Permeability In Dogs Research Articles

PSXIII-7 Effects of high fat diets on metabolic parameters and markers of colonic permeability in dogs

Abstract Obesity and nutritional factors like high-fat diets (HFD) are associated with metabolic alterations, which result from the diet, bodyweight change or both. The objective was to determine the effects of HFD consumed either at maintenance or excess energy levels on BW, metabolic parameters, and colonic permeability markers. A HFD (35% fat as fed, 4923 kcal/kg) was given for 8wks to 24 healthy Beagle dogs, 8 at 100% (G1.0) and 16 at 150% (G1.5) maintenance energy levels for each individual dog. At baseline and after 8wks, basal leptinemia, ghrelinemia, glycemia and insulinemia were measured, while postprandial glycemia and insulinemia were determined for 6h following a challenge test meal. Colonic mucosa biopsies were performed and the expression of proteins involved in permeability was analyzed using RT-rtPCR. Results were analyzed using linear mixed model (nlme package; R) with dog as a random term (data rank-transformed where residuals not normally distributed). Ethics approval was granted by the French Ministry of Research and Royal Canin’s Ethical Committee. In the G1.5 group, increases in bodyweight (15%; P < 0.001), leptinemia (5±2 vs 2±1 ng/ml; P < 0.01), and postprandial insulin AUC (10405±2316 vs 6232±1368 μU.min.ml-1; P = 0.03) were observed at 8wks compared to baseline, whereas ghrelinemia decreased (552±361 vs 633±413 pg/ml; P = 0.04). No such change were observed in the G1.0 group. In both groups at 8wks, mRNA levels of claudin 2 (G1.0: P < 0.05; G1.5: P < 0.01) and syndecan (G1.0: P < 0.01; G1.5: P < 0.01) were higher compared to baseline, while mRNA levels of TJP1 (P < 0.01) increased in the G1.5 group. The changes observed in the G1.5 group but not in the G1.0 group suggest that the metabolic and hormonal parameters are related to weight gain but not to HFD alone. Furthermore, the mechanisms behind the modifications to colonic permeability markers observed in both groups remains to be elucidated.

Regional Intestinal Permeability in Dogs: Biopharmaceutical Aspects for Development of Oral Modified-Release Dosage Forms.

The development of oral modified-release (MR) dosage forms requires an active pharmaceutical ingredient (API) with a sufficiently high absorption rate in both the small and large intestine. Dogs are commonly used in preclinical evaluation of regional intestinal absorption and in the development of novel MR dosage forms. This study determined regional intestinal effective permeability (Peff) in dogs with the aim to improve regional Peff prediction in humans. Four model drugs, atenolol, enalaprilat, metoprolol, and ketoprofen, were intravenously and regionally dosed twice as a solution into the proximal small intestine (P-SI) and large intestine (LI) of three dogs with intestinal stomas. Based on plasma data from two separate study occasions for each dog, regional Peff values were calculated using a validated intestinal deconvolution method. The determined mean Peff values were 0.62, 0.14, 1.06, and 3.66 × 10(-4) cm/s in the P-SI, and 0.13, 0.02, 1.03, and 2.20 × 10(-4) cm/s in the LI, for atenolol, enalaprilat, metoprolol, and ketoprofen, respectively. The determined P-SI Peff values in dog were highly correlated (R(2) = 0.98) to the historically directly determined human jejunal Peff after a single-pass perfusion. The determined dog P-SI Peff values were also successfully implemented in GI-Sim to predict the risk for overestimation of LI absorption of low permeability drugs. We conclude that the dog intestinal stoma model is a useful preclinical tool for determination of regional intestinal permeability. Still, further studies are recommended to evaluate additional APIs, sources of variability, and formulation types, for more accurate determination of the dog model in the drug development process.

