/tox birds yielded no growth of bacteria associated with septicemia. Mild to severe HPMP was observed in all age groups, representing 28% of examined birds. Increases in inflammatory cells observed by PEMG and BMC were positively correlated with progressive HPMP and end-stage sep/tox hepatitis in broiler chickens.

Artículo regular­Proliferación de miofibroblastos perisinusoidales hepáticos y respuesta inflamatoria sistémica que precede a la hepatitis por septicemia y toxemia (sep/tox) en pollos de engorde. La septicemia-toxemia (sep/tox) se incluye en la Categoría 1 de seguridad alimentaria del Departamento de Agricultura de los Estados Unidos. (USDA) y es la causa más común y económicamente significativa de decomisos de canales de pollos de engorde. Las lesiones hepáticas se consideran una posible consecuencia de la septicemia y de la contaminación bacteriana asociada con la canal. Por lo tanto, estas lesiones se consideran un indicador de septicemia/toxemia (hepatitis sep/tox). Este estudio se llevó a cabo para analizar las lesiones histológicas que preceden a las lesiones hepáticas muy visibles que conducen a los decomisos debido a septicemia/toxemia en la planta de procesamiento. Se utilizaron hígados de canales de pollos de engorde decomisados por los inspectores del USDA por septicemia/toxemia para establecer criterios microscópicos y generales de hepatitis en etapa terminal de la septicemia/toxemia. Después de la caracterización de la hepatitis por septicemia/toxemia, los pollos de engorde de una granja con un historial de decomisos por septicemia/toxemia se sometieron a examen post mortem e investigación bacteriológica en cuatro intervalos durante los últimos 20 días de producción. Se enviaron cinco pollos sanos y cinco clínicamente enfermos de cuatro casetas a los 18, 25, 32 y 38 días de producción (160 en total). Las lesiones microscópicas que representan la proliferación de miofibroblastos perisinusoidales hepáticos (HPMP), la granulopoyesis extramedular periportal (PEMG), la histocitosis folicular esplénica y la celularidad de la médula ósea (BMC) se clasificaron subjetivamente para cada ave, y la clasificación subjetiva se evaluó con técnicas cuantitativas digitales. La morfología de las células estrelladas hepáticas perisinusoidales y la transformación progresiva de estas células en miofibroblastos se confirmó mediante inmunohistoquímica para actina y desmina del músculo liso. Los cultivos aeróbicos de hígados y vesícula biliar de aves con septicemia/toxemia no produjeron crecimiento de bacterias asociadas con la septicemia. Se observó proliferación de miofibroblastos perisinusoidales hepáticos de leve a severa en todos los grupos de edad, lo que representa el 28% de las aves examinadas. Los aumentos en las células inflamatorias observados por granulopoyesis extramedular periportal y celularidad de la médula ósea se correlacionaron positivamente con proliferación progresiva de miofibroblastos perisinusoidales hepáticos y con hepatitis por septicemia/toxemia en etapa terminal en pollos de engorde.

Changes in cardiac biomarkers in goats naturally affected by pregnancy toxemia.

Pregnancy toxemia (PT) is considered one of the most common metabolic diseases with high impact on the production of small ruminants. The objective of this study was investigate possible myocardial damage in goats affected with PT by the determination of serum myocardial biomarkers CK-MB and cTnI. A total of 44 goats affected with PT, and 10 apparently healthy goats (control group or CG) were used in the study. In goats with PT, the serum concentrations of cTnI (0.43 ng/mL) were significantly higher than that in CG goats (0.06 ng/mL). Although CK-MB showed no significant difference, it was approximately three times higher in animals with PT. The serum concentrations of insulin were significantly lower in PT goats (5.03 ppmol/L) compared to CG goats (10.66 pmol/L). The serum concentrations of cortisol in PT goats (155.41 nmol/L) were significantly higher than that in CG goats (36.58 nmol/L). Results of this study indicate that a clinically significant myocardial damage might occur in goats affected with PT leading to significant elevations in values of cTnI and CK-MB. Therefore, these parameters could be used as a potential prognostic indicator in goats affected with this important disease.

Definition of prepartum hyperketonemia in dairy goats.

