Idiopathic immune-mediated polysynovitis in three horses.

This paper describes the clinical, laboratory and histological findings in three horses with immune-mediated polysynovitis; they had lost weight, suffered intermittent fever, were lethargic and stiff, and had effusions in several joints. Laboratory abnormalities included anaemia, leucocytosis, hyperfibrinogenaemia and hyperglobulinaemia. The diagnosis was based on the presence of a suppurative, non-septic inflammation in at least two different joints in each of the horses and the presence of immunoglobulins in the synovial membrane of one of them. The horses were treated with a combination of dexamethasone and azathioprine, and responded well to the initial treatment. Twenty months after its last re-evaluation, the first horse was being maintained on azathioprine because similar clinical signs had recurred after the cytotoxic drug was discontinued; the second horse was finishing a tapering course of prednisolone 15 months after its first examination, and the third horse was euthanased five months after it was first examined as a result of an unrelated injury.

Purpura haemorrhagica in 53 horses.

The medical records of 53 horses with purpura haemorrhagica were reviewed. Seventeen of them had been exposed to or infected with Streptococcus equi, nine had been infected with Corynebacterium pseudotuberculosis, five had been vaccinated with S. equi M protein, five had had a respiratory infection of unknown aetiology, and two had open wounds; the other 15 cases had no history of recent viral or bacterial infection. The horses were between six months and 19 years of age (mean 8.4 years). The predominant clinical signs were well demarcated subcutaneous oedema of all four limbs and haemorrhages on the visible mucous membranes; other signs included depression, anorexia, fever, tachycardia, tachypnoea, reluctance to move, drainage from lymph nodes, exudation of serum from the skin, colic, epistaxis and weight loss. Haematological and biochemical abnormalities commonly detected were anaemia, neutrophilia, hyperproteinaemia, hyperfibrinogenaemia, hyperglobulinaemia and high activities of muscle enzymes. All of the horses were treated with corticosteroids; 42 also received non-steroidal anti-inflammatory drugs and 26 received antimicrobial drugs. Selected cases received special nursing care, including hydrotherapy and bandaging of the limbs. Most of the horses were treated for more than seven days and none of them relapsed. Forty-nine of the horses survived, one died and three were euthanased, either because their severe clinical disease failed to respond to treatment or because they developed secondary complications. Two of the four non-survivors had been vaccinated against S. equi with a product containing the M protein, one had a S. equi infection and the other had a respiratory infection of undetermined aetiology.

Pulmonary coccidioidomycosis in a neonatal foal.

A 13-day-old foal with profound tachypnoea and respiratory distress was examined. Thoracic radiographs revealed a severe, diffuse miliary pattern, and the foal was markedly hypoxaemic. It failed to improve with empirical treatment, and was euthanased. Lesions associated with Coccidioides immitis infection were identified at postmortem examination, and were limited to the lower respiratory tract.

Hyperhydration prior to moderate-intensity exercise causes arterial hypoxaemia.

The second day of a 3-day event is the most physically demanding of the 3 days. If this is performed under hot and humid environmental conditions, detrimental effects on cardiovascular and thermoregulatory function and, therefore, on exercise capacity, may occur due to exercise-induced dehydration. We hypothesised that the administration of fluid equivalent to 6% of the horse's bodyweight prior to a simulated second day of a 3-day event would increase plasma volume and limit increases in core temperature. Seven Standardbred geldings underwent a training protocol prior to the study. A standardised exercise test was developed for each horse so that exercise intensity at each phase would be the same percentage of the maximal heart rate for all horses. The exercise test involved 4 phases: Phase A involved 30 min exercise at 3.7 m/s (approximately 25% VO2max); Phase B 4 min exercise at 8 m/s (approximately 60% VO2max); and Phase C 50 min at 3.7 m/s, after which there was a 10 min rest. Phase D involved 14 min at 7.3 m/s (55% VO2max). In a cross-over design, horses were grouped randomly and allocated to either exercise with no fluid (control) or approximately 26 l isotonic fluid by nasogastric tube, 120 min prior to exercise. Arterial and mixed venous blood samples were collected prior to exercise, towards the end of each of the phases and during the rest period. The administration of fluid prior to exercise resulted in a pre-exercise bodyweight gain of 21.3 +/- 1.2 kg. Hyperhydration resulted in a greater degree of arterial hypoxaemia than the control group in Phases B and D, but not in Phases A and C or at rest. During Phases B and D, mean PaO2 values in the horses that received fluid were about 15 torr lower than in the control group, but there were no differences in PaCO2 values between the 2 groups. In both arterial and mixed venous blood, pH and HCO3- were significantly lower in the group that were hyperhydrated. We concluded that the most likely cause of the more severe arterial hypoxaemia in the hyperhydrated group during the intense exercise phase was some degree of pulmonary oedema, from the extravasation of the administered fluid. Hyperhydration prior to exercise may be detrimental to respiratory function and therefore care must be taken in administration of large volumes of fluid prior to exercise.

