Association between plasma vitamin E concentration and the risk of equine motor neuron disease

Equine motor neuron disease (EMND) is a neurodegenerative disorder of the somatic lower motorneurons that results in a syndrome of diffuse neuromuscular disease in the adult horse. The aetiology of this disorder is unknown, although prior studies have suggested that a deficiency in the lipid antioxidant vitamin E (α-tocopherol) contributes to the development of EMND. This paper describes a case-control study designed to investigate the association between plasma vitamin E levels and the risk of EMND for horses. Signalment, plasma vitamin E levels at the time of referral, and information relative to dietary and management practices were collected from 53 horses diagnosed with EMND and 69 controls. The mean plasma vitamin E concentration in EMND cases was significantly lower than that of control horses. After controlling for other risk factors of EMND, there was a statistically significant association between plasma vitamin E levels and EMND, with the likelihood of the disease increasing as the vitamin E concentration decreased. These findings support the reported role of vitamin E deficiency as one of the risk factors for EMND.     

Sarcocystis myositis and vitamin E deficiency in a Gypsy Vanner stallion suspected of having equine motor neuron disease

A 3-year-old Gypsy Vanner stallion was presented for evaluation of intermittent recumbency, muscle fasciculations, weakness, low head carriage, shifting of weight between the hindlimbs and an elevated tail head. History, physical examination and serum alpha tocopherol concentrations were suggestive of vitamin E deficiency and equine motor neuron disease (EMND). Sacrocaudalis dorsalis medialis muscle biopsy identified myositis secondary to sarcocystosis. Treatment with alpha tocopherol, ponazuril and sulfadiazine/pyrimethamine resulted in significant improvement in muscle weakness and body condition with resolution of sarcocystosis and inflammation on repeat muscle biopsy. This case illustrates the importance of muscle biopsy in horses with neuromuscular disease as concurrent diseases may be present that require specific treatment for a positive outcome.     

Evaluation of the oral vitamin E absorption test in horses

An oral vitamin E absorption test used in human beings was modified for use in horses. The most appropriate techniques with which to measure gastrointestinal tract absorption of vitamin E (alpha-tocopherol) in horses were developed. Vitamin E was administered orally, and serum values of alpha-tocopherol were measured by use of high-performance liquid chromatography at 0, 3, 6, 9, 12, and 24 hours after vitamin E administration. Variables included comparison of 2 dosages (45 and 90 IU/kg of body weight), routes of administration, and absorption dynamics of 3 preparations of dl-alpha-tocopherol. Absorption of the 2 doses of dl-alpha-tocopherol acetate indicated a dose response; the area under the curve at 24 hours (AUC24) was 4.3 micrograms.h/ml for the 45-IU/kg dose and 32.2 micrograms.h/ml (P less than 0.01) for the 90-IU/kg dose. Maximal absorption was apparent when vitamin E was naturally consumed in grain, compared with administration of identical preparations by stomach tube or paste. In the same horses, dl-alpha-tocopherol and dl-alpha-tocopherol acetate plus polyethylene glycol had statistically similar absorption curves and both had significantly greater AUC24, compared with dl-alpha-tocopherol acetate; values for the 3 compounds were 23.6, 25.8, and 12.6 micrograms.h/ml, respectively. The AUC24 varied between individual horses, but time of peak value was consistently observed between 6 and 9 hours.(ABSTRACT TRUNCATED AT 250 WORDS)     

Evaluation of glucose metabolism in three horses with lower motor neuron degeneration

