The effect of P‐Glycoprotein on methadone hydrochloride flux in equine intestinal mucosa

Linardi, R. L., Stokes, A. M., Andrews, F. M. The effect of P‐Glycoprotein on methadone hydrochloride flux in equine intestinal mucosa. J. vet. Pharmacol. Therap.  36 , 43–50. Methadone is an effective analgesic opioid that may have a place for the treatment of pain in horses. However, its absorption seems to be impaired by the presence of a transmembrane protein, P‐glycoprotein, present in different tissues including the small intestine in other species. This study aims to determine the effect of the P‐glycoprotein on methadone flux in the equine intestinal mucosa, as an indicator of in vivo drug absorption. Jejunum tissues from five horses were placed into the Ussing chambers and exposed to methadone solution in the presence or absence of Rhodamine 123 or verapamil. Electrical measurements demonstrated tissue viability for 120 min, and the flux of methadone across the jejunal membrane (mucosal to submucosal direction) was calculated based on the relative drug concentration measured by ELISA. The flux of methadone was significantly higher only in the presence of verapamil. P‐glycoprotein was immunolocalized in the apical membrane of the jejunal epithelial cells (enterocytes), mainly located in the tip of the villi compared to cells of the crypts. P‐glycoprotein is present in the equine jejunum and may possibly mediate the intestinal transport of methadone. This study suggests that P‐glycoprotein may play a role in the poor intestinal absorption of methadone in vivo.     

The effects of Strongylus vulgaris parasitism on eosinophil distribution and accumulation in equine large intestinal mucosa

REASONS FOR PERFORMING STUDY: Eosinophilic granulocytes have been associated with parasite or immune-mediated diseases, but their functions in other disease processes remain unclear. Cause and timing of eosinophil migration into the equine gastrointestinal mucosa are also unknown. OBJECTIVE: To determine the effects of intestinal parasitism on eosinophils in equine large intestinal mucosa. METHODS: Large intestinal mucosal samples were collected from horses and ponies (n = 16) from the general veterinary hospital population, ponies (n = 3) raised in a parasite-free environment, ponies experimentally infected with 500 infective Strongylus vulgaris larvae and treated with a proprietary anthelmintic drug (n = 14), and a similar group of ponies (n = 7) that received no anthelmintic treatment. Total eosinophil counts and eosinophil distribution in the mucosa were determined by histological examination. A mixed model analysis was performed and appropriate Bonferroni adjusted P values used for each family of comparisons. P<0.05 was considered significant. RESULTS: There was no difference in large intestinal mucosal eosinophil counts and eosinophil distribution between ponies infected with S. vulgaris and those raised in a parasite-free environment. Experimental infection with S. vulgaris, with or without subsequent anthelmintic treatment, did not change eosinophil counts, and counts were similar to those for horses from the general population. CONCLUSIONS: Migration of eosinophils to the equine large intestinal mucosa appears to be independent of exposure to parasites. Large intestinal mucosal eosinophils may have more functions in addition to their role in defence against parasites.     

云南地区马流感病毒检测及其HA基因序列分析

马流感能引起马属动物的高度接触性呼吸道传染病。该病毒已知仅存在H7N7(马流感病毒1型)、H3N8(马流感病毒2型)两个亚型,目前仅H3N8亚型毒株在全球范围内引起马流感流行和暴发。采用A型及H3亚型特异性RT-PCR技术,检测云南疑似马流感病例鼻腔棉拭子样品;HA基因PCR扩增产物经纯化后,克隆至pMD18-T载体进行测序,并与已知代表性毒株对应序列进行比对及系统发育分析。研究发现云南马流感病毒与已知代表毒株血凝素基因核苷酸及氨基酸序列同源性分别介于91.2%~99.2%和89.5%~98.4%,属于美洲谱系(American lineage)佛罗里达亚谱系(Florida sub-lineage)毒株。     

Testicular ischaemic necrosis as a cause of equine cryptorchidism

Cryptorchidism is a fairly common pathology presented to equine surgical facilities with the cryptorchid testicle most commonly located in the abdomen or ipsilateral inguinal canal. The causes of cryptorchidism are not known, but testicular abnormalities have been suggested. Monorchidism as a cause of maldescent of one or both testicles is rare and is hypothesised to be the result of a vascular insult, similar to testicular regression in man. This case report details laparoscopic abdominal exploration of a cryptorchid horse and identification of an abnormal testicular remnant affected by ischaemic necrosis.     