Estimation of intestinal permeability in healthy dogs using the contrast medium iohexol

Iohexol is a nonradioactive marker that has been used successfully to test intestinal permeability in humans with inflammatory bowel disease. There is evidence in dogs that iohexol shares a similar permeability pathway as (51)chromium-EDTA, the gold standard marker. The objective of this study was to determine an optimal oral iohexol dosage for an intestinal permeability serum test (IPST) and to use the test to estimate intestinal permeability in healthy dogs. Eight clinically healthy dogs free of gastrointestinal, liver, and pancreatic disease were used in the study. Dosages of 0.25, 0.5, 1.0, 2.0, and 4.0 mL/kg of Omnipaque-350 (iohexol) were administered to 2 dogs at weekly intervals. Iohexol concentration was determined in serum samples obtained hourly for 6 hours after administration by high-performance liquid chromatography. Using the optimal dosage, iohexol was administered to 8 dogs twice, 6-36 days (mean 10 days) apart, and coefficients of variation (CVs) for iohexol concentration were calculated. A dosage of 2.0 mL/kg was chosen as optimal for the IPST, based on ease of iohexol detection in serum, intestinal contrast, and clinical effects of iohexol. Following administration of this dose to healthy dogs, mean (+/-SD) serum iohexol concentrations were 8.74+/-4.38, 11.89+/-5.67, 12.40+/-5.47, 9.23+/-5.54, 7.61+/-5.13, and 5.27+/-2.67 microg/mL at 1, 2, 3, 4, 5, and 6 hours after iohexol administration, respectively. CVs between the 2 test days were 28-45%. Using the iohexol dosage established in this study, the iohexol IPST was easy to perform as a marker for intestinal permeability in dogs. Further studies to establish reference intervals and evaluate the diagnostic value of the iohexol IPST in dogs with gastrointestinal disease are warranted.

Colonic permeability is higher in Great Danes compared with smaller breed-dogs

Fed the same dry diet, large dogs show poorer fecal quality than small ones. A high colonic permeability could explain a low water and electrolyte net balance leading to high fecal water content. This experiment was conducted to evaluate colonic permeability in dogs varying in body size and to determine whether colonic permeability is related to fecal sodium concentration and fecal quality. Four breeds of dogs were used: six Miniature Poodles (MP), six Standard Schnauzers (SS), six Giant Schnauzers (GS) and six Great Danes (GD). Colonic permeability was evaluated using the ratio of urinary lactulose to sucralose (L:S) after oral administration. Fecal sodium concentration was measured by flame photometry. The urinary L:S ratio was significantly lower in GD, indicating a higher colonic permeability, than in the three other breeds (0.35 ± 0.12 for GD and 0.51 ± 0.05 for MP). GD also presented the higher fecal sodium concentrations and the poorest fecal quality. The higher fecal sodium concentration observed in GD could be explained by the higher colonic permeability and both these variables could be important explanations for higher fecal moisture in large dogs.

Measurement of Intestinal Mucosal Permeability in Dogs with Lymphocytic-Plasmacytic Enteritis

Lymphocytic-plasmacytic enteritis (LPE) is a type of canine inflammatory bowel disease (IBD). One of its most probable causes is a defect in the mucosal permeability barrier. In the present study, intestinal permeability in LPE dogs was examinated to evaluate its clinical value. Twenty-nine dogs with LPE diagnosed by clinical and histological examinations were included in this study. Intestinal permeability was evaluated by measuring the ratio of the concentrations of two sugars (lactulose (L) and rhamnose (R)) with different molecular weights in urine samples after oral administration of a solution containing them. Biopsy specimens of duodenum were evaluated according to histological criteria. The urinary L:R ratio in the 29 LPE dogs (1.68 +/- 1.17, mean +/- SD) was significantly higher than that in the 10 healthy control dogs (0.75 +/- 0.38, P<0.01). In the LPE dogs, a weak correlation was observed between the histopathological grading score of the duodenum and the urinary L:R ratio (r=0.408, P<0.05). The urinary L:R ratio in the 20 dogs showing hypoalbuminemia (< 2.5 g/dl) was significantly higher than that in the 9 dogs with normal serum albumin levels > 2.5 g/dl (P<0.01). In conclusion, permeability of the intestinal mucosa as determined by the urinary L:R ratio could be a useful laboratory parameter for evaluating intestinal damage in LPE dogs.