OBJECTIVES: To determine whether increased glucose metabolism is the potential cause of the decreased plasma glucose curve determined after oral glucose tolerance testing in horses with lower motor neuron degeneration. ANIMALS: 3 horses with signs suggestive of lower motor neuron degeneration, 1 horse with malignant melanoma with multiple metastases, and an obese but otherwise healthy horse. Procedures-Glucose metabolism was assessed by use of the hyperglycemic clamp and euglycemic hyperinsulinemic clamp techniques. RESULTS: Mean rate of glucose metabolism of horses with lower motor neuron degeneration was significantly greater (mean, 3.7 times greater than control horses; range, 2.1 to 4.8 times greater) than that reported in 5 healthy control horses (41 +/- 13 micromol/kg/min vs 11 +/- 4.5 micromol/kg/min, respectively). In addition, one of the affected horses, an 8-year-old warmblood gelding, had a 5.6-times increased sensitivity to exogenously administered insulin, compared with that reported in 5 healthy control horses. Pancreatic insulin secretion was not insufficient in horses with lower motor neuron degeneration. Findings in the 2 diseased control horses were unremarkable. CONCLUSIONS AND CLINICAL RELEVANCE: Increased glucose metabolism in horses with lower motor neuron degeneration may be the cause of the decreased plasma glucose curve detected after oral glucose tolerance testing. This finding could aid in developing supportive treatments with respect to adequate glucose and vitamin E supplementation.     

Evaluation of glucose tolerance and intestinal luminal membrane glucose transporter function in horses with equine motor neuron disease

OBJECTIVE: To confirm whether the plasma glucose concentration curve obtained during oral glucose tolerance tests (OGTTs) in horses with equine motor neuron disease (EMND) is decreased, compared with that obtained in clinically normal horses, and determine whether that decrease is a result of defective glucose metabolism or intestinal glucose transport dysfunction. ANIMALS: 8 horses with EMND and 44 matched control horses. PROCEDURE: Electromyography and OGTTs were performed in all 8 affected horses and 10 control horses. Intravenous GTTs (IVGTTs) were performed in 6 affected horses and another 11 control horses. The activity and levels of jejunal luminal membrane glucose transporter (Na+ / glucose cotransporter isoform 1 [SGLT1]) were measured in 2 affected horses and 23 control horses. RESULTS: In horses with EMND, generalized neuropathy was detected via quantitative electromyography; the mean increase in plasma glucose concentration during the OGTT was significantly decreased, compared with the value in control horses. During the IVGTT the mean increase in plasma glucose concentration was significantly lower than that of control horses. The activity and levels of SGLT1 in 2 affected horses were similar to those of control horses. Diagnosis of EMND was confirmed postmortem in all affected horses. CONCLUSIONS AND CLINICAL RELEVANCE: Data suggest that the decreased plasma glucose curve obtained in horses with EMND during OGTTs (compared with control horses) is a result of overall enhanced glucose metabolism or abnormalities in the facilitated glucose transporters; definitive identification of the underlying mechanisms could aid in the development of appropriate treatments of EMND in horses.     

Changes in lipoprotein composition in horses fed a fat-supplemented diet

This study was designed to compare the effects of a fat-supplemented diet versus a traditional diet on the lipoprotein (LP) content in horses. In the first of two trials, eight two-year old horses were fed a basal diet, in which 80% of the digestible energy was supplied as chopped alfalfa hay. One group of four horses (fat-supplemented group, FS) was fed the remaining 20% of the digestible energy as corn oil, while the other group of four horses (control) was fed rolled corn to complete their diet. Blood samples were collected at the start of the experiment and every 2.5 weeks thereafter for 10 weeks. Total serum lipids were measured in both groups of horses and the lipoproteins were fractionated into very low density LP (VLDL), low density LP (LDL), and high density LP (HDL) using ultracentrifugation and agarose-column chromatography. Each LP fraction was measured for protein, cholesterol (CH) and triglyceride TG) content. Total serum lipids were increased in the FS group above the control (9.16 vs. 4.65 mg/ml, week 5). Serum cholesterol and triglyceride concentrations were elevated in the FS group, but were highly variable. Variations in lipid concentrations may have been due to variation in time of sampling during the day. In the second trial, four of the eight horses were used and divided into the same two groups; FS vs. control. After an initial sample, postprandial serial blood samples (1 hour intervals for 8 hours) were drawn at 2, 4 and 6 weeks. Upon examination of the data, Hour 3 post-feeding was chosen to represent postprandial LP values. In both trials, the horses were able to adapt to the added dietary fat. Maintenance of body weight and increased speed of lipid clearance from the blood by the end of each trial in the FS group support this statement. In the FS group, there was an increase in VLDL TG concentration, but not in LDL. This indicates an increase in VLDL TG clearance from the circulation, presumably by increased lipoprotein lipase (LPL) activity. Cholesterol concentrations were also increased in the FS horses in the LDL and HDL. The rise in cholesterol may be attributed to endogenous recycling of liver products such as bile salts, which aid in digestion and absorption and are cholesterol based. In addition, there were greater quantities of LDL and HOL produced in the FS horses as supported by the increased protein concentrations as well as larger peaks for the eluate from the gel filtration column.     