A prospective cohort study was conducted on 1,081 dairy goats from 10 commercial herds in Québec (Canada) to define prepartum hyperketonemia based on optimal blood ß-hydroxybutyrate acid threshold values for the early prediction of pregnancy toxemia (PT) and mortality in late-gestation dairy goats. All pregnant goats had blood sampled weekly during the last 5wk of pregnancy. The blood was analyzed directly on the farm for ß-hydroxybutyrate acid quantification using a Precision Xtra meter (Abbott Diabetes Care, Saint-Laurent, QC, Canada). Body condition scores on the lumbar region and sternum were noted. Each goat was classified as being at low (n=973) or high risk (n=108) of having PT by producers based on a standardized definition. The optimal threshold for predicting a PT diagnosis or mortality for each week before kidding was determined based on the highest sum of sensitivity and specificity. The association between hyperketonemia and subsequent PT was tested using a multivariable logistic regression model considering hyperketonemia at wk 4 prepartum, litter size, and body condition score at wk 4 prepartum as covariates, and herd and parturition cohort as random effects. The association between mortality and hyperketonemia was also tested using a logistic regression model accounting for the presence or absence of treatment during the last month of pregnancy. The hyperketonemia definition based on PT varied between ≥0.4 and ≥0.9mmol/L during the last 5wk prepartum. Goats affected by hyperketonemia at wk 4 prepartum and with a large litter size (≥3 fetuses) had 2.1 and 40.5 times the odds, respectively, of subsequent PT than other goats. Hyperketonemia definitions based on mortality varied between ≥0.6 and ≥1.4mmol/L during the last 4wk prepartum, and was ≥1.7mmol/L during the first week postpartum. Goats affected by hyperketonemia and treated by producers had 3.4 and 11.8 times the odds, respectively, of subsequent mortality than did other goats. These results showed that prepartum hyperketonemia could be defined in dairy goats using subsequent risks of PT or mortality during the last month of pregnancy.

Identification of the 1B vaccine strain of Chlamydia abortus in aborted placentas during the investigation of toxaemic and systemic disease in sheep.

CASE HISTORY: One hundred and forty Cheviot and 100 Suffolk cross Mule primiparous 1-2-year-old ewes, from a flock of about 700 ewes, were vaccinated with an attenuated live 1B strain Chlamydia abortus vaccine about 4 weeks before ram introduction (September 2011). Between 08 March and 01 April 2012, 50 2-year-old ewes aborted and 29 of these died, despite antimicrobial and anti-inflammatory treatment and supportive care. PATHOLOGICAL FINDINGS: Seven fetuses and three placentae from five 2-year-old ewes were submitted for pathological investigation. The aborted fetuses showed stages of autolysis ranging from being moderately fresh to putrefaction. Unusual, large multifocal regions of thickened membranes, with a dull red granular surface and moderate amounts of grey-white surface exudate were seen on each of the placentae. Intracellular, magenta-staining, acid fast inclusions were identified in Ziehl Neelsen-stained placental smears. Immunohistochemistry for Chlamydia-specific lipopolysaccharide showed extensive positive labelling of the placental epithelia. LABORATORY FINDINGS: Molecular analyses of the aborted placentae demonstrated the presence of the 1B vaccine-type strain of C. abortus and absence of any wild-type field strain. The vaccine strain bacterial load of the placental tissue samples was consistent with there being an association between vaccination and abortion. DIAGNOSIS: Initial laboratory investigations resulted in a diagnosis of chlamydial abortion. Further investigations led to the identification of the 1B vaccine strain of C. abortus in material from all three of the submitted aborted placentae. CLINICAL RELEVANCE: Timely knowledge and understanding of any potential problems caused by vaccination against C. abortus are prerequisites for sustainable control of chlamydial abortion. This report describes the investigation of an atypical abortion storm in sheep, and describes the identification of the 1B vaccine strain of C. abortus in products of abortion. The significance of this novel putative association between the vaccine strain of C. abortus and severe clinical disease is unknown. Aspects of the approach that is described are relevant to the investigation of all outbreaks of ovine abortion, irrespective of the diagnosis. Awareness of the changing role of C. abortus as a major global cause of abortion ought to reinforce the importance of monitoring of adequate biosecurity in those countries which are currently free from chlamydial abortion.