Impedance monitoring of equine intestinal motility

Myoelectrical and myomechanical activities of the distal portion of the jejunum and pelvic flexure were studied in 7 ponies, using permanently implanted monopolar and bipolar stainless steel electrodes. Dental acrylic embedded recording electrodes were surgically sutured to the serosal surface of the distal portion of the jejunum and pelvic flexure. Myoelectrically, regular spike bursts and irregular spike bursts were observed in the jejunum. Short spike bursts and long spike bursts were recorded and associated with spike potentials, using impedance recording techniques. Electrical and mechanical data could be monitored simultaneously from the same recording electrode, using separate channels on a physiograph. This method proved effective to monitor periodically myomechanical activity at the same time that myoelectrical activity was being evaluated. The recording system required fewer recording devices to be attached to the intestinal tract, was an inexpensive method of obtaining myomechanical recordings, and did not alter markedly the myoelectrical activity when mechanical activity was being monitored.     

In vitro culture of equine respiratory mucosa explants

An in vitro model of the upper respiratory tract of the horse was developed to investigate mechanisms of respiratory diseases. Four tissues of the upper respiratory tract of three horses were collected. Explants were maintained in culture at an air–liquid interface for 96 h. At 0, 24, 48, 72 and 96 h of cultivation, a morphometric analysis was performed using light microscopy, scanning electron microscopy and transmission electron microscopy. The explants were judged on morphometric changes of epithelium, basement membrane and connective tissue. Viability was evaluated using a fluorescent Terminal deoxynucleotidyl transferase-mediated dUTP Nick End Labelling (TUNEL) staining. No significant changes in morphometry and viability of any of the explants were observed during cultivation. Hence, the in vitro model may be useful to study infectious and non-infectious diseases at the level of the equine respiratory tract, with potential application to the development of vaccines and treatments for diseases of the respiratory tract.     

The use of staplers in equine intestinal surgery

Automatic suturing instruments were tested in two ponies and two horses. The TA stapler was used for bowel closure (enterotomies and gut transection) and for end-to-end 'triangulation' anastomoses. The GIA stapler was used for making side-to-side anastomoses. In one horse several enterotomies were made at different stages and the results of suturing by hand and with a stapler were compared. The utility of both types of staplers is discussed in relation to clinical evaluations and macroscopical and microscopical findings post mortem.     

Attenuation of ischaemic injury in the equine jejunum by administration of systemic lidocaine

