Characterization of differentiation antigens expressed in bovine lymphosarcomas.

To characterize the cell-surface antigens expressed in tumour cells derived from bovine leukosis and to determine their cell lineages, the immunophenotypes of the tumour cells from 13 bovine lymphosarcomas were examined with 13 monoclonal antibodies (mAbs). Of 13 cattle with lymphosarcomas, four were identified clinically as having the thymic-type sporadic bovine leukosis (SBL) and one as having the skin-type; two had enzootic bovine leukosis (EBL) and six were untypable. Flow cytometric analysis revealed that the tumour cells from nine cases were of T-cell lineage (BoCD5+, BoCD2+ or BoCD2-) and two were of B-cell lineage (MHC-II+, BoCD5+, IgM+); there were two bovine leukaemia virus-infected cattle (EBL). T-cell tumours appeared to originate from immature (BoCD4-, BoCD8-) T cells, but there was no significant relationship between clinical type (EBL, calf-, skin- and thymic-type) and tumour-cell immunophenotype.

Effects of vitamin B2 on neutrophil functions in cattle.

Vitamin B2 was intramuscularly administered to Holstein cattle and the ensuing changes in peripheral blood neutrophil function were investigated. The neutrophil count displayed a significant increase at 1-2 days after administration, while nitroblue tetrazolium reducing activity and phagocytic bactericidal activity were enhanced at 1-4 days after administration in calves and at 1-6 days after administration in adult cows. The increases in the neutrophil count and the activation of neutrophil functions were observed, being manifested at dosages of 10 mg/kg or greater for calves and 5 mg/kg or greater for adult cows.

Bovine herpesvirus meningoencephalitis association with infectious bovine rhinotracheitis (IBR) vaccine.

During 1992, on a farm in the Tokachi district of Hokkaido, Japan, approximately 20 Holstein-Friesian calves showed neuroparalysis and died within 7-10 days after routine vaccination. Six male calves, aged about 1.5 months, were submitted to our laboratory for pathological examination and diagnosed as acute or subacute necrotizing meningoencephalitis due to bovine herpes virus (BHV) infection. The main necropsy findings included a few hemorrhages or clots, and malacic lesions localized in the cortical to subcortical area of the cerebrum. Histopathological brain lesions were characterized by laminar or focal necrosis of neurons, accompanying macrophages, polymorphonuclear cell infiltration, severe astrogliosis, and perivascular cuffing in all six calves. Nuclear basophilic inclusion bodies, which showed positive reaction with immunocytochemical staining of BHV antigen, were observed in the necrotic neurons, astroglia and oligodendroglia in five affected calves. BHV antigens were also seen in the cell bodies and cell processes of the necrotic neurons, which was indicative of cell-to-cell propagation of infection. There was a general tendency for more severe lesions to be located at the cortex to subcortex of the cerebrum. Milder lesions were observed in the cerebellum and brain stem. These findings suggest that the infectious route to the cerebrum in the present cases was through the olfactory bulbs and/or along the meninges beginning from the ethmoid bone, rather than through the trigeminal ganglia route as had been emphasized in studies dealing with experimental infection.

Hereditary myopathy of the diaphragmatic muscles in Holstein-Friesian cattle.

We describe a family line with an autosomal recessive disease of muscular dystrophy of the diaphragmatic muscles in Holstein-Friesian cattle. Histopathological examination in the present cases revealed various degenerative changes in the diaphragmatic and other thoracic muscles as follows: variation in muscle fiber diameter, fiber splitting, sarcoplasmic masses, ring fiber, vacuolar and hyalinized degeneration of muscle fibers. In addition, central core-like structures were the prominent features in the diaphragmatic muscles, occupying the center of the fiber or scattered within the fiber. These pathological alterations are consistent with the diaphragmatic myopathy previously reported in Meuse-Rhine-Yssel cattle in the Netherlands. The fibers containing core-like structures consisted of three distinct zones which could be well distinguished by NADH-tetrazolium reductase activity. This activity was absent in the innermost zone, decreased in the intermediate zone, and normal or increased in the periphery. Electron microscopically, this structure appeared to be composed of focal myofibrillar degeneration beginning with streaming or disintegration of the Z disk. We discuss here the similarity between this core-like structure and the other alternative organelles that have been reported previously, and a possible defect or storage in the cytoskeleton from the findings of the Z disk abnormalities.

Muscular dystrophy of the diaphragmatic muscles in Holstein-Friesian cows.

