Haemophilia A in a female mixed-breed dog.

A spayed female mixed-breed dog was presented with excessive bleeding from a wound in the mouth. The dog had a history of self-limiting bleeding following ovariohysterectomy. A coagulation test revealed prolongation of the activated partial thromboplastin time (20.2 seconds; reference interval: 11.0-15.0 seconds), prothrombin time was normal and factor VIII activity was markedly decreased (1.9%; reference interval: >50%). The von Willebrand factor antigen concentration was 158% (reference interval: >50%). A cross-mixing test indicated that the diminished factor VIII activity was due to deficiency or dysfunction of factor VIII rather than inhibition of factor VIII activity. Based on these results, the dog was diagnosed with haemophilia A. Haemophilia A should be considered in the differential diagnosis of bleeding disorders also in female mixed-breeds dogs.

Visualization of Toxoplasma gondii stage conversion by expression of stage-specific dual fluorescent proteins.

To recognize the stage conversion of Toxoplasma gondii between tachyzoite and bradyzoite in live host cells, a transgenic T. gondii line, which expressed stage-specific red and green fluorescence, was constructed. T. gondii PLK strain tachyzoites were stably transformed with genes encoding red fluorescent protein (DsRed Express) and green fluorescent protein (GFP) under the control of tachyzoite-specific SAG1 and bradyzoite-specific BAG1 promoters, respectively. The resulting transgenic parasite was designated PLK/DUAL. When PLK/DUAL was cultured in pH 7.0 medium, the PLK/DUAL zoites expressed red fluorescence, but no detectable levels of green fluorescence were observed. The PLK/DUAL zoites reacted with anti-SAG1 antibody, but not anti-BAG1 antiserum. When PLK/DUAL was cultured under high pH conditions, or in the presence of the p38 MAPK inhibitor SB202190, a small number of zoites expressed green fluorescence and were BAG1 positive. C57BL/6J mice were infected with PLK/DUAL tachyzoites. During the acute and reactivating phase, zoites expressed red fluorescence. However, green fluorescence was not detectable. By contrast, latent cysts expressed green fluorescence. The stage-specific dual fluorescence of PLK/DUAL facilitates identification of the parasitic stage in live cells, with the advantage that fixation or immunostaining is not required.

Association of laboratory data and death within one month in cats with chronic renal failure.

OBJECTIVES: To retrospectively compare the data taken at the first visit of 34 cats with chronic renal failure surviving more than one month (surviving group) and 16 cats dying within one month (non-surviving group). METHODS: Records were collected on cats with chronic renal failure presented to a private veterinary practice in Nagoya, Japan, from March 1996 to March 2005. All cats with chronic renal failure diagnosed on the basis of case histories, clinical signs (such as, lethargy, anorexia, loss of bodyweight and vomiting) and a high plasma creatinine (>180 micromol/l) were included in the study. RESULTS: Plasma creatinine, urea nitrogen, inorganic phosphate, packed cell volume and urine protein/creatinine ratio were significantly different between cats of the surviving and non-surviving groups. In the surviving group, survival statuses were recorded, and laboratory data was obtained within one month before death in 13 cats. In the 13 cats, plasma creatinine, packed cell volume and urine protein/creatinine ratio showed significant differences between the data taken within one month before death and that taken at first visit, and only urine protein/creatinine ratio exhibited a consistent alteration (increase) in relation to first visit data. CLINICAL SIGNIFICANCE: These results indicated that plasma creatinine, urea nitrogen, inorganic phosphate, packed cell volume and urine protein/creatinine ratio were associated with death within one month and urine protein/creatinine ratio was most likely to be associated with mortality in cats with chronic renal failure.

Renal reabsorption of magnesium and calcium by cattle with renal tubular dysplasia.

