Lowered Antioxidant Status of Red Blood Cells in Post-Parturient Haemoglobinuria of Buffaloes

The antioxidant status of the red blood cells of buffaloes (n = 20) suffering from post-parturient haemoglobinuria (PPH) was assessed by comparing their tocopherol (vitamin E) and reduced glutathione contents with those of red blood cells from apparently healthy buffaloes (n = 20). The red cell tocopherol content of the diseased buffaloes (1.76±0.11 g/ml) was significantly (p<0.01) lower than that of healthy buffaloes (2.45±0.14 g/ml). This may be the first report comparing the concentration of tocopherol in the red blood cells of buffaloes suffering from PPH and apparently healthy buffaloes.There was a drastic reduction in the reduced glutathione content in the red cells of haemoglobinuric buffaloes (23.74±2.86 mg%) compared to the healthy control buffaloes (73.71±3.87 mg%). The diseased buffaloes also exhibited severe hypophosphataemia. These findings suggest that an impaired or insufficient antioxidant potential of the red blood cells in this disease in buffaloes is associated with the phosphorus deficiency.     

Haemoglobinuria and hypophosphataemia in postparturient dairy cows without dietary deficiency of phosphorus

Postparturient haemoglobinuria occurred in 11 Holstein-Friesian cows in 8 dairy herds in East Gippsland, Victoria. The affected cows had a wide range of ages, body condition and production level. Seven of 9 cows sampled were hypophosphataemic, 4 of 8 cows examined had Heinz bodies present in erythrocytes and strong ketonuria was present in 6 of 10 urine samples tested. Herdmates having similar calving dates had mostly normal serum phosphorus levels and none had Heinz bodies. There was no evidence for dietary phosphorus (P) deficiency, hypocuprosis or excessive P drain from high production being predisposing factors. There were no cruciferous plants in the diet. Haemolysis persisted in 4 cows despite treatment to raise serum P levels. An hyposthesis is proposed for the sporadic nature and clinical manifestations of the disease. This is based on mechanisms of hypophosphataemia occurring in man and mechanisms of Heinz body formation and haemolysis in other circumstances. We suggest that hypophosphataemia occurs following recovery from preparturient keto-acidosis caused by undernutrition in late pregnancy and then becoming nutritionally replete in the early postparturient period. Circulating oxidants may cause erythrocyte damage which may be predisposed to by hypophosphataemia.     

Evaluation Of An Additive Solution For Preservation Of Canine Red Blood Cells

The effect of an additive preservative solution on canine red blood cell posttransfusion viability (PTV) and on selected canine red blood cell biochemical parameters was studied. One unit (450 mL) of blood was collected from 6 clinically normal dogs into the anticoagulant citrate phosphate dextrose, centrifuged, and the plasma removed. The red blood cells were then suspended in 100 mL of a saline, adenine, dextrose, and mannitol solution and stored at 4°C. Aliquots were removed for study at 1, 10, 20, 30, 37, and 44 days. The 24-hour PTV of autologous red blood cells was determined using a sodium chromate (⁶¹Cr) label. Red blood cell concentrations of 2,3-diphosphoglycerate (2,3-DPG), adenosine-5'-triphosphate (ATP), and pH were also determined. Canine red blood cell PTV, pH, ATP, and 2,3-DPG concentrations decreased during storage ( P < .05). The PTV decreased from 94% using day 1 red blood cells to 80% and 75% using day 37 and day 44 red blood cells, respectively ( P < .05). Although the mean PTV of the day 44 stored units equaled the Food and Drug Administration (FDA) minimum standard for human red blood cells, the PTV was substandard in 75% of the day 44 units. The FDA standard was exceeded in 83% of the day 37 units. It was concluded that 37-day-old canine red blood cells preserved with a saline, adenine, dextrose, and mannitol solution are of acceptable quality for transfusion.     