Cytochrome P450 isozymes involved in the metabolism of phenol, a benzene metabolite.

Benzene is an occupational and environmental toxicant. The major health concern for humans is acute myelogenous leukemia. To exert its toxic effects, benzene must be metabolized by cytochrome P450 to phenol and subsequently to catechol and hydroquinone. Previous research has implicated CYP2E1 in the metabolism of phenol. In this study the cytochrome P450 isozymes involved in the metabolism of phenol were examined in hepatic and pulmonary microsomes utilizing chemical inhibitors of CYP2E1, CYP2B, and CYP2F2 and using CYP2E1 knockout mice. CYP2E1 was found to be responsible for only approximately 50% of 20 microM phenol metabolism in the liver. This suggests another isozyme(s) is involved in hepatic phenol metabolism. In pulmonary microsomes both CYP2E1 and CYP2F2 were significantly involved.

Hepatic and pulmonary microsomal benzene metabolism in CYP2E1 knockout mice.

Benzene is an occupational and environmental toxicant. The major health concern for humans is acute myelogenous leukemia. To exert its toxic effects, benzene must be metabolized via cytochrome P450. CYP2E1 has been identified as the most important cytochrome, P450 isozyme in hepatic benzene metabolism in mice, rats, and humans. In pulmonary microsomes CYP2E1 and members of the CYP2F subfamily are both significantly involved. In the current study CYP2E1 knockout mice and wild-type controls were used to further examine the cytochrome P450 isozymes involved in metabolism of 24 microM benzene. The results show that CYP2E1 is the most important isozyme in the liver, accounting for 96% of the total hydroxylated metabolite formation. However, in the lung CYP2E1 was responsible for only 45% of the formation of total hydroxylated metabolite. Chemical inhibitors of CYP2E1 and CYP2F2 were used to further examine the contributions of these isozymes to benzene metabolism. The results confirmed the finding that while CYP2E1 is the most important isozyme in the liver, CYP2F2 and CYP2E1 are both significantly involved in the lung.

Metabolism of the styrene metabolite 4-vinylphenol by rat and mouse liver and lung.

Styrene is a widely used chemical in the reinforced plastics industry and in polystyrene production. Its primary metabolic pathway to styrene oxide and then to styrene glycol, which is further metabolized to mandelic acid and phenylglyoxylic acid, has been well studied. However, a few studies have reported finding a minor metabolite, 4-vinylphenol (4-VP), in rat and human urine. The present studies sought to determine if the formation and metabolism of 4-VP in rat and mouse liver and lung preparations could be measured. When styrene was incubated with hepatic and pulmonary microsomal preparations, 4-VP formation could not be measured in these preparations. However, considerable 4-VP metabolizing activity, as determined by the loss of 4-VP, was observed in both mouse and rat liver and lung microsomal preparations. 4-Vinylphenol metabolizing activity in mouse liver microsomes was three times greater than that in rat liver microsomes, and activity in mouse lung microsomes was eight times greater than that in rat lung microsomes. This activity was completely absent in the absence of NADPH. Studies with cytochrome P-450 inhibitors indicated the involvement of CYP2E1 and CYP2F2. Induction of CYP2E1 by pyridine resulted in an increase in 4-VP metabolism by mouse hepatic microsomes but not by pulmonary microsomes. The metabolite(s) formed as a result of this oxidative pathway remain to be identified. In additional studies, glutathione conjugation appeared to be involved in 4-VP metabolism with the highest activity being in mouse lung, with or without the addition of NADPH.

