Insulin-like growth factor I receptor analysis of satellite cell-derived myotube membranes established from two lines of Targhee rams selected for growth rate

Ovine-derived fibroblasts were used to validate an insulin-like growth factor I (IGF-I) membrane-receptor binding assay system. Competitive binding using fibroblasts revealed that half-maximal inhibition of 125I-IGF-I binding by IGF-I was 2.3 nM. SDS-polyacrylamide gel electrophoresis analysis of specific protein-associated 125I-IGF-I was consistent with the migration of 125I-IGF-I-labeled Type I IGF receptor alpha-subunits at Mr 133,000 daltons. Further, the efficiency of two cell solubilization methods was examined and time-dependent binding equilibrium was determined for the membrane assay system. Satellite cell-derived myotubes were subsequently isolated from primary satellite cell cultures established from the semimembranosus muscles of high and low efficiency-of-gain (EOG) Targhee rams, and IGF-I receptor dynamics were measured. A membrane competitive binding study revealed that half-maximal inhibition of 125I-IGF-I binding was achieved by 1-ng IGF-I for low, and 10-ng IGF-I for high, EOG myotube membrane preparations. Kd values were similar between the high EOG (4.78 nM) and low EOG (2.95 nM) groups; however, receptor concentrations (Bmax) appeared to differ between groups. High EOG membrane receptor Bmax was 3.88 pmole/micrograms protein (19.87 pmole/micrograms DNA), whereas low EOG membrane receptor Bmax was 1.22 pmole/micrograms protein (9.28 pmole/micrograms DNA). These preliminary findings support the hypothesis that genetic selection for EOG results in altered satellite cell responsiveness to IGF-I.     

Correlation of DNA ploidy to tumor histologic grade, clinical variables, and survival in dogs with mast cell tumors.

By using flow cytometry, a retrospective analysis of the DNA content of 40 primary canine mast cell tumors and seven lymph nodes that contained metastatic mast cell tumor from 44 dogs of various breed, sex, and age was performed on formalin-fixed, paraffin-embedded samples of the tumors and nodes. These samples were chosen according to the following criteria: samples contained sufficient well-preserved tumor tissue in the paraffin block for processing, sufficient patient history data were available, clean and homogeneous cell suspensions were obtained after processing, and interpretable DNA histograms were produced on analysis. The ploidy data obtained were compared with the histopathologic grade, the anatomical site of occurrence, the clinical stage of the tumors, and the survival of the dogs. Over 70% (29/40) of the mast cell tumors were diploid. Three metastatic mast cell tumors in lymph nodes had the same ploidy status as their corresponding primary tumors. In five dogs, mast cell tumors from multiple sites in each dog displayed similar ploidy status. Of 26 dogs evaluated for survival times, 69% (18/26) had diploid tumors and 31% (8/26) had aneuploid tumors. When numbers of diploid versus aneuploid tumors were compared, no significant difference was found between any two grades, clinical stages, or anatomic sites. A significant difference (P = 0.02) was found, however, between aneuploid and diploid tumors when comparing Stage I and non-Stage I disease. The Kaplan-Meier survival plot indicated a tendency towards an increased survival within the first year in dogs with diploid versus aneuploid tumors (P = 0.06).     

Selective measurement of lipoprotein lipase and hepatic triglyceride lipase in heparinized plasma from horses.

Affinity chromatography on heparin sepharose was used to identify 2 lipolytic enzymes in heparinized plasma from horses. One enzyme was typical of hepatic triglyceride lipase (HTGL), because it was resistant to inactivation by high concentrations of NaCl, and it did not require the addition of serum for activity. The other enzyme was identified as lipoprotein lipase (LPL), because of its inactivation at NaCl concentrations in excess of 0.2M, and its dependency on addition of serum as a source of apolipoprotein C-II activator. The enzymes were purified by 347-(HTGL) and 442- (LPL) fold, with yields of 54 and 58%, respectively. The partially purified enzymes were used to design incubation conditions that gave optimal activities for each enzyme in vitro. A selective assay was then developed for direct measurement of LPL and HTGL activities in heparinized plasma from horses. Analysis of HTGL took advantage of the almost complete inactivation of LPL when serum cofactor was excluded from the assay at the NaCl concentration that gave optimal HTGL activity. Prior incubation of heparinized plasma with sodium dodecyl sulfate to inhibit HTGL was necessary for measurement of LPL, because HTGL retained 67% of its activity at the NaCl concentration required for optimal LPL activity. Activity of each enzyme was measured in heparinized plasma from 12 Shetland ponies. The mean activity +/- SD for LPL was 3.22 +/- 1.04 mumol of fatty acids/ml of heparinized plasma/h (mumol of FA/ml/h. The mean activity for HTGL was 4.9 +/- 1.56 mumol of FA/ml/h. The performance of the assay was assessed by replicate analysis of pools of each enzyme with high and low activities.(ABSTRACT TRUNCATED AT 250 WORDS)     

The contribution from hyphae, roots and organic carbon constituents to the aggregation of a sandy loam under long-term clover-based and grass pastures