Energy metabolism in canine erythrocytes associated with inherited high Na+- and K+-stimulated adenosine triphosphatase activity

Energy metabolism in canine erythrocytes associated with inherited high Na+- and K+-stimulated adenosine triphosphatase [(Na,K)-ATPase] activity (HK cells) was compared with that in normal canine erythrocytes (LK cells). Activities of some of the glycolytic enzymes in the HK cells were significantly higher than those in LK cells. The concentrations of adenosine triphosphate (ATP) and glycolytic intermediates in HK cells were almost equal to those in LK cells. Glucose utilization and lactate production by HK cells in vitro and incorporation of [32P]orthophosphate or [14C]glucose into 2,3-diphosphoglycerate in HK cells were higher than in LK cells. Radioactivity of [32P]ATP in HK cells was lower than in LK cells, but increased to approximately that of LK cells when (Na,K)-ATPase of HK cells was completely blocked by ouabain. When HK cells and LK cells were incubated in the absence of glucose, the concentration of ATP in HK cells was decreased more than that of LK cells. Although ouabain reduced the rate of decrease in ATP in HK cells, the decrease in ATP in HK cells was still 2-fold that in LK cells. The half-life of HK cells was about one-half that of LK cells. The results indicated that glycolysis is greater in HK cells than in LK cells, and that the increased glycolysis in HK cells was stimulated by an increased rate of ATP breakdown for active cation transport by the (Na,K)-ATPase and by increased degradation of ATP for some other pathway, eg, glutathione synthesis. Thus, the increased demand for ATP in HK cells might result in shortening the lifespan of HK erythrocytes.     

In vitro studies of haemolysis by Leptospira interrogans serovars pomona and ballum

Washed and unwashed red blood cells (RBC) from young calves, adult cattle, hamsters and humans were incubated with Leptospira interrogans serovars pomona and ballum. Washed cells suspended in saline were always haemolysed while unwashed cells and those which were washed and resuspended in plasma were never haemolysed, despite the presence of large numbers of organisms within the culture supernatant. Pomona produced greater haemolysis of cattle and human RBC than did ballum, but with hamster RBC ballum produced greater haemolysis than did pomona. A group of 6- to 9-month-old cattle infected with pomona showed no signs of clinical disease and RBC taken from them before infection and during the development of antibodies to pomona were haemolysed by pomona only after the cells were washed. Plasma therefore appears to have a protective function. This in vitro protective function of plasma even extended to plasma from young seronegative calves.     

低磷奶牛血清某些生化指标变化

为进一步研究低磷引起血红蛋白尿症的发病机理,对低磷奶牛血清生化指标进行了检测。采用分组对比的方法,测定各组奶牛血清若干生化指标。结果,血尿组,低磷组,对照组血清无机磷含量差异显著。     

Enhanced Lipid Peroxide Levels in the Erythrocytes of Calves with Haemoglobinuria

Eight 6–9 month old calves, showing clinical signs of intermittent haemoglobinuria, even after treatment with an antipiroplasmal drug (4,4-diamidinodiazoaminobenzene diaceturate), were examined for oxidative damage to their erythrocytes and the presence of hemoprotozoa in blood smears. Four calves without signs of haemoglobinuria served as controls. The blood smears from three of the eight calves contained piroplasms for Theileria annulata. Irrespective of the presence of piroplasms in their blood smears, the calves with haemoglobinuria had significantly (p<0.01) lower haemoglobin concentrations (Hb) and packed cell volumes (PCV). The lipid peroxide level in the erythrocytes, but not in the plasma, of calves with red urine was significantly (p<0.05) higher than that for the controls. It is concluded that haemoglobinuria, irrespective of the presence of piroplasms in blood smears, is associated with oxidative stress to erythrocytes and peroxidation of polyunsaturated fatty acids of cell membrane.     

Effect of storage on oxygen dissociation of canine blood.

Oxygen dissociation curve and adenosine triphosphate (ATP) and 2,3-diphosphoglycerate (DPG) contents in red blood cells (RBC) were determined in canine blood stored in acid citrate dextrose (ACD) and citrate phosphate dextrose (CPD) solutions. The oxygen-unloading ability decreased, as shown by the left shift of oxygen dissociation curve during storage, and the shift correlated with decreasing DPG but not decreasing ATP concentrations. After 2 weeks of storage in ACD solution, oxygen dissociation curves were shifted significantly to the left. For blood stored in CPD solution, 4 weeks was required before the shift was significant. It was concluded that canine blood collected and stored in CPD solution is more efficient than that stored in ACD solution in delivering oxygen to the tissues.     