Six Holstein-Friesian cows suffering from recurrent rumenal tympany were pathologically investigated. Macroscopical lesions associated with the clinical symptoms were confined to the diaphragmatic muscles which were pale, and stiff on palpation. Histopathological examination revealed various degenerative changes in diaphragmatic muscles as follows: variation in muscle fiber diameter, vacuolar and hyalinized degeneration of muscle fibers, fiber splitting, central core-like structures, sarcoplasmic masses and ring fibers. These characteristic features in the present cases were consistent with dystrophy of the diaphragmatic muscles in Meuse-Rhine-Yssel cattle. From these observations, it is confirmed that muscular dystrophy of the diaphragmatic muscles dose occur in Holstein-Friesian cows, although a genetic mode was not proven.

Clinicopathological observations on thoroughbred foals with enlarged thyroid gland.

Seven cases of thoroughbred foals kept on a farm in the Hidaka district of Hokkaido showed bilateral thyroid enlargement from 3 or 4 months after birth. Their serum T4 levels were remarkably low, whereas T3 levels were in the normal range. Serum T4 levels were also low in the other clinically normal foals and their dams kept together. Since the iodine content in pasture soil and grass was very low, iodine deficiency was supposed to be the cause of the disease.

Effects of active egg white product on neutrophil function in calves.

The effects of an active egg white product (AEWP) on neutrophil function in calves were investigated. Calves were administered AEWP orally at doses of 250 mg and 500 mg/kg either once or twice, with an interval of 5 days between doses in the latter case. The peripheral blood neutrophils of calves receiving a single 500 mg/kg dose displayed increased nitroblue tetrazolium reducing activity and increased intracellular killing of Staphylococcus aureus from 1 day after administration, with maximum levels being attained on the 3rd day. However, no such increase of these activities was observed after administration of 250 mg/kg. Calves receiving two doses of 500 mg/kg displayed the same changes as seen in the corresponding one-dose group, while the neutrophil activity of calves receiving 250 mg/kg also increased after the second dose. However, no increase in the peripheral blood neutrophil count was observed after AEWP administration. Thus, AEWP enhanced the nonspecific antibacterial activity of neutrophils when given to calves by the oral route.

Spontaneous diabetes mellitus in young cattle: histologic, immunohistochemical, and electron microscopic studies of the islets of Langerhans.

Pathomorphologic studies were carried out on three cases of bovine diabetes mellitus with clinical signs of polydipsia, polyuria, severe emaciation, glycosuria, persistent hyperglycemia, and decreased glucose tolerance. At necropsy, two animals had atrophy of the pancreas, whereas other visceral organs, including the endocrine organs, showed no significant changes. Microscopically, there was atrophy and reduced numbers of pancreatic islets accompanied by interlobular and interacinar fibrosis and compensatory enlargement of some remaining islets. Lymphocytes were observed commonly around and within atrophic islets and occasionally around and within enlarged islets. Vacuolar degeneration with occasional accumulation of glycogen granules was observed in the beta-cells of these enlarged islets. Immunohistochemical studies of atrophic islets demonstrated complete loss of beta-cells or only a few small beta-cells. There also was a corresponding decrease in the number of cells that stained with anti-glucagon (alpha-cells) or anti-somatostatin (delta-cells) antibodies. The vacuolated cells in the enlarged islets stained strongly with anti-insulin antibody (beta-cells). Ultrastructurally, the majority of cells in the atrophic islets had reduced cytoplasmic volume and few secretory granules, features consistent with alpha-cells. In contrast, enlarged islets that had prominent immunohistochemical staining for insulin (beta-cells) consisted of beta-cells with cytosolic edema, mitochondrial swelling, dilated smooth endoplasmic reticulum, and reduced numbers of or degranulated secretory granules. These pathomorphologic features found in cattle are similar to those found in juvenile-onset insulin-dependent diabetes mellitus in human beings and suggest autoimmune involvement in diabetes.

Bovine undifferentiated alveolar rhabdomyosarcoma and its differentiation in xenotransplanted tumors.