REASONS FOR PERFORMING STUDY: Absorption of endotoxin across ischaemic-injured mucosa is a major cause of mortality after colic surgery. Recent studies have shown that flunixin meglumine retards mucosal repair. Systemic lidocaine has been used to treat post operative ileus, but it also has novel anti-inflammatory effects that could improve mucosal recovery after ischaemic injury. HYPOTHESIS: Systemic lidocaine ameliorates the deleterious negative effects of flunixin meglumine on recovery of mucosal barrier function. METHODS: Horses were treated i.v. immediately before anaesthesia with either 0.9% saline 1 ml/50 kg bwt, flunixin meglumine 1 mg/kg bwt every 12 h or lidocaine 1.3 mg/kg bwt loading dose followed by 0.05 mg/kg bwt/min constant rate infusion, or both flunixin meglumine and lidocaine, with 6 horses allocated randomly to each group. Two sections of jejunum were subjected to 2 h of ischaemia by temporary occlusion of the local blood supply, via a midline celiotomy. Horses were monitored with a behavioural pain score and were subjected to euthanasia 18 h after reversal of ischaemia. Ischaemic-injured and control jejunum was mounted in Ussing chambers for measurement of transepithelial electrical resistance (TER) and permeability to lipopolysaccharide (LPS). RESULTS: In ischaemic-injured jejunum TER was significantly higher in horses treated with saline, lidocaine or lidocaine and flunixin meglumine combined, compared to horses treated with flunixin meglumine. In ischaemic-injured jejunum LPS permeability was significantly increased in horses treated with flunixin meglumine alone. Behavioural pain scores did not increase significantly after surgery in horses treated with flunixin meglumine. CONCLUSIONS: Treatment with systemic lidocaine ameliorated the inhibitory effects of flunixin meglumine on recovery of the mucosal barrier from ischaemic injury, when the 2 treatments were combined. The mechanism of lidocaine in improving mucosal repair has not yet been elucidated.     

In vitro transport of L-alanine by equine cecal mucosa

When sheets of mucosa from the cecum of clinically normal horses were incubated in vitro with radiolabeled L-alanine, they could accumulate this amino acid against an apparent concentration gradient after 60 to 150 minutes of incubation. The active transport system for L-alanine was on the serosal surface of the mucosal sheet only. L-Alanine accumulation at 60 minutes was partly inhibited by 20 mM glycine (P less than 0.01), 0.5 mM ouabain (P less than 0.05), and Na deprivation (P less than 0.02). Anoxia for 60 minutes increased L-alanine accumulation, but had adverse effects on cell structure and intracellular cation distributions. Transmucosal fluxes induced a small, but significant (P less than 0.05), net secretion of L-alanine, and the mean (+/- SEM) transmucosal potential difference was 7.3 +/- 0.7 mV over the period of flux measurement. It was concluded that L-alanine was accumulated by the serosal surface of the cecal mucosa, possibly to provide substrate for tissue metabolism. There was no evidence that the cecal mucosa could actively transport this amino acid from the luminal bathing medium.     

In vitro transport of cycloleucine by equine cecal mucosa

Mucosa obtained from the cecum of healthy horses and incubated in vitro with 0.1 mM cycloleucine could accumulate this amino acid against an apparent concentration gradient after 60 and 120 minutes. Accumulation by the serosal (antiluminal) surface of the tissue was 3 times greater than accumulation by the mucosal (luminal) surface after 120 minutes (P less than 0.001). Cycloleucine accumulation was significantly reduced by Na deprivation after 60 minutes (P less than 0.05) and 120 minutes (P less than 0.01) and by anoxic conditions after 120 minutes (P less than 0.05). Transmucosal flux from mucosal to serosal surface of the tissue was significantly (P less than 0.05) greater than the opposing flux, but both unidirectional fluxes were small and were largely attributed to passive processes. It was concluded that the most avid transport system for cycloleucine was on the serosal surface of the horse's cecal mucosa, and an active transport system was not evident on the mucosal surface. An active transport system for amino acids on the serosal surface could be explained by the need for crypt cells, the predominant epithelial cell type in the cecum, to obtain nutrients from blood, rather than from the intestinal lumen.     

The teniae of the equine intestinal tract.

At several locations along the equine cecum and colon, the outer longitudinal portion of the tunica muscularis is gathered into discrete bands of smooth muscle and connective tissue called "teniae". In this study, the disposition of the teniae ceci and coli was traced along the equine intestinal tract. It was discovered that, in several instances, arrays of teniae converge toward the valves and sphincters which separate the various intestinal compartments. The teniae may also provide support for and directionality to, peristaltic contraction waves. The tissue proportions of the teniae vary in different locations. The tenia libera lateralis of the ventral colon is rich in elastic connective tissue, while that of the right dorsal colon is primarily composed of smooth muscle. This may reflect the different responsibilities of these two compartments. The teniae are innervated and their smooth muscle cells are joined by many gap junctions. The connective tissue constituents afford intestinal support while yielding to intestinal distension. The smooth muscle and neural elements may foster active tenial participation in peristalsis. This premise must be tested by electrophysiological experimentation. Further experimentation is also necessary to ascertain whether injury to the teniae might predispose a horse to colic.     