Red urine from red deer grazed on pure red clover swards

Twenty-four red deer hinds with their calves were released on to a newly established pure red clover sward and, 2 days later, red staining of the tail, perineum and hocks was observed. This was presumed to be of urinary origin. Observation of micturition showed that when urine was passed, it was a normal straw colour but it turned scarlet-red about 1 hour after exposure to air. Midstream urine remained the normal colour when held under a pure nitrogen atmosphere immediately after micturition, but it turned red when held in air in the dark, suggesting that the colour change was due to an oxidative rather than a photosensitive reaction. All deer grazing red clover were affected but this did not occur in deer grazing ryegrass/white clover swards. No adverse effects were observed in the deer grazing the red clover, and calf growth was significantly higher than on ryegrass/white clover, suggesting that the red urine had no effect on health or productivity. Blood and urine analyses showed no signs of haemolysis, haematuria or haemoglobinuria. Preliminary chemical analyses suggest that the compounds involved are not those found in the urine of sheep grazing oestrogenic clover. The nature of the compounds have yet to be determined.     

Red urine from red deer grazed on pure red clover swards

Twenty-four red deer hinds with their calves were released on to a newly established pure red clover sward and, 2 days later, red staining of the tail, perineum and hocks was observed. This was presumed to be of urinary origin. Observation of micturition showed that when urine was passed, it was a normal straw colour but it turned scarlet-red about 1 hour after exposure to air. Midstream urine remained the normal colour when held under a pure nitrogen atmosphere immediately after micturition, but it turned red when held in air in the dark, suggesting that the colour change was due to an oxidative rather than a photosensitive reaction. All deer grazing red clover were affected but this did not occur in deer grazing ryegrass/white clover swards. No adverse effects were observed in the deer grazing the red clover, and calf growth was significantly higher than on ryegrass/white clover, suggesting that the red urine had no effect on health or productivity.

Blood and urine analyses showed no signs of haemolysis, haematuria or haemoglobinuria. Preliminary chemical analyses suggest that the compounds involved are not those found in the urine of sheep grazing oestrogenic clover. The nature of the compounds have yet to be determined.     

Influence of dietary mineral imbalance on the incidence of urolithiasis in Egyptian calves

The influence of certain dietary elements on the urolithiasis syndrome in cattle calves was elucidated. Calcium, phosphorus, and magnesium measurements were conducted on feed rations as well as on serum and urine samples collected from affected and normal calves. Analysis of the rations given to the animals showed phosphorus at higher levels than calcium, indicating mineral imbalance. Serum and urine of urolithic calves were characterised by high phosphorus, calcium, magnesium, urea, and creatinine levels. Physical examination of urine of affected animals showed a high degree of turbidity, a large amount of calcium carbonate, and triple phosphate as well as abundant amount of pus cells and red blood cells. The characteristic clinical symptoms of urine retention were observed. Moreover, some animals were found to urinate through an opening in front of the scrotal region.     

Identification of ticks and detection of blood protozoa in friesian cattle by polmerase chain reacton test and estimation of blood parameters in district Kasur,Pakistan

