Glucagon-like peptide-1 (GLP-1) is a polypeptide that is mainly produced by intestinal L cells and is encoded by the proglucagon gene. In this study, GLP-1 localisation was investigated in the ileum of healthy and diabetic mice by immunohistochemistry and proglucagon gene expression was assayed by reverse transcription-polymerase chain reaction. This study included 18 male Balb/c mice that were divided into diabetic, sham, and control groups. Mice in the diabetic group received 100 mg/kg of streptozotocin. Immunohistochemical expression of GLP-1 was determined using the avidin–biotin–peroxidase complex technique, and proglucagon gene expression was determined by RT-PCR. Analysis of GLP-1 immunohistochemical localisation showed that GLP-1-immunopositive cells (L cells) were present between epithelial cells in the intestinal crypts. The intensity and localisation of GLP-1 immunoreactivity were similar among the mice in all the groups. Proglucagon gene expression levels were also statistically similar among the mice in all the groups. No difference was demonstrated among the mice in the diabetic, sham, or control groups with respect to proglucagon gene expression and GLP-1 localisation in the ileum, suggesting that diabetes does not affect proglucagon gene expression in the ileum.

Veterinarians use flumequine (FLU) widely but its toxicological effects are still unclear. FLU doses of 53, 200, or 750 mg/kg were administered orally for six weeks to pubertal male rats for evaluation of their toxicity. Weight gain was poorer after seven days of exposure to FLU 750, but relative weights of the brain, adrenal and thyroid glands, and testes were notably higher. Haematological and lipid profile parameters, cardiac markers, and inorganic phosphate significantly increased in the FLU 750 group. Blood glucose, oestradiol and serum concentrations of immunoglobulins G (IgG) and E (IgE) significantly decreased after treatment. The levels of interleukins 10 (IL-10) and 6 (IL-6) fell significantly in the FLU 200 and FLU 750 groups. Cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) and cyclooxygenase-2 (Cox-2) expression amplified after treatment. Serum levels of free triiodothyronine (fT3) and free thyroxine (fT4) reduced in the FLU 200 and FLU 750 groups without changes in total T3 or T4 level. All doses of FLU significantly depressed concentrations of thyroid-stimulating hormone (TSH) and testosterone. Histopathology of thyroid glands from rats treated with FLU 750 showed degeneration and depletion of thyroid follicular epithelial cells. Expression of 8-hydroxydeoxyguanosine (8-OHdG) was increased in a dose-dependent manner in the brain, but decreased in the testes. Expression of CYP1A1 increased in the adrenal and pituitary glands. The results of this study suggest that the toxicity of FLU in rats is an effect of its disruptive influence on the pituitary-thyroid hormonal system and on the dysfunction of the immune system.

Objective: To use microarray analysis to identify genes that are differentially expressed in horses with experimentally induced osteoarthritis. Animals: 24 horses. Procedures: During arthroscopic surgery, a fragment was created in the distal aspect of the radiocarpal bone in 1 forelimb of each horse to induce osteoarthritis. At day 14 after osteoarthritis induction, horses began exercise on a treadmill. Blood and synovial fluid samples were collected before and after surgery. At day 70, horses were euthanized and tissues were harvested for RNA analysis. An equine-specific microarray was used to measure RNA expression in peripheral WBCs. These data were compared with mRNA expression (determined via PCR assay) in WBCs, cartilage, and synovium as well as 2 protein biomarkers of cartilage matrix turnover in serum and synovial fluid. Results: A metalloproteinase domain-like protein decysin-1 (ADAMDEC1), glucose-regulated protein (GRP) 94, hematopoietic cell signal transducer (HCST), Unc-93 homolog A (hUNC-93A), and ribonucleotide reductase M2 polypeptide (RRM2) were significantly differentially regulated in WBCs of horses with osteoarthritis, compared with values prior to induction of osteoarthritis. There was correlation between the gene expression profile in WBCs, cartilage, and synovium and the cartilage turnover proteins. Gene expression of ADAMDEC1, hUNC-93A, and RRM2 in WBCs were correlated when measured via microarray analysis and PCR assay. Conclusions and Clinical Relevance: Expression of ADAMDEC1, GRP94, HCST, hUNC-93A, and RRM2 was differentially regulated in peripheral WBCs obtained from horses with experimentally induced osteoarthritis. Gene expression of ADAMDEC1, hUNC-93A, and RRM2 in peripheral WBCs has the potential for use as a diagnostic aid for osteoarthritis in horses.