The concentrations of magnesium and calcium in the serum and urine and their rates of clearance were determined in cattle with renal tubular dysplasia, an autosomal recessive hereditary disease associated with a deletion of the paracellin-1 gene in Japanese Black cattle. There were no significant differences in the serum or urine magnesium concentrations between normal cattle and cattle which were heterozygous or homozygous for the condition. Serum calcium concentrations tended to be lower in the homozygous cattle, and the serum creatinine and urea nitrogen concentrations were significantly higher in the homozygous cattle. The ratio of magnesium:creatinine and the fractional excretion of magnesium were higher in cattle with the disease than in normal cattle. There were no significant differences in urine calcium concentration, the calcium:creatinine ratio, and fractional excretion of calcium between normal cattle and cattle which were homozygous or heterozygous for the condition. The creatinine clearance was significantly lower in the homozygous cattle than in normal cattle. The clearance, excretion rate, reabsorption rate and reabsorption rate:clearance ratio of magnesium in cattle with renal tubular dysplasia were significantly lower than in normal cattle. The clearance rate and reabsorption rate of calcium were also significantly lower in the affected cattle, but the excretion rate and reabsorption rate:clearance of calcium were not different between the normal cattle and the cattle homozygous for the condition. In cattle with the condition the rate of reabsorption of magnesium by the kidneys was low, but the rate of reabsorption of calcium was normal.

Pathological changes of renal tubular dysplasia in Japanese black cattle.

Pathological studies were conducted on 91 Japanese Black cattle with a hereditary disease which induced growth retardation, long hooves and renal failure. In calves one to two months old, no gross abnormalities were observed in the kidneys, but microscopical examinations revealed immature epithelia which were arranged irregularly and not attached to the basement membranes in some proximal tubules. In animals three to 36 months old, the kidneys had shrunk perceptibly and had grey-white radial streaks; microscopically they showed severe interstitial fibrosis with round-cell infiltration in the outer zone of the medulla and cortex, and reductions in the numbers of glomeruli and tubules. In the fibrotic areas there were immature epithelia with an irregular arrangement, and the basement membrane of the tubules was thickened. It was concluded that renal tubular dysplasia was the primary lesion of the disease, and that interstitial fibrosis and reductions in the numbers of nephrons were secondary lesions.

E7070, a novel sulphonamide agent with potent antitumour activity in vitro and in vivo.

E7070 (N-(3-Chloro-7-indolyl)-1,4-benzenedisulphonamide) was selected from our sulphonamide compound collections via antitumour screening and flow cytometric analysis. Following treatment with E7070, the cell cycle progression of P388 murine leukaemia cells was disturbed in the G1 phase. The cell-killing effect on human colon cancer HCT116 cells was found to be time-dependent. In the panel of 42 human tumour cell lines, E7070 showed an antitumour spectrum that was distinct from those of other anticancer drugs used in clinic. Animal tests using human tumour xenograft models demonstrated that E7070 could cause not only tumour growth suppression, but also tumour regression in three of five colorectal and two of two lung cancers. In the HCT116 xenograft model, E7070 was shown to be superior to 5-FU, MMC and CPT-11 (irinotecan). Furthermore, complete regression of advanced LX-1 tumours was observed in 80% of E7070-treated mice. All of these observations have promoted this drug to clinical evaluation.

Clinical features of renal tubular dysplasia, a new hereditary disease in Japanese Black cattle.

A new hereditary disease characterised by renal failure, poor growth and long hooves in Japanese Black cattle (wagyu) has been recognised in a region of central Japan since 1990. The number of calves affected has increased gradually, with the incidence reaching 17 of 485 (3.51 per cent) in 1995. Almost all the calves were slightly undersized at birth, and repeatedly had diarrhoea during the neonatal period. They began to show signs of growth retardation with proportional body and elongation of the hooves from about two to five months of age, but they had an almost normal or only slightly decreased appetite. The concentrations of urea nitrogen, creatinine and inorganic phosphorus in serum were high, and the affected calves excreted diluted urine frequently. Among 25 cases, the urine of 21 contained occult blood, 24 contained protein and two contained glucose. In 29 calves observed for 30 to 130 days, the course of the disease varied; in 21 of them it remained unchanged, six became gradually worse and two became severely debilitated and died. The disease was diagnosed as renal tubular dysplasia by histopathological examination.

Relaxing and contracting activities of heartworm extract on isolated canine abdominal aorta.