Renal tubular acidosis in horses (1980-1999).

Renal tubular acidosis (RTA) is characterized by altered renal tubular function resulting in hyperchloremic metabolic acidosis. The purpose of the study was to describe RTA in 16 horses. No breed or sex predilection was found. The mean age at onset of the disease was 7 years of age. The type of diet had no apparent effect on development of RTA. The most common clinical signs were depression, poor performance, weight loss, and anorexia. Initial blood work revealed a marked hyperchloremic metabolic acidosis in all horses and a compensatory respiratory response in most horses. Sixty-three percent (10/16) of the horses had some evidence of renal damage or disease. Initial treatment consisted of large amounts of sodium bicarbonate given intravenously and orally for the prompt correction of the acidosis. Response to treatment was largely dependent on the rate of sodium bicarbonate administration. Long-term oral supplementation with NaHCO3 was required for the maintenance of normal acid-base status in individual horses. Recurrence of RTA was noted in 56% (9/16) of the horses. Horses with evidence of renal disease had multiple relapses. RTA should be considered as a differential diagnosis in horses with vague signs of depression, weight loss, and anorexia. The pathogenesis of RTA in horses remains uncertain, but prompt recognition and early aggressive intravenous sodium bicarbonate therapy followed by long-term oral supplementation seem to be important to successful management.

Cytochromes P450 involved with benzene metabolism in hepatic and pulmonary microsomes.

Benzene is an occupational hazard and environmental toxicant found in cigarette smoke, gasoline, and the chemical industry. The major health concern associated with benzene exposure is leukemia. The toxic effects of benzene are dependent on its metabolism by the cytochrome P450 enzyme system. Previous research has identified CYP2E1 as the primary P450 isozyme responsible for benzene metabolism at low concentrations, whereas CYP2B1 is involved at higher concentrations. Our studies using microsomal preparations from human, mouse, and rat indicate that CYP2E1 is the P450 isozyme primarily responsible for benzene metabolism in lung and in liver. CYP2B isozymes have little involvement in benzene metabolism by either lung or liver. Our results also indicate that isozymes of the CYP2F subfamily may play a role in benzene metabolism by lung.

Metabolism of styrene by human liver and lung.

In mice, styrene is pneumotoxic, and there is some evidence of tumorigenicity. This toxicity is thought to be related to its bioactivation to styrene oxide in lung. To determine if human tissues have this capacity, the metabolism of styrene to styrene oxide was measured in human liver and lung microsomal preparations. Hepatic microsomes metabolized styrene to styrene oxide, but lung microsomes had essentially no activity. However, microsomes from both tissues metabolized benzene to phenol. The data suggest that human lung has low styrene metabolizing activity and may be much less of a target organ than in mouse.

Metabolism of styrene oxide to styrene glycol in enriched mouse clara-cell preparations.

Styrene is a widely used chemical that has been shown to cause lung tumors in mice but not in rats. Styrene toxicity appears to be related to its bioactivation to styrene oxide, and this occurs almost exclusively in Clara cells. An important pathway in the detoxification of styrene oxide is via epoxide hydrolase to yield styrene glycol. When mouse Clara cells were incubated with racemic styrene oxide, R-styrene glycol was the predominant metabolite, giving an R/S ratio of 3.6. When the pure styrene oxide enantiomers were used as substrates, the corresponding styrene glycols were the predominant but not exclusive metabolites. Activity was slightly higher with the S-styrene oxide than with the R-styrene oxide. Addition of reduced glutathione to the incubation medium resulted in an increase in epoxide hydrolase activity, perhaps by decreasing oxidative stress. Mouse Clara cells thus show the capacity for detoxifying styrene oxide.

Metabolism of styrene by mouse and rat isolated lung cells.

