Establishing a laboratory animal model from a transgenic animal: RasH2 mice as a model for carcinogenicity studies in regulatory science

Transgenic animal models have been used in small numbers in gene function studies in vivo for a period of time, but more recently, the use of a single transgenic animal model has been approved as a second species, 6-month alternative (to the routine 2-year, 2-animal model) used in short-term carcinogenicity studies for generating regulatory application data of new drugs. This article addresses many of the issues associated with the creation and use of one of these transgenic models, the rasH2 mouse, for regulatory science. The discussion includes strategies for mass producing mice with the same stable phenotype, including constructing the transgene, choosing a founder mouse, and controlling both the transgene and background genes; strategies for developing the model for regulatory science, including measurements of carcinogen susceptibility, stability of a large-scale production system, and monitoring for uniform carcinogenicity responses; and finally, efficient use of the transgenic animal model on study. Approximately 20% of mouse carcinogenicity studies for new drug applications in the United States currently use transgenic models, typically the rasH2 mouse. The rasH2 mouse could contribute to animal welfare by reducing the numbers of animals used as well as reducing the cost of carcinogenicity studies. A better understanding of the advantages and disadvantages of the transgenic rasH2 mouse will result in greater and more efficient use of this animal model in the future.     

Protothecosis as a cause of chronic diarrhoea in a dog

The clinical form of the protothecosis in animals is most commonly observed in countries with a warm and moist climate, only a few reports describing cases of this infection in cooler areas of the word exist. In the case of large bowel infection in dogs, organisms colonise the lamina propria and submucosa causing severe necrotizing ulcerative or haemorrhagic enterocolitis. In this report the intestinal form of protothecosis in 1.5-year-old, male, mongrel dog with chronic hemorrhagic diarrhoea is described. History revealed that the dog spent some time in the countryside and afterwards diarrhoea with fresh blood appeared. The results of morphological and biochemical blood analysis were normal and stool examination did not reveal the presence of parasites. Treatment with anti-inflammatory doses of prednisone, metronidazole and enrofloxacin followed by sulphasalazine resulted in a short period of improvement, but was followed by deep deterioration of animal status. Because of the relapse diagnostic laparotomy was performed and tissue samples of the colon and jejunum were obtained for histopathology. On the basis of the clinical signs, exploratory laparotomy findings and histopathology the diagnosis of canine intestinal prototecosis was made and medical treatment was recommended.     

Canine periodontitis: the dog as an important model for periodontal studies

Periodontal disease (PD) refers to a group of inflammatory diseases caused by bacterial plaque in the periodontium and ranges from an early stage (gingivitis) to an advanced stage (periodontitis). It is a multifactorial disease that results from the interaction of the host defence mechanisms with the plaque microorganisms. Early detection, diagnosis and treatment are essential in the control of this disease. PD has an enormous impact on human and veterinary medicine due to its high prevalence. The most common animal PD models use dogs and non-human primates, although other animals (rats, mice, hamsters, rabbits, miniature pigs, ferrets, and sheep) have also been employed. Dog models have contributed significantly to the current understanding of periodontology. The most important clinical aspects of canine PD are considered in this review and the various animal models are examined with an emphasis on the role of the dog as the most useful approach for understanding human PD and in the development of new therapeutic and preventive measures.     

Phenotypic analysis of hepatic T lymphocytes in a dog with chronic hepatitis

We evaluated hepatic T lymphocyte phenotypes in a dog with chronic hepatitis. Before treatment, numerous CD3+ lymphocytes were demonstrated in the liver, and the ratio of CD4+/CD8+ was remarkably high (2.96; reference range, 0.33+/-0.12). After treatment, CD3+ lymphocyte infiltration in the liver was reduced, and the ratio of CD4+/CD8+ decreased to 0.31. Therefore, hepatic T lymphocytes, especially CD4+ lymphocytes, might play a central role in the pathogenesis of this dog with chronic hepatitis.     

A systematic study of Tupaia as a model for human acute hepatitis B infection

The molecular features of hepatitis B virus (HBV) infection, eradication, and pathogenesis are poorly understood, partly due to the lack of an adequate animal model that faithfully reproduces the course of infection. Although Tupaia belangeri were previously recognized as HBV-susceptible animals, the course of infection in adult tupaias remains obscure. Herein, we performed a longitudinal study and demonstrated that adult tupaias were efficiently infected (90% infection rate) with 10⁸ copies of the HBV genome. HBV replicated vigorously, produced high levels of covalently closed circular DNA (cccDNA) in hepatocytes, and released hepatitis B surface antigen (HBsAg), hepatitis Be antigen (HBeAg), and HBV DNA into the serum at day 9 post-inoculation (p.i.), which then decreased on day 15 p.i. The kinetics were consistent with the expression of liver HBsAg and HBeAg, as determined with immunohistochemistry. The viral products in serum at day 9 and 15 p.i. represented de novo synthesized viral products, as treatment with a viral entry inhibitor completely abolished these products from the serum. Viral clearance and serological conversion occurred at day 21 p.i. and were accompanied by elevated alanine transaminase (ALT) levels and liver pathology, such as inflammatory infiltration and hepatocyte ballooning degeneration. Although ALT levels eventually returned to normal levels by day 42 p.i., the liver pathology persisted until at least day 120 p.i. The HBV infection process in tupaia, therefore, exhibits features similar to that of human acute HBV infection, including viral replication, viral eradication, ALT elevation, and liver pathology. Thus, adopting the tupaia model to study host-HBV interactions presents an important advance which could facilitate further investigation and understanding of human HBV infection, especially for features like cccDNA that current small-animal models cannot effectively model.     