Serum zinc, iron and copper status during early gestation in sows fed a folic acid-supplemented diet

The purpose of this trial was to determine whether an addition of folic acid to a commercial diet would affect serum Zn, Fe and Cu status in sows between weaning and 30 d of gestation. At weaning, 162 sows were assigned randomly to six groups and housed in individual cages fitted on a slatted floor. There were six treatments according to a 2 X 3 factorial arrangement: two levels of supplementary folic acid (0 and 5 mg/kg of diet) and three treatments to stimulate ovulation (none, flushing and pregnant mare serum gonadotropin [PMSG] i.m. injection). Control groups were fed a commercial-type diet, and folic acid-treated groups were fed the same diet supplemented with 5 mg/kg of pteroylglutamic acid. All sows were mated twice within 7 d after weaning. Of the 162 animals originally selected, 123 sows were pregnant and used in this trial. Serum folates, Zn, Cu and Fe were measured at weaning, mating and 30 d of gestation. Serum Cu, Zn and folates increased between weaning and mating, and then decreased to 30 d of gestation. Supplementing the commercial diet with folic acid elevated serum folates between weaning and d 30 of gestation (P less than .001). Folic acid supplementation also was associated with a higher level of serum Zn at 30 d of gestation. Supplemental folic acid had no effect on the pattern of serum Cu and Fe throughout the experiment.(ABSTRACT TRUNCATED AT 250 WORDS)     

Concentrations of trace minerals in the spinal cord of horses with equine motor neuron disease

OBJECTIVE: To compare concentrations of trace minerals in the spinal cord of horses with equine motor neuron disease (EMND) with those of horses without neurologic disease (control horses). ANIMALS: 24 horses with EMND and 22 control horses. PROCEDURE: Spinal cord trace mineral concentrations in horses with EMND and control horses were analyzed by use of inductively coupled plasma atomic emission spectroscopy (calcium, phosphorus, sodium, potassium, magnesium, copper, iron, manganese, nickel, zinc, aluminum, cobalt, and chromium), atomic absorption spectrophotometry (lead and cadmium), flameless atomic absorption (mercury), and fluorometry (selenium). RESULTS: Copper concentration was significantly higher in the spinal cord of horses with EMND, compared with control horses; spinal cord concentrations of all other trace minerals were similar between groups. CONCLUSION AND CLINICAL RELEVANCE: Among spinal cord trace minerals investigated in the study, only copper concentrations were significantly different between horses with EMND and horses without neurologic disease, which suggests that copper may be involved in the pathogenesis of EMND. An hypothesis of oxidative injury in this disease is supported by the finding of increased copper concentrations in the spinal cord and by low vitamin E concentrations reported by other researchers.     

Trichinella in horses: a low frequency infection with high human risk

After the initial report in 1976 of a trichinellosis epidemic caused by the consumption of infected horsemeat, 12 other outbreaks have been described in Europe. Since the first serious human outbreak several experiments have confirmed the susceptibility of horses to Trichinella species and the rapid disappearance of specific antibodies in this host that prevents the use of serological methods for routine screening. A review of the distribution of parasite burdens in muscles of naturally or experimentally infected horses indicates that the tongue is the most likely sample to contain detectable numbers of Trichinella larvae in low level infections. Requirements for testing of horsemeat are specified in legislation of the European Union, and other recommendations are published elsewhere. The EEC directives have evolved into very specific requirements which specify the testing of at least 5g of tongue, masseter or diaphragm per horse using a pooled digestion assay. More recently, France has revised the requirement for sample size to 10g for horsemeat originating from countries with high prevalence of Trichinella. To address the continuing outbreaks of human trichinellosis due to infected horsemeat, the development and implementation of a quality assurance system for testing is being considered.     