The study was carried out to detect Theileria annulata, the causative agent of theileriosis, and Babesia bovis, the causative agent for babesiosis, in Friesian cattle by PCR and conventional blood smear examination. One hundred blood samples obtained from diseased Friesian cattle kept on private livestock farms at Pattoki, District Kasur, Pakistan were collected in addition to 20 blood samples obtained from non-diseased animals. The disease manifestations observed clinically included high fever, swelling of sub mandibular and sub scapular lymph nodes, weakness, increased respiration and pulse, anorexia, loss of condition and rough hair coat. Neurologic sign of in coordination was also seen in weak animals. Signs of lacrimation, pale conjunctiva, diarrhoea, dyspnea and frothy nasal discharge were observed in only one animal. Clinically nine animals showed signs of haemoglobinuria. Diagnosis of bovine theileria and babesia species was based on finding many intraerythrocytic piroplasms of both blood protozoa with clinical signs associated with anaemia, lymph node hyperplasia and haemoglobinuria. One hundred samples of ticks were also collected for identification of vector. Results showed that the prevalence of Hyalomma tick was highest (15%) followed by Boophilus (12%), Haemaphysalis (5%) and Rhipicephalus (3%). The blood smear examination showed 21% (21/100) samples positive for blood parasites out of which 66.6% (14/ 21) samples were positive for theileriosis while 42.8% (9/21) were positive for babesiosis. It was also recorded that 66.66% (6/9) samples were positive for B.bigemina while 33.33% (3/9) were positive for B.bovis. The results showed that 60% (60/100) samples were positive for blood parasites by PCR test. Out of these 60% (36/60) were positive for T.annulata while 33.33% (20/60) were positive for babesia. The specificity and sensitivity of PCR test was higher than blood smear examination. The blood parameters in haemoparasites infection were also analyzed and the results showed significant decrease in total erythrocyte count and haemoglobin while MCV, MCH values increased and MCHC was slightly less than normal indicating macrocytic hypochromic anaemia.     

Severe hypophosphataemia associated with the management of hyperlipaemia in a miniature pony

This report describes a case of severe hypophosphataemia associated with the management of hyperlipaemia in a miniature pony following colic surgery. Clinical signs attributed to hypophosphataemia included obtundation, anorexia, tachycardia, tachypnoea and generalised muscle fasciculations. Hyperlipaemia was managed with enteral and partial parenteral nutrition; insulin was also administered to control hyperglycaemia after the initiation of caloric support. Specific therapy for hypophosphataemia consisted of parenteral potassium phosphate at 0.03 mmol/kg bwt/h (i.v.). The pony made a full recovery without further complications. Hypophosphataemia may be an under‐recognised clinical problem in certain populations of critically ill equids, such as those with hyperlipaemia and receiving insulin as part of their management. The routine measurement of phosphate concentration in these cases is recommended.     

当归对鸡血液指标的影响研究

为探讨当归对鸡血液指标的影响,试验用高、中、低3个剂量的当归煎剂分别给正常和贫血鸡进行灌服,测定试验鸡的红细胞数、血红蛋白含量、红细胞压积和血沉值,并与正常及贫血不用药组进行比较.贫血鸡模型利用环磷酰胺制造.结果发现:当归能显著提高正常鸡的红细胞数、血红蛋白含量、红细胞压积和血沉值;同时当归能够逆转环磷酰胺所致鸡的红细胞数、血红蛋白含量、红细胞压积和血沉值,使其恢复至正常水平.     

Efficacy of oral calcium and/or sodium phosphate in the prevention of parturient paresis in cows

The goal of this study was to investigate the efficacy of calcium chloride, sodium phosphate or a combination of these two substances administered orally immediately postpartum for the prevention of parturient paresis in cows. Thirty-two cows that had had parturient paresis at the previous calving, and in which serum biochemistry had shown hypocalcaemia and hypophosphataemia, were used in the study. The cows were transferred to the Department of Farm Animals, University of Zurich, five days before their expected due dates. On a randomized trial, the cows were given calcium chloride, sodium phosphate, both substances or no treatment (controls) via a stomach tube immediately postpartum and 12 hours later. The cows were monitored for 96 hours during which time blood was collected on a regular basis for the determination of total calcium, ionized calcium, inorganic phosphorus and magnesium concentrations. Of the 32 cows treated, 19 (59%) had parturient paresis and 13 (41%) did not. The incidence of parturient paresis did not differ significantly among the groups although there was a tendency for a lower incidence in cows treated with both calcium chloride and sodium phosphate. The various treatments had no apparent effect on serum calcium concentration. The concentration of inorganic phosphorus increased significantly in cows treated with sodium phosphate compared with the controls. The results of this study showed that cows treated with both calcium chloride and sodium phosphate orally tended to have a lower incidence of parturient paresis. Further investigation into multiple administrations of oral calcium chloride and sodium phosphate, started before parturition, for the prevention of parturient paresis is required.     