Detection of endotoxin in cases of equine colic

The Limulus amoebocyte lysate assay was used to test for the presence of endotoxin in 37 clinical cases of equine colic. Positive plasma titres were detected in 10 cases and the presence of endotoxin was significantly correlated with a high heart rate, a high packed cell volume and a poor prognosis. High levels of endotoxin were detected in gut contents taken from several sites in the gastrointestinal tract of normal horses.     

Orally administered phenylbutazone causes oxidative stress in the equine gastric mucosa

Phenylbutazone (PBZ) is widely used in equine medicine, and its side effects on the gastrointestinal tract are well known. The inhibition of prostaglandins and the oxidative stress induced by nonsteroidal anti‐inflammatory drugs (NSAIDs) are described as mechanisms of gastric mucosal injury in humans. In horses, only the secondary effect of changes in cyclooxygenases is related to gastric mucosal injury. The objective of this study was to evaluate the effect of PBZ on certain antioxidative/oxidative parameters of the gastric mucosa. The concentrations of antioxidants and oxidants (superoxide dismutase, SOD; catalase, CAT; nitric oxide, NO; total glutathione, GSH; myeloperoxidase, MPO; and malondialdehyde, MDA), PGE₂ levels, and the ulcerative lesions score were assessed. The results demonstrated decreased levels of antioxidant variables, increased levels of oxidant variables, and alterations in the prostaglandin E₂ (PGE₂), myeloperoxidase (MPO), and glutathione (GSH) levels. In conclusion, PBZ induces oxidative stress in the gastric glandular mucosa of horses by changing the antioxidant–oxidant balance of this surface, which might be regarded as another mechanism of injury in the horse stomach.     

Bacteriologic examination of equine fecal flora as a diagnostic tool for equine intestinal clostridiosis

The fecal flora of 56 clinically healthy and 23 sick horses were examined bacteriologically for counts of Clostridium perfringens, molds, coliforms, alpha- and beta-hemolytic streptococci, and microbes belonging to genus Bacillus, as well as for the presence of Salmonella spp. Of the healthy horses, 85.7% had a C perfringens count less than 10(1) colony-forming units/g of feces. Of the healthy horses, lowest counts were found in race-horses. Of the sick horses, equine intestinal clostridiosis was diagnosed in 2 horses with large C perfringens counts (10(4) to 10(7) colony-forming units/g) and with acute diarrhea. The 7 isolates of C perfringens were identified as serotype A. Salmonella spp were not detected from any of the horses. The study indicated that diagnosing equine intestinal clostridiosis based on the determination of the fecal C perfringens count was suitable.     

Factors for prognostic use in equine obstructive small intestinal disease

Twenty horses with small intestinal obstructions requiring surgery were evaluated prospectively. Ten horses lived (group 1) and 10 died (group 2). Eight of the horses in group 1 had simple obstruction and 7 of the horses in group 2 had strangulation obstruction. There was a significant difference (P less than 0.001) between the mean intraluminal hydrostatic pressure in horses of groups 1 and 2 (6.3 cm H2O and 15 cm H2O, respectively). The mean peritoneal fluid protein concentration in horses of groups 1 and 2 (2.8 mg/dl and 5.4 mg/dl, respectively) also differed significantly between groups (P less than 0.01). Histologic evaluation of the intestinal specimens from horses of group 1 (n = 3) and group 2 (n = 6) revealed more severe mucosal lesions in group 2. The measured values that were not significantly different between the 2 groups included PCV, total serum protein content, WBC count, anion gap, and duration of colic before admission. It was concluded that peritoneal fluid protein concentration and intraluminal hydrostatic pressure in small intestinal obstruction may be used as adjuncts to diagnosis and as prognostic indicators.     