The neutrophil plasma membrane has a major role in migration, phagocytosis, and destruction of microorganisms. Neutrophils isolated from blood and mammary secretions were homogenized, and the plasma membrane fraction was isolated on discontinuous sucrose gradient (20, 32, and 50%). Purity of plasma membrane preparation was determined by use of marker enzyme analysis. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the membrane proteins was performed under reducing conditions for polypeptide characterization. The membrane proteins were also labeled with 125I externally, using 1,3,4,6-tetrachloro-3 alpha-6 alpha-diphenylglycouril, and proteins were separated by SDS-PAGE and autoradiographed. Compared with whole cell homogenate, the plasma membrane fraction obtained at the 20/32% interface was enriched for the marker enzymes, 5'-nucleotidase (16-fold), alkaline phosphatase (5.5-fold), and total phosphatase (26-fold). The membrane fraction had minimal specific activity for beta-glucuronidase (0.4-fold), compared with whole cell homogenate. Plasma membrane protein yield was about 500 micrograms/ 10(9) bovine blood neutrophils. The SDS-PAGE of plasma membrane proteins revealed 25 protein bands, of which 13 were major bands. There were 3 distinct bands (18, 36, and 65 kd) in the plasma membrane-enriched fraction (20/32 interface) that were not seen in other fractions (30/50% and pellet). Further, 125I-labeling identified 5 distinct protein bands (205, 140, 65, 35, and 30 kd). Blood and mammary neutrophils had similar polypeptide patterns, except that 36- and 65-kd bands were more prominent for blood neutrophils than for mammary neutrophils.

The antigenic profile of Ehrlichia canis, E risticii, E sennetsu, and E equi was investigated by the use of protein (western) immunoblot technique. Results of analysis of serum from acutely and chronically infected animals indicated that the 4 Ehrlichia species share a unique 25-kD polypeptide in addition to other peptides. Immune sera from dogs inoculated with E canis recognized a wide range of E canis polypeptide antigens, as determined by western blot analysis. A larger number of E sennetsu polypeptides were detected when homologous antiserum and antiserum to E equi were used. The latter antiserum did not recognize antigens of E canis or E risticii. Antisera to E canis, E risticii, and E sennetsu detected E equi antigens. Data indicate that a 25-kD protein is a common antigen among the species of the genus Ehrlichia and that the ascending order of abundance of immunodominant determinants in the 4 species of Ehrlichia studied would be: E risticii leads to E equi leads to E sennetsu leads to E canis. Implications of these findings for diagnosis of ehrlichial infections and prophylaxis are evident.

Polypeptides from whole cell preparations of Pasteurella multocida serotypes A:12 and A:3 were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and transferred to nitrocellulose paper. Antigens were detected by immunoblot analysis, using sera from 3 groups of rabbits. Sera were obtained from rabbits inoculated intranasally with P multocida serotype A:12 or A:3, from rabbits maintained in a rabbitry with enzootic P multocida A:12 infection, and from rabbits maintained in a rabbitry with enzootic P multocida A:3 infection. Immunoblot analyses of pre- and postinoculation sera from experimentally infected rabbits, using serotype A:12 antigen, revealed 3 polypeptides with approximate molecular mass of 28, 30, and 37 kDa that consistently detected antibodies after P multocida-induced infection. Sera from rabbits naturally infected with either serotype, tested against serotype A:12 and A:3 antigens, detected the same polypeptides in both serotypes. Thus, immunologic reactivity to these polypeptides may be useful for serologic detection of P multocida infection.