Modulation of Glycolysis and the Pentose Phosphate Pathway Influences Porcine Oocyte In Vitro Maturation

Glycolytic and pentose phosphate pathway (PPP) activities were modulated in porcine cumulus–oocyte complexes (COCs) during in vitro maturation (IVM) by the addition of inhibitors or stimulators of key enzymes of the pathways to elucidate their relative participation in oocyte maturation. The activities of glycolysis and PPP were evaluated by lactate production per COC and by the brilliant cresyl blue test, respectively. Glucose uptake per COC and the oocyte maturation rate were also evaluated. Lactate production, glucose uptake and the percentage of oocytes reaching metaphase II decreased in a dose‐dependent manner in the presence of the pharmacological (NaF) or the physiological (ATP) inhibitors of glycolysis (p < 0.05). The addition of the physiological stimulator of glycolysis (AMP) caused no effect on lactate production, glucose uptake or the meiotic maturation rate. The pharmacological (6‐AN) and the physiological (NADPH) inhibitors of PPP induced a dose‐dependent decrease in the percentage of oocytes with high PPP activity and in the nuclear maturation rate (p < 0.05). The physiological stimulator of PPP (NADP) caused no effect on the percentage of oocytes with high PPP activity. The glycolytic and PPP activities of porcine COCs and maturational competence of oocytes seem to be closely related events. This study shows for the first time the regulatory effect of ATP and NADPH as physiological inhibitors of glycolysis and PPP in porcine COCs, respectively. Besides, these pathways seem to reach their maximum activities in porcine COCs during IVM because no further increases were achieved by the presence of AMP or NADP.     

Clinicopathological changes and effect of imidocarb therapy in dogs experimentally infected with Babesia canis

In this study one spleen-intact dog (A) and two splenectomised dogs (BSE, CSE) were infected with Babesia canis. All animals developed an acute disease characterised by fever, haemoglobinuria and anaemia, the latter being more severe in the splenectomised dogs. Fever and parasitised red blood cells were detected for three days after imidocarb treatment in the splenectomised animals. Haematological abnormalities included regenerative anaemia, thrombocytopenia and leukopenia (due to neutropenia and lymphopenia) in the acute phase, soon followed by leukocytosis, neutrophilia and left shift a few days later. Acute hepatopathy was detected in all dogs with elevated ALT activity, which was more seriously altered in the splenectomised dogs. Diffuse changes in liver structure and hepatomegaly were seen by ultrasonography. Liver biopsy and histology revealed acute, non-purulent hepatitis in the splenectomised dogs. Both splenectomised dogs were successfully cured after collection of 400 ml highly parasitised blood, proving that large-amount antigen production is possible with rescuing the experimental animals. Whole blood transfusion, imidocarb and supportive care with infusions, antipyretics, glucocorticoids and diuretics were applied. The spleen-intact dog clinically recovered after receiving supportive treatment, with no imidocarb therapy. Microbial infections developed in both splenectomised animals (BSE: haemobartonellosis, CSE: osteomyelitis caused by Escherichia coli), probably as a consequence of immunosuppression after splenectomy and glucocorticoid therapy.     

Evaluation of Canine Red Blood Cells Stored in a Saline, Adenine, and Glucose Solution for 35 Days

An additive solution for the storage of red blood cells was evaluated for use in dogs. Blood collected from 6 dogs was processed into packed red blood cells and stored for 35 days in the additive solution Nutricel (Miles, Inc, Pharmaceutical Division, West Haven, CT). Packed red blood cells stored in citrate-phosphate-dextrose-adenine (CPDA-1; Fenwal Laboratories, Baxter Health Care Corp, Deerfield, IL) also were evaluated for comparison. Red blood cell 2,3-diphosphoglycerate (2,3-DPG) concentration, adenosine triphosphate (ATP) concentration, percentage hemolysis, and pH were determined. The red blood cell post-transfusion viability (PTV) after 35 days of storage was assessed with both single-labeled chromium 51 (⁵¹Cr) and double-labeled technetium 99m/chromium 51 (^99mTc/⁵¹Cr) techniques. Mean ATP concentration and percentage hemolysis of the cells stored in Nutricel were 1.1 μmol/g hemoglobin (Hb) and 0.28% respectively and did not differ significantly (P < .05) from the values of 1.0 μmol/g Hb and 0.33% from the CPDA-1-stored red blood cells. The mean pH of red blood cells stored in Nutricel was 6.19, which was significantly lower than the pH of 6.47 for cells stored in CPDA-1. The mean 2,3-DPG concentration of red blood cells stored in Nutricel was significantly higher at 10.1 μmol/g Hb than the 2,3-DPG concentration of 3.4μmol/g Hb for cells stored in CPDA-1. The mean PTV of canine red blood cells stored in Nutricel for 35 days was 85% with ⁵¹Cr and 90% with ^99mTc/⁵¹Cr. This was significantly higher than the mean PTVs of 38% and 36% for the CPDA-1 stored cells as assessed with ⁵¹Cr and ^99mTc/⁵¹Cr techniques, respectively. It was concluded that 35-day-old canine red blood cells stored in Nutricel are of acceptable quality for transfusion purposes.     