Prepartum administration of recombinant bovine somatotropin (rBST) on adaptation to subclinical ketosis of the ewes and performance of the lambs / Administração pré-parto de somatotropina bovina recombinante (rBST) na adaptação da cetose subclínica de ovelhas e no desempenho dos cordeiros

The aim of this study was to determine the effect of prepartum rbST injection on the metabolic profile of pregnant ewes induced to subclinical ketosis, as well as the metabolism until seven days of life and weight gain until seven weeks of life of the lambs. Twenty seven pregnant ewes of the pantaneiro genetic group were used, divided into two groups: rbST group (n = 14) and control group (n = 13). The rbST group received two applications of 1 mg/kg of rbST, at 97 and 111 days gestation, while the control group received placebo injections. There were significant differences between groups in levels of GGT in the ketosis post induction period and BHB concentrations in the postpartum period. Concentrations of glucose, urea, phosphorus, albumin, cholesterol, AST, NEFA and insulin were not different between dams from the two groups in different periods of the study (P>0.05). There was an effect of rbST on body weight observed already at fourteen days of life (P<0.0001), there was an increase in serum phosphorus levels at birth of lambs (P=0.0014), and albumin at seven days of life (P = 0.0014) of the lambs, with no difference between groups for the other metabolites. Therefore, the use of rbST was effective in increasing the weight of the lambs until the seventh week of life. In addition, rbST treatment had positive effects on the dam metabolism with reduction of liver overload, as indicated by decreased GGT after ketosis induction and decreased BHB at the postpartum period.

O objetivo deste estudo foi determinar o efeito da administração pré-parto de rbST sobre o perfil metabólico de ovelhas induzidas à cetose subclínica, assim como sobre o metabolismo até sete dias de vida e ganho de peso até sete semanas de vida dos cordeiros. Vinte e sete ovelhas prenhas do grupo genético pantaneiro foram divididas em dois grupos: grupo rbST (n = 14) e grupo controle (n = 13). O grupo rbST recebeu duas aplicações de 1mg/kg de rbST, aos 97 e 111 dias de gestação, ao passo que o grupo controle recebeu injeções de placebo. Houve diferenças significativas entre os grupos nos níveis de GGT no período de pós-indução de cetose e concentrações de BHB no período pós-parto. As concentrações de glicose, ureia, fósforo, albumina, colesterol, AST, NEFA e insulina não foram diferentes entre os grupos nos diferentes períodos do estudo (P>0,05). Houve um efeito de rbST no peso corporal dos cordeiros já observado nos 14 dias de vida (P<0,0001), verificou-se um aumento dos níveis séricos de fósforo ao nascimento de cordeiros (P=0,0014) e albumina de sete dias de vida (P=0,0014 ), e não houve diferença entre os grupos para os outros metabólitos. Portanto, a utilização de rbST foi eficaz em aumentar o peso dos cordeiros até a sétima semana de vida. Além disso, a aplicação de rbST teve efeitos positivos no metabolismo com a redução da sobrecarga do fígado, como indicado pela diminuição da GGT após a indução da cetose e diminuição de BHB no período pós-parto.

Prolapso vaginal e uterino em ovelhas / Uterine and vaginal prolapse in ewes

O presente trabalho objetivou realizar um estudo retrospectivo sobre os prolapsos vaginal e uterino em ovelhas atendidas no Serviço de Clínica de Bovinos e Pequenos Ruminantes (CBPR) da FMVZ/USP no período compreendido entre 2000 a 2010, no qual, foram atendidas 56 ovinos com problemas inerentes ao sistema reprodutivo, dessas, 25 apresentaram prolapso vaginal ou uterino (44,6%). O prolapso vaginal total foi o de maior frequência (72%). As ovelhas acometidas, em sua maioria, possuíam idade superior a quatro anos (64%), eram sem raça definida (44%) ou da raça Ile de France (40%). As manifestações clínicas observadas durante a maioria dos atendimentos foram: taquipnéia, taquicardia, mucosas oculares avermelhadas indicando estado de toxemia, decúbito esternal ou lateral, apatia e anorexia. O tratamento instituído para todos os casos foi a limpeza, desinfecção e reintrodução do órgão prolapsado. A sutura de Bühner foi feita em 84% dos casos e a histeropexia em um caso (4%). A evolução foi satisfatória em 80% dos casos atendidos, nos demais casos (20%) observou-se óbito da fêmea acometida. Do total de óbitos, os prolapsos vaginais foram responsáveis por 60% (3/5) e os prolapsos uterinos por 40% (2/5). A etiologia dos prolapsos não foi definida nos casos atendidos, sendo esses associados com o período pós-parto em sua maioria (56%), provavelmente associados com quadros de hipocalcemia, altas concentrações séricas de estrógeno e hipertonia uterina. Além disso, a predisposição genética não pode ser descartada.