Storage and mobilization of muscle glycogen in exercising horses fed a fat-supplemented diet

In a switchback experiment, six mature mares were fed a control and a fat-supplemented diet while being exercised in a galloping regimen. After three weeks adaptation to each diet, horses performed an exercise test (ET) consisting of four, 600-m gallops. Muscle biopsies were obtained before and after the ET, and blood samples were taken before, during and throughout recovery from the ET. Resting glycogen concentration in the biceps femoris muscle increased (P<.05) from 15.77 mg/g wet tissue when the horses were fed the control diet to 22.89 mg/g when they were fed the fats-supplemented diet. During the ET, the amount of glycogen mobilized by the muscle increased (P<.05) from 6.99 mg/g when the horses were fed the control diet to 13.09 mg/g when they were fed the fat-supplemented diet. When the horses were fed the fat-supplemented diet, they galloped faster (P<.09), at a constant heart rate, during the last two gallops of the ET. Thus, adapting exercising horses to a fat-supplemented diet increased muscle glycogen concentrations, which appeared to enhance their performance past the anaerobic threshold.     

Calcium metabolism of pregnant rats fed a vitamin D-depleted diet

The effects of vitamin D (VD) deficiency on calcium (Ca) metabolism during pregnancy were evaluated in rats fed VD-repleted (VD-repleted rats) and VD-depleted (VD-depleted rats) diets. In the VD-depleted rats, the plasma concentrations of 1,25-dihydroxyvitamin D and Ca decreased severely, whereas the parathyroid hormone concentrations increased. The Ca contents of the feces of the VD-depleted rats were higher than those of the VD-repleted rats. The fetal Ca contents of the VD-repleted and VD-depleted rats increased continuously, but that of the VD-depleted rats was lower. These data reveal that VD deficiency during pregnancy induces severe hypocalcemia due to reduced intestinal absorption of Ca and elevated fetal demand for Ca.     

Change in blood antioxidant status of horses moved from a stable following diagnosis of equine motor neuron disease

The antioxidant status of 10 horses living in stable 1 where 2 cases of equine motor neuron disease had previously been diagnosed was assessed before and 9 weeks after moving to another stable. Duration of residence in stable 1, subsequent moving, or both, significantly affected several parameters of the antioxidant status.     

Relative bioavailability of organic bis-glycinate bound copper relative to inorganic copper sulfate in beef steers fed a high antagonist growing diet