Effect of mitochondrial uncoupling and glycolysis inhibition on ram sperm functionality

Studies have demonstrated the importance of mitochondria to sperm functionality, as the main source of ATP for cellular homoeostasis and motility. However, the role of mitochondria on sperm metabolism is still controversial. Studies indicate that, for some species, glycolysis may be the main mechanism for sperm energy production. For ram sperm, such pathway is not clear. Thus, we evaluated ram sperm in response to mitochondrial uncoupling and glycolysis inhibition aiming to assess the importance of each pathway for sperm functionality. Statistical analysis was performed by the SAS System for Windows, using the General Linear Model Procedure. Data were tested for residue normality and variance homogeneity. A p < .05 was considered significant. Groups treated with the mitochondrial uncoupler Carbonyl cyanide 3 chlorophenylhydrazone (CCCP) showed a decrease in the percentage of cells with low mitochondrial activity and high mitochondrial membrane potential. We also observed that the highest CCCP concentration promotes a decrease in sperm susceptibility to lipid peroxidation. Regardless the lack of effect of CCCP on total motility, this substance induced significant alterations on sperm kinetics. Besides the interference of CCCP on spermatic movement patterns, it was also possible to observe such an effect in samples treated with the inhibitor of glycolysis (2‐deoxy‐d ‐glucose, DOG). Furthermore, treatment with DOG also led to a dose‐dependent increase in sperm susceptibility to lipid peroxidation. Based on our results, we suggest that the glycolysis appears to be as important as oxidative phosphorylation for ovine sperm kinetics as this mechanism is capable of maintaining full motility when most of the cells have a low mitochondrial membrane potential. Furthermore, we found that changes in the glycolytic pathway trough glycolysis inhibition are likely involved in mitochondrial dysfunction and sperm oxidative unbalance.     

The halothane gene, energy metabolism, adenosine monophosphate-activated protein kinase, and glycolysis in postmortem pig longissimus dorsi muscle

The presence of the halothane gene results in PSE meat. However, the exact mechanisms linking the halothane gene and the incidence of PSE meat remain unclear. We hypothesize that the presence of the halothane gene accelerates energy consumption in postmortem muscle, which activates adenosine monophosphate-activated protein kinase (AMPK), leading to enhanced glycolysis and PSE meat. To test our hypothesis, energy status, AMPK activity, and glycolysis in the postmortem LM of the halothane gene carrier and halothane-negative pigs were compared. The results showed that the presence of the halothane gene accelerated energy depletion in postmortem muscle immediately after exsanguination, leading to rapid and early depletion of ATP, as shown by an increase in the (adenosine monophosphate + inosine monophosphate):ATP ratio in postmortem LM. In addition, an early AMPK activation was observed in LM from halothane carriers. The fructose-2,6-diphosphate concentration in postmortem LM was well correlated with AMPK activation. To be a potent stimulator of phosphofructose kinase, the increase in fructose-2,6-diphosphate is expected to activate phosphofructose kinase, a key enzyme controlling glycolysis, leading to enhanced glycolysis and early accumulation of lactic acid. In summary, this study showed that the presence of the halothane gene induced early energy depletion, which could be a primary reason causing AMPK activation, leading to accelerated glycolysis and an increased incidence of PSE meat. However, AMPK might also be activated by other mechanisms besides energy depletion, which warrants further studies.