This study aimed to conduct a retrospective study on vaginal and uterine prolapse in sheep seen at the Clinic and Surgery on Cattle and Small Ruminants (CBPR) at University of São Paulo, from 2000 to 2010. During this period, 56 sheep were treated with problems of the reproductive system. Of these, 25 ewes had vaginal or uterine prolapse (44.6%). The total vaginal prolapse was the most frequently (72%). The majority of sheep that was affected were 4 years old (64%); most were mixed breed (44%) and 40% were Ile de France. Main clinical signs were increased cardiac and respiratory rates, congested ocular mucosa, sternal or lateral recumbence, apathy and anorexia, suggesting toxemia. The treatment of all cases was the cleaning and disinfection of the prolapsed organ and its reintroduction. The Bühner suture was made in 84% of the cases. The uterus fixation was made in one case (4%). Recovered was observed in 80% of the cases and 20% of the patients died. Vaginal prolapse corresponded to 60% of the deaths and uterine prolapse to 40%. The etiology of the prolapses had not been defined, but most cases (56%) occurred during the postpartum period, probably associated with hypocalcaemia, high serum concentrations of estrogen or uterine hypertonia. Furthermore, a genetic predisposition on affected sheep cannot be discharged.

Acid-base and electrolyte status during early induced pregnancy toxaemia in goats.

Eleven Murciano-Granadina goats in late pregnancy were separated into two groups (1) control (n=6) and (2) fasting for 72 h to induce pregnancy toxaemia (n=5). Venous blood was taken daily to determine acid-base and electrolyte parameters. Significant decreases in blood pH, bicarbonate concentration and base excess, and a significant increase in anion gap were observed after 24h of fasting. These changes were significantly correlated with non-esterified fatty acid concentration. No significant changes were observed in pCO(2), and electrolyte or lactate concentrations. Clinical signs of pregnancy toxaemia in fasted goats appeared by 72h post-fasting. These signs and the changes in acid-base balance disappeared once feed was reintroduced. Blood pH, bicarbonate concentration, base excess and anion gap could be indicators of early pregnancy toxaemia in goats.

Distinct physiologic and inflammatory responses elicited in baboons after challenge with Shiga toxin type 1 or 2 from enterohemorrhagic Escherichia coli.

Shiga toxin-producing Escherichia coli is a principal source of regional outbreaks of bloody diarrhea and hemolytic-uremic syndrome in the United States and worldwide. Primary bacterial virulence factors are Shiga toxin types 1 and 2 (Stx1 and Stx2), and we performed parallel analyses of the pathophysiologies elicited by the toxins in nonhuman primate models to identify shared and unique consequences of the toxemias. After a single intravenous challenge with purified Stx1 or Stx2, baboons (Papio) developed thrombocytopenia, anemia, and acute renal failure with loss of glomerular function, in a dose-dependent manner. Differences in the timing and magnitude of physiologic responses were observed between the toxins. The animals were more sensitive to Stx2, with mortality at lower doses, but Stx2-induced renal injury and mortality were delayed 2 to 3 days compared to those after Stx1 challenge. Multiplex analyses of plasma inflammatory cytokines revealed similarities (macrophage chemoattractant protein 1 [MCP-1] and tumor necrosis factor alpha [TNF-alpha]) and differences (interleukin-6 [IL-6] and granulocyte colony-stimulating factor [G-CSF]) elicited by the toxins with respect to the mediator induced and timing of the responses. Neither toxin induced detectable levels of plasma TNF-alpha. To our knowledge, this is the first time that the in vivo consequences of the toxins have been compared in a parallel and reproducible manner in nonhuman primates, and the data show similarities to patient observations. The availability of experimental nonhuman primate models for Stx toxemias provides a reproducible platform for testing antitoxin compounds and immunotherapeutics with outcome criteria that have clinical meaning.

An outbreak of enterotoxaemia at livestock farm during subtropical summer.