Distribution of the neurokinin-1 receptor in equine intestinal smooth muscle

REASON FOR PERFORMING STUDY: Tachykinins have profound effects on equine intestinal motility, but the distribution of the neurokinin receptors (NKRs) through which they act is unknown. This study reports the distribution of one of these receptors, the neurokinin-1 receptor (NK1R), in smooth muscle throughout the equine intestinal tract. OBJECTIVES: To quantify the distribution of the NK1R, based upon mRNA expression, in smooth muscle of different regions of the equine intestinal tract. METHODS: Nine regions of the intestinal tract were sampled in 5 mature horses. Total RNA was isolated from smooth muscle and reverse transcribed; NK1R mRNA was then quantified using real-time PCR. RESULTS: NK1R mRNA was found at all levels of the sampled intestinal tract. The smooth muscle of the proximal small intestine and the ventral colon exhibited the highest level of NK1R mRNA expression in the equine intestinal tract. CONCLUSIONS: Tachykinins probably affect intestinal contractility and propulsion in the proximal small intestine and in the ventral colon.     

早期断奶仔猪肠黏膜的营养调控

肠黏膜的发育与成熟状况是制约幼畜快速生长的关键因素。早期断奶仔猪肠黏膜免疫力较低,容易遭受饲粮抗原的刺激而引起肠黏膜过敏性损伤,诱发仔猪断奶后腹泻。通过肠黏膜营养调控技术,补充肠黏膜能量营养, 促进肠黏膜的发育与成熟,修补断奶引起的肠黏膜损伤,可以从根本上减轻仔猪断奶后腹泻的发生。本文对早期断奶引起的肠黏膜形态变化以及有利于修复受损伤肠黏膜的营养物和调控因子进行了综述。     

Characterization of xenobiotic metabolizing enzymes in bovine small intestinal mucosa

Virkel, G., Carletti, M., Cantiello, M., Della Donna, L., Gardini, G., Girolami, F., Nebbia, C. Characterization of xenobiotic metabolizing enzymes in bovine small intestinal mucosa. J. vet. Pharmacol. Therap. 33 , 295–303. The intestinal mucosa plays a capital role in dictating the bioavailability of a large array of orally ingested drugs and toxicants. The activity and the expression of several xenobiotic metabolizing enzymes were measured in subcellular fractions from the duodenal mucosa of male veal calves and beef cattle displaying a functional rumen but differing in both age (about 8 months vs. 18 to 24 months) and dietary regimens (i.e., milk replacer plus hay and straw vs. corn and concentrated meal). Intestinal microsomes showed cytochrome P450 (CYP) 2B, 2C‐ and 3A‐mediated activities and the presence of the corresponding immunorelated proteins, but no proof of CYP1A expression and/or functions could be provided. Intestinal microsomes were also active in performing reactions typically mediated by carboxylesterases (indophenylacetate hydrolysis), flavin‐containing monooxygenases (methimazole S‐oxidation), and uridindiphosphoglucuronyltransferases (1‐naphthol glucuronidation), respectively. Cytosolic fractions displayed the glutathione S‐transferase (GST)‐dependent conjugation of 1‐chloro‐2,4‐dinitrobenzene; besides, the GST‐mediated conjugation of ethacrinic acid (GSTπ) or cumene hydroperoxide (GSTα) was matched by the presence of the corresponding immunorelated proteins. Conversely, despite the lack of measurable activity with 3,4‐dichloronitrobenzene, a protein cross reacting with anti‐rat GSTμ antibodies could be clearly detected. Although, as detected by densitometry, CYPs and GST isoenzymes tended to be more expressed in beef cattle than in veal calf preparations, there was a general poor correlation with the rate of the in vitro metabolism of the selected diagnostic probes.