At necropsy, a 7-year-old Holstein cow, clinically diagnosed via rectal palpation as having malignant abdominal neoplasia was found to have many metastatic tumors of various sizes in the abdominal and thoracic cavities and in the liver, lung, kidney, adrenal, uterus, and lymph nodes. These tumors were examined by histologic and immunohistochemical methods using anti-human myoglobin, anti-bovine myoglobin, and anti-desmin sera. Tumors were diagnosed as undifferentiated alveolar rhabdomyosarcomas. The tumors in the abdominal and thoracic cavities differed histologically from the metastases in organs. The former tumors consisted of only undifferentiated cells, most of which did not stain positively for desmin or myoglobin. The metastatic tumors contained a few rhabdomyoblastic cells that stained positively for desmin and myoglobin. Tumors tissues from the cow were transplanted and propagated through six passages in athymic nude mice. After one passage, the transplanted tumor histologically resembled those from the abdominal serosa and consisted of cells with scanty cytoplasm with an alveolar arrangement. Cells from this tumor stained positively for desmin and negatively for myoglobin. After two passages through nude mice, tumor cells were of two distinct histologic types: those cells with scanty cytoplasm and elongated cells with plentiful eosinophilic cytoplasm. Both of these cells stained positively for both desmin and myoglobin. After six passages, cross striations were detected in neoplastic cells by electron microscopy. These findings illustrate that a highly undifferentiated bovine rhabdomyosarcoma, in which most of the cells in the original tumor lacked desmin and myoglobin, became better differentiated and stained positively for desmin and myoglobin after serial transplantation in nude mice.

Efficacy of colostral immunoglobulins for therapeutic and preventive treatments of calf diarrhea.

Pooled colostral milk samples were collected from apparently healthy Holstein cows. Immunoglobulin (Ig) was partially purified from the colostral milk and concentrated by freeze drying. The Ig powders had neutralizing antibody titers of 1:6,250 to 1:31,250 against bovine rotavirus (RV), and reacted with four RV proteins of VP2, VP4, VP6, and VP7. No therapeutic effect of Ig powder showed on clinically serious diarrhea with RV in dairy and beef calves. However, the Ig powder had a preventive effect on the diarrhea in beef calves when it was given soon after calving.

Studies on serum tocopherol and selenium levels and blood glutathione peroxidase activities in lambs with white muscle disease.

For the investigation of the cause of white muscle disease (WMD), tocopherol (Toc) and selenium (Se) levels and blood glutathione peroxidase (GSH-Px) activities were examined using lambs with WMD and their ewes. Serum Se levels of 4 lambs with WMD were low under 30 ppb, lambs showing very low levels below 15 ppb. The serum Se level was correlated with blood GSH-Px activity showing remarkably low activities in the lambs with WMD. Se contents in the organs of lambs with WMD were lower than those of control lambs, and particularly liver Se contents were deficient levels below 50 ppb. Serum Toc levels were normal, but alpha-Toc contents in organs showed very low levels, especially in the liver. The serum Toc and Se levels and blood GSH-Px activities of their ewes and other sheep kept in the same farm revealed similar results to those of lambs with WMD. Feedstuffs supplied on the farm showed the deficient level of the Se content below 50 ppb and a very low level of alpha-Toc. It was concluded that WMD of lambs in Hokkaido was nutritional muscular dystrophy resulted from deficiencies of Toc and Se to their ewes.

Primary structure of bovine lactate dehydrogenase-A isozyme and its synthesis in Escherichia coli.

Eleven cDNA clones encoding lactate dehydrogenase(LDH)-A isozyme were isolated from a bovine lymphocyte cDNA library, and the nucleotide sequences of three of the clones (pLDH5, pLDH9 and pLDH12) were determined. With the exception of variation in the 5' portion, two cDNA clones (pLDH9 and pLDH12) appeared to contain the full-length cDNA of 1786 bp, consisting of the protein-coding sequence (996 bp), the 5'- and the 3'-untranslated regions and the poly(dA) tail. The predicted amino acid (aa) sequence of bovine LDH-A (332 aa) showed 96.7% homology with that of pig LDH-A. The protein-coding cDNA region (1650 bp) was inserted into an Escherichia coli expression vector ptac11 and expressed. The protein synthesized in E. coli showed enzyme activity of LDH and was identified by cellogel electrophoresis as LDH-5 isozyme whose subunit M chain is the product of the LDH-A gene.

Origin of abomasum gas in the cows with displaced abomasum.

The gas composition of the rumen and the abomasum were investigated in 71 cows including 25 cows with displaced abomasum. The ratio of methane (CH4) to CO2 in the abomasum gas was higher than that in the rumen. In the incubated mixtures of fresh rumen fluid, CH4 was detected at a high concentration, while a small amount of CO2 and CH4 was found in incubated mixtures of fresh abomasum fluids. Hence, it was suggested that the gas in the abomasum originated not from the abomasum itself but from the rumen. In cows fed with mixed roughages and concentrates, the rumen CO2:CH4 ratio was higher compared with cows fed with roughages alone. Marked changes in rumen gas composition were presumed to be associated with changes in abomasum gas composition. Consequently, it is possible that feeding a large quantity of concentrates may increase the flow rate of the ingesta through the forestomach, so that the return rate of gas separated from the ingesta in the abomasum to the forestomach may also increase. Therefore, the frequent accumulation of a large amount of gas with concomitant changes in the total volatile fatty acid (VFA) concentration in the abomasum caused abomasal atony and subsequently displaced abomasum.