Styrene is pneumotoxic in mice. It is metabolized by pulmonary microsomes of both mouse and rat to styrene oxide (SO), presumed to be the toxic metabolite of styrene, and known to be genotoxic. To determine which pulmonary cell types are responsible for styrene metabolism, and which cytochromes P450 are associated with the bioactivation of styrene, we isolated enriched fractions of mouse and rat Clara and type II cells in order to determine the rate of styrene metabolism, with and without chemical inhibitors. Mouse Clara cells readily metabolized styrene to SO. Diethyldithiocarbamate, a CYP2E1 inhibitor, caused less inhibition of SO formation in Clara cells isolated from mice than previously found with pulmonary microsomes. As in microsomes, 5-phenyl-1-pentyne, a CYP2F2 inhibitor, inhibited the formation of both enantiomers. alpha-Naphthoflavone, a CYP1A inhibitor, did not inhibit SO formation in Clara cells. alpha-Methylbenzylaminobenzotriazole, a CYP2B inhibitor, exhibited minimal inhibition of SO production at 10 microM and less at 1 microM. The microsomal and isolated cell studies indicate that CYP2E1 and CYP2F2 are the primary cytochromes P450 involved in pulmonary styrene metabolism. Styrene metabolizing activity was much greater in Clara cells than in type II pneumocytes, which demonstrated essentially no activity. Styrene-metabolizing activity was several-fold higher in the mouse than in rat Clara cells. The more pneumotoxic and genotoxic form, R-SO, was preferentially formed in mice, and S-SO was preferentially formed in rats. These findings indicate the importance of Clara cells in styrene metabolism and suggest that differences in metabolism may be responsible for the greater susceptibility of the mouse to styrene-induced toxicity.

Species comparison of hepatic and pulmonary metabolism of benzene.

Benzene is an occupational hazard and environmental toxicant found in cigarette smoke, gasoline, and the chemical industry. The major health concern associated with benzene exposure is leukemia. Studies using microsomal preparations from human, mouse, rabbit, and rat to determine species differences in the metabolism of benzene to phenol, hydroquinone and catechol, indicate that the rat is most similar, both quantitatively and qualitatively, to the human in pulmonary microsomal metabolism of benzene. With hepatic microsomes, rat is most similar to human in metabolite formation at the two lower concentrations examined (24 and 200 microM), while at the two higher concentrations (700 and 1000 microM) mouse is most similar in phenol formation. In all species, the enzyme system responsible for benzene metabolism approached saturation in hepatic microsomes but not in pulmonary microsomes. In pulmonary microsomes from mouse, rat, and human, phenol appeared to competitively inhibit benzene metabolism resulting in a greater proportion of phenol being converted to hydroquinone when the benzene concentration increased. The opposite effect was seen in hepatic microsomes. These findings support the hypothesis that the lung plays an important role in benzene metabolism, and therefore, toxicity.

Effects of frusemide on electrolyte and acid-base balance during exercise.

This study was undertaken to evaluate the effects of frusemide on the concentration of plasma electrolytes and the relationship between changes in electrolyte concentration and the simultaneous changes in acid-base balance in arterial and venous blood during intense exercise. Five exercise-conditioned Thoroughbred horses were exercised on a high-speed treadmill at a slope of 10% at speeds known to exceed VO2max. Horses participated in 3 randomised exercise trials in which they received either placebo (control), low-dose frusemide (0.5 mg/kg bwt), or high-dose frusemide (1.0 mg/kg) 4 h prior to exercise. Arterial and mixed venous blood samples were drawn before, during and after intense exercise. Packed cell volume (PCV), plasma protein, plasma electrolyte, plasma lactate concentrations and acid-base balance were determined. Frusemide at both dosages induced a significant metabolic alkalosis associated with a decrease in plasma çhloride[ in both arterial and mixed venous blood. There was a highly significant correlation between the strong ion difference (SID) and bicarbonate[ in all blood samples. The effects of frusemide on electrolyte concentration and acid-base balance were largely due to changes in SID.

Weak acid-concentration Atot and dissociation constant Ka of plasma proteins in racehorses.