Water-stable macro-aggregate size fractions (>2.0 mm, 1.0–2.0 mm, 0.5–1.0 mm and 0.25–0.5 mm) and non-aggregated soil from a sandy loam under long-term clover-based pasture and from grass pasture were analysed to determine the role of acid- and water-extractable carbohydrate C, total hyphal length, microbial biomass, organic C and total and mycorrhizal root length in stabilization of the aggregates. Aggregates were examined by scanning electron microscopy (SEM) and the particle-size distribution of the size fractions was also determined. Macro-aggregation increased under grass, relative to clover-based pasture; however, the properties of the aggregate fractions measured did not reflect this difference. Microbial-biomass C, extractable-carbohydrate C, hyphal length, total and mycorrhizal root length and organic C content of the soils were poorly correlated with macro-aggregation. Within the aggregates, the proportion of 250–1000-km sand was smaller and clay, silt and fine sand (20–250 μm) were greater relative to non-aggregated soil, suggesting that the >250-μm sand in the non-aggregated soil limited the stabilization of macro-aggregates. Under SEM, no enmeshment of aggregates by hyphae and roots was apparent. Although 50–160 m hyphae g^?1 soil was found within the aggregates, calculations showed that on average only 5 to 13 lengths of hyphae were associated with each 250-μm cube of soil within the aggregates, and suggested little potential to stabilize the aggregates by enmeshing. On average, all >2.0-mm aggregates contained less than 3.6 mm of roots and less than 50% by weight of <2.0-mm aggregates contained a single length of root. The findings cast doubt about the role of hyphae and fine roots in the stabilization of macro-aggregates through an enmeshing mechanism in sandy soils.     

Production and characterization of monoclonal antibodies to budgerigar fledgling disease virus major capsid protein VP.

Eleven hybridoma cell lines producing monoclonal antibodies (MAbs) against intact budgerigar fledgling disease (BFD) virions were produced and characterized. These antibodies were selected for their ability to react with BFD virions in an enzyme-linked immunosorbent assay. Each of these antibodies was reactive in the immunofluorescent detection of BFD virus-infected cells. These antibodies immunoprecipitated intact virions and specifically recognized the major capsid protein, VP1, of the dissociated virion. The MAbs were found to preferentially recognize native BFD virus capsid protein when compared with denatured virus protein. These MAbs were capable of detecting BFD virus protein in chicken embryonated cell-culture lysates by dot-blot analysis.     

Comparative susceptibility of caponized and uncaponized tom turkeys to Pasteurella multocida.

When susceptibility to virulent Pasteurella multocida was compared, there was no significant (P greater than 0.05) difference between caponized and uncaponized tom turkeys. Neither was there any significant (P greater than 0.05) difference between the surviving caponized and uncaponized toms in the development of serum anti-P. multocida antibody. However, at 28 weeks of age, the average live body weight of the caponized toms was significantly (P less than 0.05) lower than that of the uncaponized toms. Turkeys were caponized when 9 weeks old, and different groups were exposed to P. multocida when 13, 18, 23, and 28 weeks old.     

Tuberculosis in wild seals and characterisation of the seal bacillus

SUMMARY Tuberculosis was diagnosed in 3 otariid seals found dead on beaches at 3 locations on the south coast of Western Australian between May 1990 and March 1991. This confirms that tuberculosis is present in the 2 native seals (Neophoca cinerea and Arctocephalus forsteri) in Western Australian waters. Mycobacterium sp isolated from the lungs of 2 of the seals were studied to determine the similarity of the strains to each other, to the strains isolated during 1986 from Australian sea lions and New Zealand fur seals kept in captivity at a marine park near Perth, Western Australia, and to a strain isolated in 1988 from a seal trainer who worked with the infected captive seals for 3 years. After restriction endonuclease analysis (REA) with the endonucleases Bst Ell, Bcl I and Pvu II, one of the wild seal strains appeared to have identical DNA fragment patterns to the strains from the captive seals and the seal trainer. The other wild seal isolate had identical REA profiles using Bst EII and Bcl I, but a minor difference was detected using Pvu II. Differences in these isolates were more clearly seen in restriction fragment length polymorphisms after hybridisation with two DNA probes. The secretory protein MPB70, present in M bovis, was not detected in wild seal isolates using sodium dodecyl sulphate polyacrylamide gel electrophoresis and Western blotting techniques. Analysis of protein and DNA fragment profiles indicated that seal tuberculosis isolates form a unique cluster within the M tuberculosis complex.     

The internet and the future of equine practice.

The Internet is a fast-growing technology that allows information (text as well as images) to be shared across the world in an instant. As computer systems, software, and Internet services become less expensive options, the world becomes more experienced and comfortable with these technologies. In the foreseeable future, equine veterinarians are likely to be better educated about diseases and treatments, better connected with specialists for consultations on difficult cases, able to respond to client communications more efficiently, able to accurately educate clients, and able to communicate with colleagues from across the country and the world. Additionally, there is likely to be a time when using the Internet and computers is profitable in all these ways as well as economically. Improving doctor-client relationships and patient care are goals for all veterinarians, and the Internet holds the technology to render these now and in the future.