51Cr‐EDTA Absorption Blood Test: An Easy Method for Assessing Small Intestinal Permeability in Dogs

The 51Cr‐EDTA test is a valuable clinical tool for screening intestinal diseases in dogs. The test is performed by calculating the percentage of recovery from urine of a PO‐ingested dose of 51Cr‐EDTA after 6 or 24 hours. Careful urine collection is a practical limitation of this test in dogs, and our goal was to develop a simpler test that measures 51Cr‐EDTA in blood. A 51Cr‐EDTA absorption test was simultaneously performed on urine and serum 43 times in healthy Beagle Dogs. Timed blood samples were withdrawn, and urine was collected during a 6‐hour period. Percentages of the ingested dose were then calculated in urine and serum. The mean ± standard deviation (range) percentage in urine after 6 hours was 14.07 ± 8.72% (3.81‐34.18%), whereas results in serum from samples taken at 2, 3, 4, 5, and 6 hours were 0.49 ± 0.45% (0.02‐2.13%), 0.75 ± 0.52% (0.03‐1.89%), 0.82 ± 0.57% (0.13‐2.21%), 0.70 ± 0.53% (0.12‐1.99%), and 0.47 ± 0.44% (0.11‐1.79%), respectively. The results for blood specimens showed good concordance with those for urine, especially for the samples taken at 4 hours (r= 0.89). Moreover, the correlation between urine and blood was better when the sum of the percentages of the recovered analyte from various blood samples was compared with urine. The correlation coefficient when summing 4 blood samples was excellent (r= 0.97) and remained excellent when summing only 2 blood samples taken at 3 and 5 hours (r= 0.95) or at 3 and 4 hours (r= 0.94). We conclude that a serum 51Cr‐EDTA test determined by summing successive blood samples provides an easier means of estimating small intestinal permeability in dogs and gives results comparable to those of the 6‐hour urine test.

Highly enhanced permeability of blood-brain barrier induced by repeated administration of endothelin-1 in dogs and rats.

The effects of intracisternal administration of endothelin-1 on blood-brain barrier permeability were examined in dogs and rats. Single doses of endothelin-1 elevated blood-brain barrier permeability about two times compared with control groups in both species. However, repeated dosing with endothelin-1 at a 24 hr intervals caused a highly enhanced disruption of blood-brain barrier permeability in dogs, but not in rats, whereas the repeated administration with a 48 hr interval markedly increased the blood-brain barrier permeability in both species of animals (dogs: 923%, rats: more than 661%). Moreover, this abnormally enhanced permeability of blood-brain barrier in dogs was completely blocked by pretreatment with the endothelin ET-A receptor selective antagonist, S-0139, administered prior to either the first or second dosing with endothelin-1. From these results, we conclude that a repeated attack of endothelin-1 from the adventitial site of brain blood vessels produces a severe disruption of blood-brain barrier permeability through an endothelin ET-A receptor-mediated process.

Milrinone improves arterial oxygenation in dogs with acute lung injury induced by oleic acid.

The aim of the study was to investigate effects of milrinone on pulmonary permeability in dogs with acute lung injury induced by oleic acid. To induce acute lung injury, we administered 0.08 mg/kg of oleic acid to 19 adult mongrel dogs and then measured hemodynamic parameters and performed blood gas analysis. An injection of oleic acid depressed the mean arterial pressure, cardiac index, and arterial oxygenation. Dogs were divided into three groups: six received a bolus of milrinone (50 microg/kg) followed by a continuous (0.5 microg/kg/min, low-dose), seven received a bolus (100 microg/kg) followed by a continuous (1.0 microg/kg/min; i.e., a low-dose twice; high-dose), and six no milrinone (control). Milrinone administration improved the cardiac index and arterial oxygenation and simultaneously depressed the intrapulmonary shunt fraction and the extravascular thermal lung water as extravascular water content of the lung. These changes produced by milrinone are different according to the doses. In conclusion, milrinone acts on the capillary endothelium and inhibits an accumulation in the extravascular water content of the lung, which may induce an improvement in arterial oxygenation. Milrinone may also improve arterial oxygenation through an inhibition of platelet aggregation and chemical mediators that are released from platelets. The latter mechanism also may improve arterial oxygenation, and the exact property responsible for causing the effect of milrinone has not yet been identified.