Effects of dihydroheptaprenol on neutrophil functions in calves.

Microemulsified dihydroheptaprenol (DHP) was administered intramuscularly to clinically healthy calves at doses of 4, 6 and 8 mg/kg body weight. Marked increase of neutrophil counts was observed 0.5, 1 and 2 days after the injection at a dose of 6 mg/kg. Nitroblue tetrazolium reducing activity of neutrophils was also obviously enhanced 1, 2 and 3 days after DHP administration. Phagocytic killing of Staphylococcus aureus by neutrophils was further enhanced markedly 1, 2 and 3 days after the drug injection at 6 mg/kg.

Studies on serum tocopherol, selenium levels and blood glutathione peroxidase activities in calves with white muscle disease.

For the purpose of clarifying the cause of white muscle disease (WMD) in calves, tocopherol and selenium levels and blood glutathione peroxidase (GSH-Px) activity were measured on 10 calves with WMD and nine of their dams. The main clinical symptoms of the 10 calves with WMD were motor disturbances including recumbency and stiffness. Serum enzyme activities (GOT, GPT, CPK, LDH) in calves with WMD increased markedly, and this increase was also observed in some of their dams. Serum tocopherol levels of calves with WMD were low, 70% of which showing deficient levels of less than 70 micrograms/100 ml. Serum selenium levels of all the calves were lower than 35 ppb, indicating a deficiency, and were accompanied by low blood GSH-Px activity. alpha-Tocopherol and selenium concentrations in organs were very low. Dams of calves with WMD showed low serum tocopherol levels, 22% of which indicating deficient levels below 150 micrograms/100 ml. Serum selenium levels in dams showed a marked decrease to under 20 ppb, and also low blood GSH-Px activity. Feedstuffs supplied in the farms to affected calves indicated very low alpha-tocopherol contents (below 3 mg/100g DM) and low selenium concentrations below 50 ppb in DM. It was concluded that WMD in calves was attributable to nutritional muscular dystrophy caused by deficiencies in tocopherol and selenium in feedstuffs supplied to their dams.

Studies on serum selenium and tocopherol in white muscle disease of foal.

In order to clarify the cause of white muscle disease (W.M.D.) in foals, tocopherol and selenium concentrations in serum and glutathione peroxidase activities in blood were measured. Examination was made on the samples from horses affected with W.M.D., the foal kept with them in the same stable, the foals kept in the stables without affected foals, and respective mares. The heavy-breed horses in Fukuoka prefecture and Tokachi district were also examined for comparison. Serum tocopherol levels of these foals were normal because after intake of colostrum. Mares of affected foals showed lower tocopherol levels than other examined mares (p less than 0.01). Serum selenium levels of all foals were below 65 ppb, showing deficient levels. The mares of affected foals had lower selenium levels than other mares (p less than 0.01). There was a good correlation between serum selenium concentration and blood glutathione peroxidase activity (r = 0.81). Selenium levels in the liver of affected foals were lower than the foals which succumbed with other diseases. Diet supplied in the stables with affected foals showed low alpha-tocopherol and selenium contents. These findings suggest that W.M.D. in foals is attributed to nutritional muscular dystrophy caused by tocopherol and selenium deficiency of their mares in late gestation period.

Clinical and biochemical findings in bovine cerebrocortical necrosis produced by oral administration of amprolium.

Purposing to get some hints on cause and early diagnosis for cerebrocortical necrosis (CCN), CCN was produced in three healthy calves by the oral administration of amprolium. All three calves showed central nervous signs characterized by ataxic gait, clonic spasm, astasia and opisthotonus, from 24 to 49 days after the start of daily administration of 321-418 mg/kg amprolium. They showed bradycardia from about 20 days before the appearance of the nervous signs, which was supposed to be a finding of primary change and to be useful for early diagnosis of CCN. At necropsy of the two calves, large necrotic lesion was found in the cerebral cortex, and tissue thiamine levels decreased significantly, especially in cerebrum and cerebellum. In the other calf, injection with 25 mg thiamine tetrahydrofurfuryl disulfide (TTFD) was proved to be effective for the recovery of clinical signs. No significant changes in thiamine level were recorded in the whole blood, but those in erythrocytes decreased slightly at about a week before the appearance of the clinical signs. No significant alteration of thiamine excretion was observed in urine. Those findings suggest that CCN in calves is caused by thiamine deficiency and that the blood thiamine levels cannot be used for diagnosis of CCN.