Effect of adult heartworm (HW) crude extract on isolated canine abdominal aortic strips precontracted with noradrenaline was examined by recording isometric changes in tension. HW extract caused contraction of the aortic strip at a low concentration (LC) and its relaxation at a high concentration (HC). In aortic strips without endothelium, LC extract elicited a contraction similar to that in the strips with endothelium, whereas HC extract failed to produce any relaxation but instead produced a contraction. The relaxing effect of HC extract was blocked after treatment with 300 microM NG-nitro-L-arginine methyl ester hydrochloride, with reversal by additional treatment with 3 mM L-arginine. It was also markedly reduced or abolished after treatment with 3 microM oxyhemoglobin or 1 microM methylene blue. Fractionation of HW extract by high-performance liquid chromatography revealed that the relaxing and contracting activities are due to different substances in the extract. The results indicate that HW extract contains 2 different vasoactive substances, 1 causing contraction of canine abdominal aorta via a direct action on the smooth muscle, and the other its relaxation indirectly by releasing nitric oxide from endothelial cells. These vasoactive substances might play a role in HW extract-induced shock in dogs, and in the pathogenesis of HW infection.

Comparison of heartworm extract-induced shock and endotoxin-induced shock in dogs by determination of serum tumor necrosis factor concentrations.

OBJECTIVE: To compare the mechanisms of heartworm (HW) extract-induced shock and endotoxin-induced shock in dogs by determination of serum tumor necrosis factor (TNF) concentrations. ANIMALS: 11 mixed-breed dogs (7 without and 4 with HW infections). PROCEDURE: Eight dogs were treated with 2 ml of HW extract IV, and 3 dogs were given endotoxin (Escherichia coli lipopolysaccharide [LPS]) at 40 or 400 microg/kg of body weight, IV. Changes in clinical and hematologic findings and serum TNF concentrations were examined from before treatment to 120 minutes after treatment in dogs given HW extract or from before treatment to 180 minutes after treatment in dogs given LPS. Tumor necrosis factor concentration was determined by cytotoxic assay, using WEHI-164 murine sarcoma cells, and plasma endotoxin concentration was determined in 2 dogs treated with HW extract, using the endotoxin-specific chromogenic test. RESULTS: Eight dogs developed shock 3 to 16 minutes after HW extract treatment. Rectal temperature did not change during examination. Serum TNF concentration was detected at a low concentration only 60 and 120 minutes after HW extract treatment, and plasma endotoxin was not detected during examination. In dogs treated with LPS, rectal temperature increased to > 40 C in 2 of 3 dogs, and serum TNF concentration began to increase 30 minutes after LPS treatment, reaching a maximum concentration by 60 minutes. CONCLUSIONS: The cause and mechanism of HW extract-induced shock may be different from those of endotoxin-induced shock, because TNF, which was a pivotal mediator in endotoxin-induced shock, increased minimally in serum of dogs treated with HW extract.

Role of histamine in heartworm extract-induced shock in dogs.

OBJECTIVE: To determine whether heartworm (HW) extract-induced shock in dogs is consistent with anaphylactic shock by examining the role of histamine. ANIMALS: 6 mixed-breed dogs (3 without and 3 with HW infections) and 4 specific pathogen-free (SPF) Beagles. PROCEDURE: Four experiments were performed as follows: 1) 6 mixed-breed dogs were treated IV with 2 ml of HW extract, and plasma histamine concentrations were determined; 2) 4 SPF dogs were treated IV with 2 ml of HW extract and examined for shock; 3) sera from 6 dogs of experiment 1 and from 4 SPF dogs of experiment 2 that were obtained before HW extract treatment were tested for heterologous passive cutaneous anaphylaxis (PCA), using rabbits during a sensitization period of 48 to 72 hours; and 4) mast cell degranulation by HW extract was tested, using rat mesentery and canine cultured mast cells. RESULTS: Experiment 1: 6 dogs developed shock, and plasma histamine concentrations increased significantly from 0.3 +/- 0.2 (mean +/- SD) ng/ml before HW extract treatment to 44.6 +/- 68.9 ng/ml at the onset of shock; experiment 2: all SPF dogs developed shock and had an increase in plasma histamine concentrations; experiment 3: sera from mixed-breed dogs without HW infection and from SPF dogs had negative PCA reactions; experiment 4: HW extract degranulated rat mesentery mast cells and released histamine directly from canine mast cells. CONCLUSIONS AND CLINICAL RELEVANCE: Results of our study indicate that an unknown mast cell-degranulating substances contained in HW extract may degranulate mast cells directly, consequently releasing histamine that may participate in the onset of shock in HW extract-induced shock in dogs.