Contribution of endothelin-1 to disruption of blood-brain barrier permeability in dogs.

We examined the effect of intracisternal application of endothelin-1 (ET-1) on the permeability of fluorescein into the cerebrospinal fluid (CSF) in beagle dogs in order to evaluate its role in disruption of blood-brain barrier (BBB) permeability seen in the subarachnoid hemorrhage animal model. Intracisternal application of their autologous blood for producing a canine two-hemorrhage model revealed an enhanced fluorescein permeability into the CSF together with the development of cerebral vasospasm. A single dose of ET-1 (40 pmol/animal) significantly increased penetration of fluorescein compared with that in normal dogs. Although its magnitude was much less than that in the two-hemorrhage model after the first administration of ET-1, the second challenge of the same dose of ET-1 with a 48-h interval produced marked disruption of BBB permeability similar to those in the animal model. Moreover, the ET-1-induced enhancement of fluorescein permeability into the CSF was completely prevented by intracisternal pretreatment with an endothelin ET(A)-receptor selective antagonist, S-0139 (0.03 mg/kg), as were the ET-1-induced cerebral vasoconstriction and behavioral changes as previously reported. Thus, we conclude that ET-1 acting on the adventitial site of brain in dogs contributes to the disruption of BBB permeability via endothelin ET(A)-receptor mediation.

Histamine-releasing properties of T-3762, a novel fluoroquinolone antimicrobial agent in intravenous use. II. Dermovascular permeability-increasing effect and action on peritoneal mast cells.

To predict the actions of T-3762, a newly developed fluoroquinolone antimicrobial agent, as well as ciprofloxacin (CPFX) and ofloxacin (OFLX), on injection sites when dosed parenterally, their ability to increase cutaneous vascular permeability in dogs and to release histamine from rat peritoneal mast cells was examined. CPFX and OFLX increased cutaneous vascular permeability in concentrations ranging from 16 to 32 microg/ml, while T-3762 was inactive at 2000 microg/ml. The vascular permeability-increasing activities of these drugs were inhibited efficiently by pretreatment with a combined dose of diphenhydramine and cimetidine. CPFX induced histamine release from rat mast cells in a dose-dependent manner, whereas T-3762 was ineffective. Therefore, it is concluded that fluoroquinolone antimicrobial agents may have the ability to cause an increase in cutaneous vascular permeability by releasing histamine from mast cells at the injection site when administered parenterally, and that T-3762 has minimum activity among the agents tested in this study.

The influence of total body hyperthermia on brain haemodynamics and blood-brain barrier in dogs.

This study was designed to examine the influence of total body hyperthermia (TBHT) using an extracorporeal circuit with a heat exchanger on the cerebral blood flow (CBF), intracranial pressure (ICP), brain tissue pH, cerebral autoregulation and blood-brain barrier (BBB) permeability in dogs. The rectal temperature of the dow was raised to 41.5 degrees C, maintained at 41.5-42.0 degrees C for 2 hours (HT period) and then reduced to normothermia by cooling. Regional CBF was measured by the hydrogen clearance method before heating, during the HT period and after cooling. ICP and brain tissue pH were monitored during the TBHT treatment. Autoregulation of the CBF during the HT period was assessed by measuring the regional CBF and the ICP in a state of induced hypo- or hypertension. The influence of TBHT on BBB permeability was examined using an immunohistochemical technique. The regional CBF increased from 38.1 +/- 6.5 (mean +/- SD) to 49.1 +/- 9.8 ml/100 g/min and the ICP from 10.3 +/- 4.2 to 16.8 +/- 3.4 mmHg when TBHT was raised. These returned to normal values after cooling. The regional CBF and the ICP changed in parallel with drug-induced changes of mean arterial blood pressure during the HT period. These changes suggest that autoregulation of the CBF is paralysed during the HT period. Brain tissue pH decreased rapidly when the rectal temperature exceeded 41.0 degrees C. The pH was 7.18 +/- 0.05 during the HT period and was relatively stable. The pH returned to a normal value after cooling. Immunopositive stain for albumin was not observed in heated brain tissue except for the normally leaky pineal gland and the choroid plexus, indicating preservation of BBB during TBHT. These results suggest that brain oedema may occur easily due to paralysed cerebral autoregulation when the arterial blood pressure fluctuates excessively, so arterial blood pressure must be controlled strictly during TBHT.

