Earnestine P. Holmes

Insulin resistance in equine digital vessel rings: An in vitro model to study vascular dysfunction in equine laminitis

REASONS FOR PERFORMING STUDY One of the causes of equine laminitis is hyperinsulinaemia, which may be associated with endothelial dysfunction and insulin resistance of vessels. HYPOTHESIS AND OBJECTIVES Insulin resistance can be induced in palmar digital vessels by continued exposure to insulin in vitro. The objective was to evaluate this in vitro model for future studies. METHODS Palmar digital vessel segments were collected immediately after euthanasia from horses with normal insulin/glucose blood values. Four arterial and 4 venous rings (3 mm wide) were prepared and each ring mounted in a tissue bath, containing Tyrodes solution at 37°C, 2 g tension was applied and the rings allowed to equilibrate for 45 min. Of the 4 rings of each vessel type, one was used as a control. One each of the remaining 3 rings was used for incubation with insulin (to induce resistance), wortmannin (to block PI3-kinase) and PD-098059 (to block MAP-kinase), respectively, for 30 min. After the incubation period, the rings were contracted with phenylephrine. When the response reached a plateau, a single dose of insulin was added to the baths and the response of each ring monitored for 30 min. RESULTS Insulin relaxed the control rings and those treated with PD 098059 but contracted those pretreated with insulin and wortmannin. Normal relaxation responses of the rings were converted to contractions by insulin resistance. Insulin resistance was confirmed by the qualitative response of insulin-incubated and wortmannin-incubated rings. CONCLUSIONS This study demonstrated successful induction of insulin resistance in both arterial and venous rings. It also suggested that the MAP-kinase pathway plays a minor role in controlling vasomotor tone under normal physiological conditions. POTENTIAL RELEVANCE The study suggests that the induction of insulin resistance in equine palmar digital vessel rings is reliable and provides a good in vitro model for studying the vascular insulin resistance which may occur in equine laminitis.

Effect of potential therapeutic agents in reducing oxidative stress in pulmonary tissues of recurrent airway obstruction-affected and clinically healthy horses.

REASONS FOR PERFORMING STUDY To determine and compare the reactive oxygen and nitrogen species (ROS and RNS) in pulmonary tissues of horses affected with recurrent airway obstruction (RAO) and clinically healthy horses, and to evaluate the effectiveness of potential therapeutic agents in reducing ROS and RNS in the tissues of these horses. OBJECTIVES We hypothesised that RAO-affected horses would have high levels of reactive species and that the test agents would reduce them. The objectives were as follows: 1) to determine the level of ROS and RNS in pulmonary tissues (bronchial and arterial rings) of RAO-affected and clinically healthy horses; and 2) to determine the ability of pentoxifylline, pyrrolidine-dithiocarbamate and a combined use of endothelin A and B receptor antagonists (BQ123 and BQ788, respectively) in reducing reactive species. METHODS Arterial and bronchial rings were collected from the diaphragmatic lung lobe of each horse immediately after euthanasia. The levels of ROS and RNS were measured in control tissues and those incubated with test agents, using an electron paramagnetic resonance instrument. RESULTS The levels of ROS and RNS were significantly greater in arterial and bronchial tissues of RAO-affected than of clinically healthy horses. Pentoxifylline and endothelin antagonists reduced both ROS and RNS in tissues from RAO-affected horses. Basal levels of reactive species in clinically healthy horses were not affected by these agents. No difference in the level of reactive species was observed between arterial and bronchial tissues. CONCLUSIONS Horses affected by RAO had higher ROS and RNS than clinically healthy horses. Pentoxifylline and endothelin antagonists effectively reduced ROS and RNS in pulmonary tissues of RAO-affected horses. POTENTIAL RELEVANCE The study suggested a potential use for pentoxifylline and endothelin antagonists in treating RAO-affected horses. As endothelin is involved in physiological functions, therapeutic use of its antagonists is cautioned.

Multixenobiotic resistance mechanism in the catfish intestine

Abstract A mechanism similar to the multidrug resistance phenomenon displayed by mammalian tumor cells has been identified in a variety of lower species, and in a variety of tissues, including the gastrointestinal tract. Xenobiotic efflux transport activity, and the location in membranes of the gastrointestinal tract luminal surfaces, indicates that P-glycoprotein (pgp) may act in modulating the bioavailability of dietary xenobiotics in aquatic animals. The objectives of this study were to investigate the dietary inducibility, depuration, and transport abilities of pgp in the catfish intestine. For this purpose, catfish underwent dietary induction with benzo(a)pyrene, β-naphthoflavone, and 3,4,3′,4′-tetrachlorobiphenyl. Plasma membrane vesicles preparations were used for protein quantitation, sodium dodecyl sulfate-polyacrylamide gel electrophoresis, western blot, and immunohistochemical detection with C219 monoclonal antibody. Transport activity was tested using [3H]-vinblastine (VBL) as a substrate. Results have shown that a catfish intestinal protein which cross-reacts with C219 antibody is inducible following dietary xenobiotic exposure, and that lower levels of this protein are still present 16 days after termination of exposure. In addition, VBL transport was shown to be ATP dependent and inhibited by verapamil. The location, function and responsiveness of pgp to dietary inducers suggest that pgp may play a role in the bioavailability and disposition of dietary xenobiotics.