Effect of E7010 on liver metastasis and life span of syngeneic C57BL/6 mice bearing orthotopically transplanted murine Colon 38 tumor.

PURPOSE: E7010 is an orally active sulfonamide antitumor agent showing good activity against various subcutaneously inoculated rodent tumors and human tumor xenografts. The purpose of this study was to evaluate the effect of E7010 on liver metastasis and life span of mice bearing orthotopically transplanted murine Colon 38 tumor. METHODS: Orthotopic transplantation of murine Colon 38 tumor as intact tissue yielded hepatic metastasis with a high incidence in about 1 month in C57BL/6 mice, and the mice died in about 2 months with cachexia. In this model, the maximum tolerated dose of E7010 (100 mg/kg per day) was administered orally on various schedules, including for 14 days or daily until death, starting at 14 days after transplantation, or for 8 days from 21 days after transplantation. RESULTS: E7010 showed tumor growth inhibition (T/C=40%) at the orthotopic site similar to that at the subcutaneous site (T/C = 32%) when administered from 14 days after transplantation. When E7010 was started from 21 days after transplantation, it significantly decreased the number of hepatic metastases (control 17.1+/-20.8, E7010 2.6+/-5.3), although inhibition of tumor growth at the orthotopic site was only moderate (T/ C=60%). The administration of E7010 until death produced a significant increase in life span (control 49.8+/-8.9 days, E7010 62.5+/-6.1 days). Although the tumor weight of the E7010-treated group on the day of death was similar to that of the untreated group (control 1.166+/-0.507 g, E7010 1.211+/-0.632 g), there were significantly fewer liver metastases in the E7010-treated group (control 41.3+/-31.1, E7010 2.0+/-2.0). CONCLUSION: E7010 suppressed tumor growth at both primary and metastatic sites and increased life span in an orthotopic transplantation model of murine Colon 38 tumor in syngeneic C57BL/6 mice. Hepatic metastasis was inhibited more effectively than the growth of the primary tumor.

Serum growth hormone and insulin-like growth factor-1 concentrations in Japanese black cattle with growth retardation.

Serum concentrations of growth hormone (GH) and insulin-like growth factor-1 (IGF-1) were determined in 5 calves in the same lineage with growth retardation. They had normal appetites, activities, body proportion, and laboratory test results. Calves with growth retardation had higher serum GH concentrations and lower serum IGF-I concentrations. These findings suggested defects in the GH-IGF-1 axis, such as in the GH-receptor.

Hemodynamic alterations in dogs with shock induced by intravenous injection of heartworm extract.

To elucidate one way of the shock mechanisms, the hemodynamic alterations were examined in 7 dogs with heartworm (HW) extract-induced shock. The first alteration observed after injection of HW extract was a decrease in right ventricular end-diastolic pressure (RVEDP). After that, left ventricular (LV) end-diastolic pressure, LV systolic pressure, and LV dp/dt fell significantly, followed by a decrease in the cardiac output of all dogs to below the detectable level (1.00 l/min). Since RVEDP depends on blood flow into the right ventricle, the decrease in RVEDP means a reduction in venous return. Therefore, this study showed that the first trigger of a decrease in blood pressure in HW extract-induced shock is the reduction in venous return.

[ALL complicated by obstructive jaundice due to choledocholithiasis after unrelated bone marrow transplantation].

A 22-year-old man with acute lymphocytic leukemia underwent allogeneic bone marrow transplantation (BMT) from an unrelated donor in October 1996. In April 1997, he suddenly developed severe abdominal pain with nausea and vomiting. The diagnosis was obstructive jaundice associated with gallstones in the gallbladder and common bile duct. The patient underwent laparoscopic cholecystectomy and endoscopic removal of the stones in the common bile duct. The major component of the gallstones was bilirubinate calcium. Although the pathogenesis of gallstones after BMT remains unclear, several factors including impaired contractility of the gallbladder, hemolysis, changes in bile composition, and biliary tract infection may play important roles.