Present investigations were carried out on 10 dead animals including eight in lambs, one in goat kid and one in calf during subtropical summer at a local farm. The weather was hot and humid with rain occurring during the period. The history suggests an association of weather and concentrate/lush green diet/fodder with occurrence of the disease. The most consistent clinical signs reported were no interest in feeding, herding in a corner with head down, diarrhea of low degree and temperature around 102 degrees F. At postmortem examination, the most consistent findings were swollen soft kidneys, hydropericardium, congested and edematous lungs, congested liver, myocardial hemorrhages and ballooning of intestines. The histopathological examination revealed the most striking changes in kidney of vacuolation in renal tubular epithelial cells and increased Bowman's space in the glomeruli. The histopathological examination of liver revealed congestion. Lungs revealed congestion and edema. The urine from urinary bladder collected showed high glucose. The deaths in these animals were probably due to enterotoxaemia type D.

[Effectiveness and disposition of the newly developed cephalosporin cefquinome in puerperal septicemia and toxemia in gilts]. / Untersuchungen zur Wirksamkeit und Verträglichkeit des neu entwickelten Cephalosporins Cefquinom bei an puerperaler Septikämie und Toxämie erkrankten Sauen.

Epizootiological, clinical, bacteriological and haematological studies were carried out to assess the effectiveness of the recently developed cephalosporin preparation Cefquinome in the treatment of the puerperal septicaemia and toxaemia syndrome. Cefquinome was administered at three different doses (1, 2 and 4 mg/kg BW) to 188 sows with feverish puerperal illness. Amoxicillin (7 mg/kg BW) was used as a control drug. In 41% of cases endometritis was a monoinfection whereas in 70% of mammary infections mixed infections were diagnosed. Results showed that for therapy of puerperal septicaemia and toxaemia Cefquinome at doses of 2 mg/kg BW and 4 mg/kg BW is clearly more effective than the control drug Amoxicillin and Cefquinome at its lowest dose of 1 mg/kg BW.

[Clinical examinations of gilts with puerperal septicemia and toxemia]. / Klinische Erhebungen bei Sauen mit puerperaler Septikämie und Toxämie.

In a clinical examination on 78 gilts suffering from feverish puerperal illness the signs were documented. 42 gilts without puerperal disturbances out of the same farms (n = 22) were used as a control. Data for statistics were recorded in minute-books, listing 38 parameters, partly divided in three or four subunits. Within the first 24 hours after parturition 60% of the probands were registered, the other gilts up to the 72nd hour after parturition. 42.3% of the patients beared for the first or second time. Duration of partus averaged more than six hours in 85.9% of patients with consequent puerperal illness, whereas probands of the same age in the control group finished parturition in less than three hours (78.8%; p < or = 0.01). Frequency of obstetrical intervention measured 27% in the group of patients, in the group of probands 9.5% (p < or = 0.05). Gilts with following puerperal illness delivered 1.1 piglets more than healthy individuals (p < or = 0.05) and showed an increased stillbirth rate (p < or = 0.01). Signs of clinical interest in puerperal illness were increase of body temperature (p < or = 0.01), cardiac rate (p < or = 0.01) and respiratory frequency (p < or = 0.01). Approximately 75% of the patients showed anorexia, 66% abnormal faecal consistency. 24.4% of the diseased animals showed exclusively signs of mastitis, in 29.5% there was a combination of mastitis and inflammatory affection of the genital system to be diagnosed, in 46.1% of the cases a solitary infection of the reproductive tract was to be stated. Predominantly E. coli, followed by Staphylococcus spp. and Streptococcus spp. were isolated from the genital tract. The results of this study emphasize the clinical necessity to differentiate between isolated mastitis and puerperal septicaemia respectively toxaemia in cases of feverish puerperal illness. Puerperal septicaemia and toxaemia can, but do not have to be associated with mastitis in gilts.

Management of a femoral fracture complicated by clostridial myositis.

A clinical case of clostridial myositis secondary to a comminuted femoral fracture is described. This case is unusual because, despite the severe degree of obvious muscle necrosis and gas production, the dog had minimal signs of systemic toxicity. Union of the fracture was achieved but six months postoperatively muscular contracture had resulted in permanent stifle extension.

Evaluation of plasma alpha-2-macroglobulin and interactions with tumour necrosis factor-alpha in horses with endotoxemic signs.