Responses of Guinea-Pig Lung Parenchymal Strips to Tracheobronchial Lavage Fluid from Horses affected with Summer Pasture-Associated Obstructive Pulmonary Disease

The response of parenchymal strips from guinea-pig lungs to tracheobronchial lavage fluid (TBLF) collected from 8 normal horses and from 8 affected with summer pasture-associated obstructive pulmonary disease (SPAOPD) was determined. TBLF was collected during the summer (July) and winter (February) seasons. The serum/TBLF urea nitrogen ratio was used to standardize the mediator concentration in the TBLF. Four strips were used from each guinea-pig. The first strip did not receive any antagonist and served as the control. The second, third and fourth strips received antagonists of PGE2, LTD4 and PAF, respectively at 10–6 mol/L for 30 min. The tissues were then precontracted with a dose of histamine (10–5 mol/L) and their responses to 1 ml of TBLF were determined. The study showed that TBLF obtained in the summer from unaffected horses produced a significantly greater relaxation than that from the affected horses, whereas TBLF obtained in the winter from unaffected or affected horses did not cause a significantly different degree of relaxation. Among the antagonist-treated strips, only those exposed to the PGE2 blocker showed a significant reduction in the relaxation caused by TBLF obtained in the summer from SPAOPD horses. This suggests that PGE2 is an important mediator present in the summer in the TBLF from horses affected with SPAOPD.

Sedative and analgesic effects of medetomidine in beagle dogs infected and uninfected with heartworm

The sedative and analgesic effects of medetomidine were evaluated in heartworm-infected (HW+) and uninfected (HW–) beagle dogs by intravenous (IV) and intramuscular (IM) administration of 30 µg/kg and 40 µg/kg doses, respectively. Posture, response to noise and the pedal reflex were monitored. A procedure for mock radiographic positioning was performed to evaluate its overall clinical use. Observation times were 0, 15, 30, 60, 90, 120 and 180 min. In addition, the times from injection until the dog could not stand on its feet (down time), from lateral to sternal recumbency (sternal recumbency time), and from sternal recumbency to rising again (rising time) were also noted.Medetomidine produced rapid sedation and analgesia by both routes. Down times for the IM and IV routes were similar, which verified the manufacturers recommended doses. The HW+ dogs had shorter down times, probably owing to increased blood flow to the brain caused by adrenergic alpha-2 activity. Sternal recumbency and rising times did not differ between the groups, suggesting a similar metabolism. Sedation and analgesia were adequate for performing the procedure in all dogs. HW– dogs showed less resistance to handling during the procedure than HW+ dogs. Overall, medetomidine seems to be a suitable agent for short-term chemical restraint in dogs, even with subclinical heartworm infestation.

Bioavailability and biotransformation of 3,4,3′,4′-tetrachlorobiphenyl (TCB) in in situ intestinal preparations of uninduced and TCB induced channel catfish

Abstract Diet is a major route of exposure of polychlorinated biphenyls for aquatic animals living in contaminated environments. This study investigated systemic bioavailability and intestinal metabolism of [ 14 C]- 3,4,3′,4′-tetrachlorobiphenyl (TCB) in channel catfish, relative to TCB micelle solubility and TCB preexposure, using in vitro methods and in situ isolated intestinal preparations. Results showed that micelle fatty acid composition affected TCB solubilization in vitro, as well as systemic bioavailability of [ 14 C]-TCB delivered in situ. Systemic bioavailability of [ 14 C]-TCB was reduced by TCB dietary preexposure at two doses. TCB was scantily metabolized in the catfish intestine to 2-OH-TCB in the in situ preparation. High-dose TCB preexposure compositely induced aryl hydrocarbon hydroxylase (AHH) activity compared to other treatments, but had no significant effect upon intestinal cytochrome P450 content and glutathione-S-transferase (GST) activity. These results suggest that TCB partitioning and bioavailability may be influenced by dietary fatty acid composition and TCB preexposure. Furthermore, these results indicate that the intestine can minimally influence the metabolite profile of absorbed TCB.