A deletion of the paracellin-1 gene is responsible for renal tubular dysplasia in cattle.

Various hereditary diseases analogous to particular human heritable diseases have been identified in cattle. Investigation of these cattle diseases will provide useful information regarding the pathogenesis of the corresponding human diseases. Renal tubular dysplasia is an autosomal recessive disease of Japanese black cattle characterized by renal failure and growth retardation. We have previously mapped the locus responsible for the disease within a region on bovine chromosome 1. In the present study, we further typed additional markers in this region and found that a genomic segment of bovine chromosome 1 including the microsatellite marker BMS4009 was deleted in the affected animals. Construction of a physical map covering this region with BAC clones and comparison of the nucleotide sequences of this region between normal and affected animals revealed that a region of 37 kb including exons 1 to 4 of the bovine paracellin-1 gene was deleted in the affected animals. The paracellin-1 gene, which is the causative gene for human renal hypomagnesemia with hypercaciuria and nephrocalcinosis, encodes a tight junction protein of renal epithelial cells. Therefore, we concluded that deletion of the paracellin-1 gene is responsible for renal tubular dysplasia of cattle, and the cattle disease could be a good model for the human disease.

A focused compound library of novel N-(7-indolyl)benzenesulfonamides for the discovery of potent cell cycle inhibitors.

A series of compounds containing an N-(7-indolyl)benzenesulfonamide pharmacophore was synthesized and evaluated as a potential antitumor agent. Cell cycle analysis with P388 murine leukemia cells revealed that there were two different classes of potent cell cycle inhibitors; one disrupted mitosis and the other caused G1 accumulation. Herein described is the SAR summary of the substituent patterns on this pharmacophore template.

Plasma renin activities, angiotensin II concentrations, atrial natriuretic peptide concentrations and cardiopulmonary function values in dogs with severe heartworm disease.

Relationships among plasma renin activities (PRA), plasma angiotensin II (ATII) concentrations, atrial natriuretic peptide (ANP) concentrations and cardiopulmonary function values were examined in dogs with ascitic pulmonary heartworm disease and acute- and chronic-vena caval syndrome (CS). PRA, plasma ATII concentration and plasma ANP concentration tended to be higher or were significantly higher in dogs with ascites, acute- and chronic-CS. PRA correlated significantly with plasma ATII concentration, WBC count, ALP activity, plasma concentrations of urea nitrogen, creatinine, sodium, potassium, and chloride, right ventricular endodiastolic pressure and right atrial pressure. Plasma ATII concentration correlated significantly with WBC count, plasma concentrations of urea nitrogen, sodium, and potassium, right ventricular endodiastolic pressure and right atrial pressure. Plasma ANP concentration did not correlate with PRA or ATII concentration, but correlated significantly only with pulmonary arterial pressure.

A general method for acylation of indoles at the 3-position with acyl chlorides in the presence of dialkylaluminum chloride.

[reaction--see text] Indoles are selectively acylated at the 3-position in high yields on treatment with a wide variety of acyl chlorides in CH(2)Cl(2) in the presence of diethylaluminum chloride or dimethylaluminum chloride. The reaction proceeds under mild conditions and is applicable to indoles bearing various functional groups without NH protection.

Homozygosity mapping of the locus responsible for renal tubular dysplasia of cattle on bovine chromosome 1.

Renal tubular dysplasia is a hereditary disease of Japanese black cattle showing renal failure and growth retardation with an autosomal recessive trait. In the present study, we mapped the locus responsible for the disease (RTD) by linkage analysis with an inbred paternal half-sib pedigree obtained from commercial herds. By analyzing segregation of microsatellite markers in the half-sibs, significant linkage was observed between the RTD locus and markers on bovine Chromosome (Chr) 1 with the highest lod score of 11.4. Homozygosity mapping with the inbred pedigree further defined the localization of the RTD locus in a 4-cM region between microsatellite markers BMS4003 and INRA119. Mapping of the RTD locus on bovine Chr 1 will facilitate cloning and characterization of the gene responsible for this disease.