The electrophoretic position and behavior of the native and activated forms of equine plasma alpha-2-macroglobulin (alpha 2M) were characterized and compared to human alpha 2M by nondenaturing polyacrylamide-gel electrophoresis (PAGE). Plasma alpha 2M was also compared between 6 normal horses and 6 horses with clinical signs of colic and endotoxemia due to volvulus or enteritis. Native and activated forms of alpha 2M were quantified by PAGE and densitometry. Binding of radio-labeled recombinant human tumour necrosis factor-alpha (125I-rhTNF-alpha) to native and activated forms of equine alpha 2M was also evaluated by autoradiography and densitometry of PAGE. Equine plasma alpha 2M migrated as a single band at a position equivalent to native human alpha 2M. Methylamine-reacted equine plasma samples resulted in faster migration of alpha 2M in a similar position to activated human alpha 2M. However, in methylamine-reacted equine plasma, an intermediate alpha 2M band was consistently present between the bands corresponding to native and activated alpha 2M. Amounts of plasma alpha 2M were similar in normal and endotoxemic horses, and remained in the electrophoretically slow or unreacted native form. The vast majority of 125I-rHuTNF-alpha did not bind to alpha 2M or other equine plasma proteins. 125I-rHuTNF-alpha bound weakly to both native and fast methylamine-reacted equine forms of alpha 2M, although binding was better to the activated form. This study indicates that: (1) equine plasma alpha 2M behaves similarly to human alpha 2M on PAGE, (2) plasma alpha 2M of horses can be activated to electrophoretically fast forms, but it is neither activated nor depleted during endotoxemia, and (3) the binding interactions between equine alpha 2M and TNF-alpha are too low to implicate equine alpha 2M as a regulator of TNF-alpha during endotoxemia in horses.

The effects of glibenclamide, a blocker of K+ ATP-sensitive potassium channels, on diaphragmatic fatigue during endotoxaemia in pigs.

An in vivo porcine model of endotoxaemia was used to study the effects of glibenclamide, a K+ ATP-sensitive potassium channel blocker. Escherichia coli lipopolysaccharides (LPS, 70 micrograms/kg, i.v., as a bolus) were infused into anaesthetized, mechanically ventilated, indomethacin-treated pigs. After 120 min of endotoxaemia, glibenclamide was administered (10 mg/kg, i.v., over 5 min) to half the pigs. The strength at different frequencies of stimulation (10, 20, 30, 50 Hz, 20 V,) 1 s) and the endurance capacity (10 Hz, 20 V, 30 s) of the diaphragm were evaluated after 120 min of endotoxaemia and 5, 10, 20 and 30 min after drug infusion. Glibenclamide transiently increased the blood pressure without changing the decreased cardiac output and at the same time further impaired the diaphragmatic activity. The reduced ability of the diaphragm to generate force in response to different electrical stimulations was shown by a significant reduction in strength. The endurance index decreased 5 min after glibenclamide infusion, returning to the pre-glibenclamide values by 150 min. These results indicate that glibenclamide modifies the activity of vascular smooth muscle and of the diaphragm.

Mediators and vascular effects in response to endotoxin.

Recent experimental findings indicate that endotoxin (i.e. lipopolysaccharide) interacts with specific membrane receptors localized to mononuclear phagocytic cells and neutrophils. Binding of endotoxin to these cells, together with endotoxin-induced activation of host vascular endothelium, initiates a series of signal transduction events that culminate in release of numerous biochemical mediators. The latter include cytokines, platelet-activating factor, thromboxane A2, prostaglandins, leukotrienes, nitric oxide, proteases, toxic O2 radicals, and vasoactive amines. These mediators orchestrate complex biological interactions and amplification signals that lead to cardiopulmonary dysfunction and multi-organ failure within 4-6 h of experimental infusion of endotoxin into animals. The pathophysiological changes include decreased cardiac output, systemic hypotension, decreased blood flow and O2 delivery to tissues, intense pulmonary vasoconstriction and hypertension, bronchoconstriction, increased permeability, pulmonary oedema, ventilation-to-perfusion inequalities, hypoxaemia, and haemoconcentration. Metabolic alterations include increased blood lactate and pyruvate, metabolic acidosis, hyperkalaemia and hypoglycaemia. Potential therapeutic modalities for treatment of endotoxaemia/septic shock include specific antagonists directed against lipopolysaccharide, cytokine, and platelet-activating factor receptors, monoclonal antibodies directed against cytokines and lipid A/core polysaccharides of endotoxin, antiproteases, and agents that block release of toxic O2 and arachidonic acid metabolites.