To assess the relative bioavailability of bis-glycinate bound Cu, 90 Angus-cross steers (265 ± 21 kg) were blocked by body weight (BW) to pens with GrowSafe bunks and randomly assigned to dietary treatments (14 to 18 steers/treatment): 0 mg supplemental Cu/kg dry matter (DM; CON), 5 or 10 mg supplemental Cu/kg DM as Cu sulfate (CS5; CS10) or bis-glycinate bound Cu (GLY5; GLY10). Steers received a high antagonist growing diet (analyzed 4.9 mg Cu/kg DM, 0.48% S, and 5.3 mg Mo/kg DM). Steers were weighed at the beginning (days 1 and 0) and end (days 125 and 126) of the trial to determine average daily gain (ADG) and gain:feed (G:F). Blood was collected from all steers on days 0, 28, 56, 84, and 126. Liver samples were collected on days −3 or −2 and day 123 or 124. Data were analyzed using ProcMixed of SAS (experimental unit = steer; fixed effect = treatment and block). Plasma Cu was analyzed as repeated measures (repeated effect = day). Plasma and liver Cu concentrations were regressed against total Cu intake using ProcGLM to calculate relative bioavailability of GLY. Final BW and overall ADG were greatest for CS5 and CS10 and least for CON and GLY5 (P = 0.01). Overall, DMI was not affected by treatment (P = 0.14), but overall G:F tended to be greatest for CS5, CS10, and GLY5 and least for CON (P = 0.08). Total and supplemental Cu intake was greatest for steers supplemented either source at 10 mg Cu/kg DM and least for CON (P < 0.01). However, total and supplemental Cu intake was greater for CS5 than GLY5 (P < 0.01). Final liver Cu concentrations were greatest for CS10, least for CON, CS5, and CS10, and intermediate for GLY10 (P < 0.01). Final plasma Cu was greatest for steers supplemented either source at 10 mg Cu/kg DM (P < 0.01). Relative bioavailability of GLY was 82% compared to CS based on liver Cu (P < 0.01) but did not differ based on plasma Cu (P = 0.60). The lesser bioavailability of GLY relative to CS could be due to a high concentration of dietary antagonists and lower solubility of GLY (68.9% relative to CS) in pH conditions (5.2) similar to the ruminal pH of a beef animal consuming a high concentrate diet. Future studies should examine the effects of bis-glycinate bound Cu fed in blended combination with inorganic Cu sulfate to determine the most effective blend of sources for feedlot cattle experiencing varying amounts of dietary Cu antagonists.     

Antioxidative and oxidative status in muscles of pigs fed rapeseed oil, vitamin E, and copper

The susceptibility of a given muscle tissue to lipid oxidation may not only depend on the presence of unsaturated fatty acids and the balance between antioxidants and prooxidants, but also on the composition of the skeletal muscle. In the present study, the effects of dietary supplementation of vitamin E (dl-alpha-tocopheryl acetate) and copper in combination with a high level of monounsaturated fatty acids were examined with regard to the antioxidant concentration and the susceptibility to lipid oxidation of two muscles, longissimus (LD) and psoas major (PM), representing different oxidative capacity. In addition, fatty acid profiles of the backfat and the intramuscular lipids, as well as fresh meat quality traits, were studied. Pigs were allotted to a 3x3 factorial experiment with three levels of dl-alpha-tocopheryl acetate (0, 100, and 200 mg/kg of feed) and three levels of copper (0, 35, and 175 mg/kg of feed) added to a diet containing 6% rapeseed oil. A basal diet (without rapeseed oil) was added to the experimental design, giving a total of 10 dietary treatments. Muscle alpha-tocopherol concentrations increased (P<.001) with increasing dl-alpha-tocopheryl acetate in the feed. The antioxidative status was higher in PM than in LD, when considering the concentration of alpha-tocopherol (P<.001) and the activity of antioxidant enzymes (superoxide dismutase, P<.001; glutathione peroxidase, P = .06). Supplemental copper did not give rise to any deposition of copper in muscle tissue or backfat, but the antioxidant status of PM increased. The susceptibility to lipid oxidation was reduced in LD with increasing dietary dl-alpha-tocopheryl acetate and in PM with increasing dietary copper. Supplemental dl-alpha-tocopherol acetate improved the water-holding capacity of LD (P = .005) and PM (P = .003). The fatty acid composition of the backfat and the triglyceride fraction of the intramuscular fat became more unsaturated with the addition of rapeseed oil to the feed. Higher intakes of monounsaturated fatty acids due to the rapeseed oil were also reflected in the phospholipid fraction of the intramuscular fat, but no influence on the proportion of saturated fatty acids was seen. The susceptibility to lipid oxidation of PM was lower for pigs on the rapeseed oil-based diet than for those on the basal diet. The energy metabolic status of the muscles and the accumulation of calcium by the sarcoplasmic reticulum were not influenced by the dietary treatments, but there were differences between muscle types. The addition of rapeseed oil to the diet reduced the muscular content of glycogen (LD, P = .02; PM, P = .06) and elevated the plasma concentration of free fatty acids (P = .05). Overall, dietary fat, dl-alpha-tocopherol acetate, and copper affected the oxidative status of pig muscles, and the results differed depending on muscle type.     