The plasma proteins are a significant contributor to the total weak acid concentration as a net anionic charge. Due to potential species difference, species-specific values must be confirmed for the weak acid anionic concentrations of proteins (Atot) and the effective dissociation constant for plasma weak acids (Ka). We studied the net anion load Atot of equine plasma protein in 10 clinically healthy mature Standardbred horses. A multi-step titration procedure, using a tonometer covering a titration range of PCO2 from 25 to 145 mmHg at 37 degrees C, was applied on the plasma of these 10 horses. Blood gases (pH, PCO2) and electrolytes required to calculate the strong ion difference ([SID] = [(Na(+) + K(+) + Ca(2+) + Mg(2+))-(Cl(-) + Lac(-) + PO4(2-))]) were simultaneously measured over a physiological pH range from 6.90-7.55. A nonlinear regression iteration to determine Atot and Ka was performed using polygonal regression curve fitting applied to the electrical neutrality equation of the physico-chemical system. The average anion-load Atot for plasma protein of 10 Standardbred horses was 14.89 +/- 0.8 mEq/l plasma and Ka was 2.11 +/- 0.50 x 10(-7) Eq/l (pKa = 6.67). The derived conversion factor (iterated Atot concentration/average plasma protein concentration) for calculation of Atot in plasma is 0.21 mEq/g protein (protein-unit: g/l). This value compares closely with the 0.24 mEq/g protein determined by titration of Van Slyke et al. (1928) and 0.22 mEq/g protein recently published by Constable (1997) for horse plasma. The Ka value compares closely with the value experimentally determined by Constable in 1997 (2.22 x 10(7) Eq/l). Linear regression of a set of experimental data from 5 Thoroughbred horses on a treadmill exercise test, showed excellent correlation with the regression lines not different from identity for the calculated and measured variables pH, HCO3 and SID. Knowledge of Atot and Ka for the horse is useful especially in exercise studies and in clinical conditions to quantify the mechanisms of the acid-base disturbances occurring.

Effects of inhibitors of CYP1A and CYP2B on styrene metabolism in mouse liver and lung microsomes.

Much of the toxicity of styrene is associated with its bioactivation to styrene oxide. Both liver and lung have been shown to carry out this metabolic step, but there are differences reported as to which isomers of cytochrome P450 are responsible for this biotransformation in various species and tissues. CYP2E1, CYP2F, CYP2B, CYP1A1/2 and CYP2C11 have all been implicated. In the current study, alpha-naphthoflavone (alphaNF) and alpha-methylbenzylaminobenzotriazole (MBA), selective inhibitors of CYP1A and CYP2B, were used to ascertain the contributions of these isomers to styrene metabolism in mouse hepatic and pulmonary microsomes. AlphaNF did not inhibit styrene metabolism with microsomal preparations from either tissue. This indicates that CYP1A is unimportant in the metabolism of styrene to styrene oxide. MBA at a very low concentration of 1 microM inhibited the hepatic metabolism of benzyloxyresorufin (a CYP2B substrate) by 87% but caused only a 16 to 19% inhibition of R- and S-styrene oxide formation. This demonstrates that CYP2B plays a minor role in styrene metabolism. At 10 microM, MBA caused an even greater inhibition of styrene metabolism but at that level it also inhibited p-nitrophenol hydroxylation, a CYP2E1-dependent reaction, suggesting a loss of selectivity for this inhibitor at higher concentrations.

Effects of concentrated electrolytes administered via a paste on fluid, electrolyte, and acid base balance in horses.