A series of experiments was performed in vitro and in vivo to determine the influence of FK-565, a heptanoyl tripeptide, on lymphocyte and macrophage function in swine. Compared with values for control cultures, mitogen-stimulated lymphocyte blastogenesis and interleukin-2 production were unaffected in cells preincubated with 0.1, 1.0, and 10.0 microgram of FK-565/ml. Natural killer cell activity was increased by preincubation with 1.0 microgram of FK-565/ml; however, this increase was not statistically significant. In vitro treatment of porcine alveolar macrophages with FK-565 did not enhance cytolytic activity or bactericidal activity. In in vivo experiments, FK-565 given orally to pigs at concentrations of 6 or 60 microgram-kg-l.-d-1 for 5 days did not affect lymphocyte blastogenesis, interleukin-2 production, or alveolar macrophage bactericidal activity. A trend toward increased natural killer cell activity was evident in pigs treated with FK-565. In contrast, pigs treated with 6 microgram-kg-1.-d-1 had significantly (P less than 0.01) decreased alveolar macrophage cytolytic activity. These data indicate that at the dosages tested, FK-565 is not a suitable immunomodulator for enhancement of nonspecific immunity in swine.

Objective—To evaluate and compare circulating concentrations of islet amyloid polypeptide (IAPP), insulin, and glucose in nondiabetic cats classified by body condition score (BCS) and in cats with naturally occurring diabetes mellitus. Animals—109 (82 nondiabetic, 21 nonketoacidotic diabetic, and 6 ketoacidotic diabetic) cats. rocedures—Cats were examined and BCSs were assessed on a scale of 1 to 9. After food was withheld for 12 hours, blood was collected and plasma concentrations of IAPP and serum concentrations of insulin and glucose were measured. Differences in these values were evaluated among nondiabetic cats grouped according to BCS and in diabetic cats grouped as ketoacidotic or nonketoacidotic on the basis of clinicopathologic findings. Correlations were determined among variables. Results—In nondiabetic cats, BCS was significantly and positively correlated with circulating IAPP and insulin concentrations. Mean plasma IAPP concentrations were significantly different between cats with BCSs of 5 and 7, and mean serum insulin concentrations were significantly different between cats with BCSs of 5 and 8. Serum glucose concentrations were not significantly different among nondiabetic cats. Mean IAPP concentrations were similar between nonketoacidotic diabetic cats and nondiabetic cats with BCSs of 8 or 9. Mean IAPP concentrations were significantly reduced in ketoacidotic diabetic cats, compared with those of nondiabetic cats with BCSs of 6 through 8 and of nonketoacidotic diabetic cats. Conclusions and Clinical Relevance—Results indicated that increased BCS (a measure of obesity) is associated with increased circulating concentrations of IAPP and insulin in nondiabetic cats.

To evaluate the transplacental transfer of Theileria sergenti infection in cattle, we used DNA probes to detect T. sergenti in 6 pregnant cows and their calves. All the animals were monitored by parasitologic, serologic, and polymerase chain reaction (PCR) assays for a predicted 875-base-pair (bp) DNA product and a 684-bp amplicon detected by nested PCR in the blood and spleens of aborted fetuses. An open reading frame (ORF) starting at nucleotide 170 and terminating at position 1021 was shown to code for a polypeptide of 283 amino acid residues. All 6 dams and 5 calves were positive for T. sergenti in all tests. One calf was positive only with nested PCR. We conclude that transplacental transmission of T. sergenti is a significant problem. The relevance of the data in the programmed introduction of new (especially pregnant) animals into established clean herds needs serious consideration with regard to control of theileriosis and other tickborne diseases.

Pili from 11 distinct serotypes of Bacteroides nodosus were examined for diversity of pilin polypeptide subunits among serotypes and for purity of the pilin preparations. The pilin of all 11 samples was shown to be homogeneous. Mean +/- SD molecular weight of the pilin of 7 serotypes (A198, IV, V, VI, IX, XVII, and XVIII) was 18,500 +/- 100. The pilin of serotypes I, III, and VIII had molecular weight of 17,600, 19,400, and 19,000, respectively. Serotype XV differed greatly from the other 10 serotypes in that 2 distinct polypeptide bands with molecular weight of approximately 7,800 and 6,200 were detected. We suggest that these 2 low molecular weight bands resulted from proteolytic cleavage of the pilin protein.