Effect of topically applied 2% pilocarpine and 0.25% demecarium bromide on blood-aqueous barrier permeability in dogs

Summary The effect of topically applied 2% pilocarpine and 0.25% demecarium bromide on aqueous humor flare was evaluated while treating normal eyes of dogs 3 times daily for 8 days. Fifteen clinically normal Beagles were allotted to 2 drug treatment groups, and flare was measured/using laser flaremetry. Pilocarpine caused an increase of flare to 167 photon counts (pc)/ms at 7 hours, compared with the nontreated control eye mean value 3.7 pc/ms. By 31 hours, flare had decreased to 70 pc/ms with 3 daily topical treatments. By 55 hours, the flare measurement was in the normal range. Intraocular pressure (iop) decreased in the treated eyes, compared with the non-treated eyes, and maximal decrease in iop was observed at the same time as maximal flare. Demecarium induced a similar increase in flare and decrease in iop, with peak effect seen at 31 hours. At that time, the flaremetry result was 28 pc/ms, compared with 3.4 pc/ms in the nontreated control eyes. After 8 days of 3 times daily treatment, flare values had returned to normal in eyes of dogs in both treatment groups. The transient increase in aqueous humor protein concentration did not result in ocular or visual damage, and long-term changes were not seen. The amount of flare increase attributable to topical application of pilocarpine was greater than the increase in human eyes when measured by use of laser flaremetry.

A novel HPLC method for the simultaneous quantification of monosaccharides and disaccharides used in tests of intestinal function and permeability

The quantification of monosaccharides and disaccharides used as probes in intestinal function and permeability tests can be technically demanding, detracting from the value of this approach to the indirect assessment of intestinal damage. In this study, a procedure is described for the simultaneous quantification of rhamnose, lactulose, 3-O- methyl- d-glucose and xylose in urine by HPLC using an anion exchange column with pulsed amperometric detection. This method is relatively fast and simple to perform, requiring no pre-treatment of urine samples or post-column derivatization. Accuracy and precision of determinations are illustrated by analytical recoveries (mean percentage ± S.D., C.V., n = 30) for multiple batch analyses of a diluted urine sample containing 20 mg/l of rhamnose (100 ± 6.8, 6.2%), lactulose (100 ±6.1, 5.5%), 3-O- methyl- d-glucose (98 ± 5.9, 5.5%) and xylose (104 ± 7.1, 6.5%). Linearity of standard curves indicated that the lower limit for accurate quantification was 0.1mg/l for all four sugars. Urinary recoveries following oral administration of these sugars to control dogs were determined as a baseline for the investigation of intestinal damage in this species and comparison of chromatograms illustrated enhanced permeability in dogs with gluten-sensitive enteropathy.

Enhanced intestinal permeability to 51Cr-labeled EDTA in dogs with small intestinal disease

Summary Intestinal permeability in dogs with small intestinal disease was measured by quantitation of 24-hour urinary excretion of 51Cr-labeled EDTA following intragastric administration. Permeability was high in dogs with a variety of naturally acquired small intestinal diseases including wheat-sensitive enteropathy of Irish Setters, small intestinal bacterial overgrowth, and giardiasis, and permeability was decreased after successful treatment. These findings indicate that the assessment of intestinal permeability may be a useful technique for detecting small intestinal disease and for monitoring the efficacy of treatment in dogs.

Increase in pulmonary capillary permeability in dogs exposed to 100% O 2

Increase in pulmonary capillary permeability in dogs exposed to 100% O 2

Endotoxin-pretreated neutrophils increase pulmonary vascular permeability in dogs.