OBJECTIVES: To test effectiveness of an electrolyte paste in correcting fluid, electrolyte and acid base alterations in response to furosemide administration. ANIMALS: 6 Standardbreds. PROCEDURES: Horses received electrolyte paste or water only (control). The paste was given orally 3 hours after furosemide administration (1 mg/kg of body weight, IM). Water was given ad libitum soon after the paste and 3 hours after furosemide administration to treated and control groups, respectively. Paste Na+, K+, and Cl- composition was approximately 2,220, 620, and 2,840 mmol, respectively. The PCV and plasma concentrations of total protein ([TP]), [Na+], [K+], [Cl-]), and bicarbonate ([HCO3-]) were determined, and urinary fluid and electrolyte excretion, fecal water, and body weight changes were measured. RESULTS: At the end of a 6-hour period, the paste-treated group had higher water consumption, which resulted in lower plasma [TP]; net electrolyte losses also were substantially less. With paste administration, [Na+] was approximately 2 mmol/L above a prefurosemide value of 137.3 mmol/L; control horses had values similar to the prefurosemide value. Plasma [Cl-] remained at the prefurosemide value, but values in control horses decreased by 7 mmol/L with water consumption. Plasma [K+] remained approximately 0.8 mmol/L below prefurosemide values in both groups. Venous [HCO3-] returned to prefurosemide values after paste administration, but alkalosis persisted in control horses after consumption of water only. Body weight loss was less after paste administration. CONCLUSIONS: Administration of electrolyte paste is advantageous over water alone in restoring fluid, electrolyte, and acid base balance after fluid and electrolyte loss attributable to furosemide administration.

Minimal effects of acrylonitrile on pulmonary and hepatic cell injury enzymes in rats with induced cytochrome P450.

Acrylonitrile (AN) has many industrial applications but is a known carcinogen in animals and a suspect human carcinogen. Its toxicity is generally associated with its bioactivation, the initial step of which is epoxidation by cytochrome P450. While the hepatotoxicity and pneumotoxicity of AN in naive rats is generally low, the purpose of this study was to investigate the pneumotoxicity and hepatotoxicity of AN in adult male Sprague-Dawley rats and evaluate interactions with agents that may alter its metabolism. Five agents, phenobarbital, beta-naphthoflavone, pyridine, ethanol, and acetone, were administered prior to AN as inducers of CYP2B, CYP1A, and CYP2E1. Pneumotoxicity was measured as increases in y-glutamyltranspeptidase (GGT) and lactate dehydrogenase (LDH) in bronchoalveolar lavage fluid (BALF). Hepatotoxicity was measured as increases in serum sorbitol dehydrogenase (SDH). AN (1 mmol/kg ip) had little effect on liver or lung, even when given following most of the inducing agents. AN (1.5 mmol/kg) caused an increase in GGT, but had little effect on SDH or LDH. Acetone plus AN caused an increase in mortality and some indication of pneumotoxicity, but lung and liver were histologically normal. Thus AN alone even at a high dose had no effect on the liver or lung and minimal effects following induction of cytochrome P450 by acetone.

Electrolyte disturbances in foals with severe rhabdomyolysis.

Marked electrolyte abnormalities characterized by profound hyperkalemia, hyponatremia, hypocalcemia, and hyperphosphatemia were noted in 4 neonatal foals with acute rhabdomyolysis and pigmenturia. In 2 foals, rhabdomyolysis developed 4-6 days after admission for dysmaturity, and in 2 foals, rhabdomyolysis was evident on presentation. Rhabdomyolysis was a consequence of selenium deficiency with or without vitamin E deficiency, possibly combined with increased oxidant stress due to sepsis or hypoxia and reperfusion injury after parturition. Foals gained from 7 to 15% of their initial body weight within 48 hours of developing rhabdomyolysis. Three of the foals developed cardiac arrhythmias characterized by spiked T waves and decreased-amplitude P waves. Postmortem examination of 2 foals revealed extensive myodegeneration and renal tubular nephrosis; renal cortical necrosis with myocardial necrosis was noted in 1 foal. Destruction of the major intracellular compartment (intracellular fluid [ICF]) through extensive myonecrosis combined, in some cases, with myoglobinuric renal insufficiency produced major fluid shifts and life-threatening electrolyte derangements. With the major ICF compartment disrupted, hyperkalemia was most effectively treated using mineralocorticoids, loop diuretics, and ion exchange resins to enhance elimination. In addition, i.v. calcium, glucose, insulin, and sodium bicarbonate were administered, which helped redistribute potassium to the ICF. Severe rhabdomyolysis should be included in the differential diagnoses of hyperkalemia, hyponatremia, hypocalcemia, and hyperphosphatemia in neonatal foals.