Effect of a preparation of Saccharomyces cerevisiae on microbial profiles and fermentation patterns in the large intestine of horses fed a high fiber or a high starch diet

Eight horses were allotted into pairs consisting of one cecum- and right ventral colon-fistulated animal and one cecum-fistulated animal. They were fed daily at the same level of intake either a high-fiber (HF) or a high-starch (HS) diet without or with 10 g of a Saccharomyces cerevisiae preparation, in a 4 x 4 Latin square design. The HS diet provided a starch overload (i.e., 3.4 g starch x kg(-1) BW x meal(-1)) while maintaining a high amount of fiber intake (i.e., dietary NDF/starch ratio was 1.0). A 21-d period of adaptation to the treatments occurred before cecal and colonic contents were withdrawn 4 h after the morning meal to count total anaerobic, cellulolytic, and lactic acid-utilizing bacteria, lactobacilli, and streptococci. Lactic acid, volatile fatty acids, ammonia concentrations, and pH were measured on cecal and colonic fluid samples collected hourly during the first 12-h postfeeding. When the HS diet was fed, the concentration of total anaerobic and lactic acid-utilizing bacteria increased (P < 0.001), whereas that of cellulolytic bacteria decreased (P < 0.05) in the cecum. The concentration of lactobacilli and streptococci increased (P < 0.001) in the cecal and colonic contents. These alterations of the microbial profiles were associated with decreases (P < 0.001) of pH, (acetate + butyrate)/propionate ratio and with an increase (P < 0.001) of lactic acid concentration. Supplementing the S. cerevisiae preparation increased (P < 0.01) the concentration of viable yeast cells, averaging 4.3 x 10(6) and 4.5 x 10(4) cfu/mL in the cecal and colonic contents, respectively. Yeast supplementation had almost no effect on microbial counts in the cecum and colon. The supplementation of S. cerevisiae appeared to modify (P < 0.05) pH, concentrations of lactic acid and ammonia, molar percentages of acetate and butyrate with the HS diet and [(acetate + butyrate)/propionate] ratio when the HF diet was fed. The effects of the S. cerevisiae preparation were greater in the cecum than in the colon, which coincided with the abundance of yeast cells. When the digestion of starch in the small intestine was saturated, the effect of the addition of a S. cerevisiae preparation appeared to limit the extent of undesirable changes in the intestinal ecosystem of the horse.     

Cellular immune responses in pigs fed a vitamin E- and selenium-deficient diet.

The effects of dietary restriction of vitamin E (Vit E) and selenium (Se) on lymphocyte proliferation, natural killer (NK) cell activity, antibody-dependent cell-mediated cytotoxicity (ADCC), and on burst respiratory response of stimulated granulocytes as measured by chemiluminescence (CL) were studied in pigs. Six male weanling pigs were maintained for 25 d on a torula yeast-based diet containing no measurable amount of alpha-tocopherol and less than .02 mg of Se per kilogram of feed. Six others received the same basal diet supplemented with 33 IU of DL-alpha-tocopheryl acetate and .2 mg of Se per kilogram of feed. All pigs were inoculated with Salmonella typhisuis on d 21 of the feeding period and killed on d 25. Tests to measure cellular immune functions were performed on cells isolated from blood samples taken on d 21 and 25. After 21 d of feeding, lymphocyte blastogenesis responses to phytohemagglutinin, concanavalin A, and pokeweed mitogen in pigs fed the Vit E- and Se-deficient diet were normal compared with the response in pigs fed the supplemented diet. Moreover, the cytotoxic activity of NK cells, the ADCC response, and the CL response of granulocytes were not affected. After 25 d, a marked suppression of lymphocyte response to mitogens occurred in pigs fed the Vit E- and Se-deficient diet when the cells were cultured in the presence of autologous serum. When fetal bovine serum replaced autologous serum in the cultures, no suppression was observed. No effect on NK activity and ADCC was observed, whereas the CL peak response of granulocytes tended to be higher in pigs fed the deficient diet.(ABSTRACT TRUNCATED AT 250 WORDS)