Endotoxin causes pulmonary vascular neutrophil sequestration and injures the lung. Whether this is primarily due to a direct effect of endotoxin on the endothelium or is mediated by an action on the neutrophil is unclear. Canine neutrophils, isolated on plasma-Percoll gradients in vitro, were incubated with Salmonella enteriditis endotoxin, washed, and injected via wedged pulmonary arterial catheters into discrete lung zones of anesthetized dogs, whereas untreated neutrophils were administered into contralateral control lung zones. 113mIn-transferrin was administered intravenously 2 h before the animals were killed. Morphometry and extravascular protein accumulation were assessed at 4 h. Endotoxin treatment of neutrophils ex vivo induced a two- to three-fold increase in neutrophils in these lung zones (0.096 +/- 0.012 vs. 0.05 +/- 0.002 neutrophils/alveolar septal intercept, P less than 0.05). Extravascular-to-intravascular protein ratios in zones receiving endotoxin-treated neutrophils were significantly increased compared with control zones (0.146 +/- 0.02 vs. 0.079 +/- 0.02, P less than 0.05). Because complement fragments increase injury to endothelium in vitro, exogenous C5 fragments were administered to other dogs before administration of neutrophils but failed to significantly increase the extravascular protein signal (0.154 +/- 0.03 vs. 0.124 +/- 0.04). In summary, endotoxin treatment of neutrophils leads to neutrophil sequestration and increased pulmonary extravascular protein accumulation. C5 fragments failed to further enhance the protein accumulation. These data are consistent with an effect of endotoxin on the neutrophil to initiate neutrophil-endothelial interaction and subsequent lung injury.

Increase in pulmonary capillary permeability in dogs exposed to 100% O2.

Changes in pulmonary capillary filtration induced by hyperoxia were investigated in 15 dogs. After 12 h of normobaric hyperoxic exposure, animals were anesthetized and artificially ventilated with 100% O2. A pulmonary lymphatic vessel was cannulated, and lymph flow and protein content were measured together with pulmonary and systemic hemodynamics. An increase in pulmonary capillary filtration was found when compared with reference data (normoxic dogs in similar conditions) gathered from available literature: lymph flow increased from 21.8 +/- 13.4 to 125.2 +/- 131.6 microliter/min, and the lymph-to-plasma protein concentration ratio increased from 0.67 +/- 0.08 to 0.78 +/- 0.08. To characterize the mechanisms involved, left atrial pressure was increased in two stages (approximately 10 and approximately 25 mmHg). The results clearly indicated an increase in pulmonary capillary permeability as evidenced by a decrease of the minimal estimate of the protein reflection coefficient from 0.62 +/- 0.05 to 0.42 +/- 0.05.

Effect of phorbal myristate acetate (PMA) on pulmonary vascular resistance and permeability in dogs: R.C. Allison, E.M. Hernandez, and A.E. Taylor. Departments of Medicine and Physiology, University of South Alabama College of Medicine, Mobile, AL 36688

Effect of phorbal myristate acetate (PMA) on pulmonary vascular resistance and permeability in dogs: R.C. Allison, E.M. Hernandez, and A.E. Taylor. Departments of Medicine and Physiology, University of South Alabama College of Medicine, Mobile, AL 36688

The Role of the Arachidonic Acid Meta-bolites in Endotoxin-induced Increase in Lung Vascular Permeability in Dogs

The Role of the Arachidonic Acid Meta-bolites in Endotoxin-induced Increase in Lung Vascular Permeability in Dogs

Effects of fibrinogen in vivo on dentine permeability in the dog

Dentine permeability measurements were made each hour for 6 h in tooth cavities prepared in control dogs or dogs which had been pretreated with a purified fraction of snake venom to deplete their plasma of fibrinogen. The control dogs exhibited a progressive fall in dentine permeability (~ 50 per cent reduction in 6 h); the permeability in the fibrinogen-depleted dogs only fell 15 per cent in 6 h. The results implicate plasma proteins, particularly high molecular-weight proteins such as fibrinogen, as agents which reduce dentine permeability in vivo.