Dacryocystomaxillorhinostomy for chronic dacryocystitis in a dog

A 10-year-old female spayed Vizsla had intermittent mucoid ocular discharge from the right eye for 7 years. History, clinical findings, imaging studies, and culture and histopathology results confirmed chronic dacryocystitis with granuloma. A dacryocystomaxillorhinostomy was performed to preserve the functional portions of the nasolacrimal system remaining in this patient, as well as to promote healing of the lacrimal sac granuloma and secondary infection. Complete resolution of the clinical abnormalities was achieved, and the dog remains healthy 3 years postoperatively.     

Effects of exercise and human contact on animal welfare in a dog shelter

The aim of the study is to investigate the reduction of stress in dogs in municipal shelters through easy-to-implement activities, ie, 25-minute sessions of exercise and human contact, that do not require a significant investment in terms of funding, staff or time. The results demonstrate that the dogs taking part in these sessions have lower salivary cortisol levels (F=121.42; P<0.05) and achieve better scores on a behaviour test (t(17)=4.27; P=0.001). It can therefore be affirmed that the exercise and human contact protocol proposed in the present study diminishes stress and improves the welfare of dogs housed in shelters.     

Farmed deer: a large animal model for stress

In the early classic period it was suggested by Heracleitus that a static unchanged state was not the natural condition, but rather that the capacity to undergo constant change was intrinsic to all living things. After that, Empedoeles, circa 500BC, proposed that for living organisms to survive they require a harmonious mixture of elements. Hippocrates, around 400 BC, developed this concept further by suggesting that health is a state of harmonious balance of these elements, equating disease to a state of disharmony and introducing the concept that nature heals disease (Noysvn Fyseiw/atrui). Soon after, Epicurus extended these ideas to include among these healing forces, the mind, writing that imperturbability of mind is desirable.Claude Bernard introduced the concept of the milieu interieur or the internal physiologic environment, describing its constancy as essential in an external environment typified by its variability. Walter Cannon described Bernard’s constancy as Homeostasis and identified the “fight or flight reflex,” linking the adaptive response and catecholamine secretion, with extreme levels of activation-producing pathology.Selye outlined four stages of the stress response: 1) the “alarm reaction” characterized by an immediate activation of the sympathetic-adrenomedullary axis (SA); 2) a “resistance phase” characterized by hypothalamic pituitary-adrenal axis (HPA) activation; 3) a stage of adrenal hypertrophy, gastrointestinal ulceration, along with thymic and lymphoid atrophy; and 4) an exhaustion phase and finally death (1).     

Lymphoma as a model for chronic illness: effects on adrenocortical function testing

Many dogs with chronic illness have serum biochemical abnormalities consistent with hyperadrenocorticism (HAC). Lymphoma (LSA) is a chronic disease of dogs. The purpose of this study was to evaluate adrenocortical screening test results in dogs with LSA to evaluate their specificity. Criteria for inclusion in the study included a diagnosis of LSA, an expected survival time of 16-56 weeks, no glucocorticoid treatment beyond 4 weeks after the initiation of chemotherapy, no evidence of HAC, and owner consent. Post-ACTH stimulation plasma cortisol concentrations (PACs), urine cortisol : creatinine (UC : Cr) ratios, and maximal left adrenal width measurements were performed at the time of LSA diagnosis before the initiation of chemotherapy and at 16, 24, 32, 40, and 52 weeks or until the loss of remission or the development of another disease. Ten dogs met the criteria for inclusion. Forty-two PACs were performed; 1 abnormal, 2 borderline, and 39 normal values were detected. Thirty-five maximal left adrenal width measurements were obtained; 0 abnormal, 5 borderline, and 30 normal measurements were detected. Thirty-six UC : Cr ratios were obtained, with 26 abnormal, 4 borderline, and 6 normal values detected and 9 of 10 dogs having at least 1 abnormal value. These data suggest that in dogs with LSA, the UC : Cr ratio frequently is abnormal and may not be a specific test for HAC, or it may be the most sensitive test for increases in cortisol secretion due to chronic illness. Maximal left adrenal width measurements and PACs were almost always normal and may be more specific for HAC or less sensitive for demonstrating chronic increases in cortisol secretion.     

Field assessment of dog as sentinel animal for plague in endemic foci of Madagascar

The epidemiology of Yersinia pestis, the causative agent of plague, involves vectors and reservoirs in its transmission cycle. The passive plague surveillance in Madagascar targets mainly rodent and fleas. However, carnivores are routinely surveyed as sentinels of local plague activity in some countries. The aim of this study is to assess the use of domestic dog (Canis familiaris) as sentinel animal for field surveillance of plague in a highly endemic area in Madagascar. Cross-sectional surveys of plague antibody prevalence in C. familiaris were conducted in endemic areas with contrasting histories of plague cases in humans, as well as a plague free area. Rodent capture was done in parallel to evaluate evidence for Y. pestis circulation in the primary reservoirs. In 2 sites, dogs were later re-sampled to examine evidence of seroconversion and antibody persistence. Biological samplings were performed between March 2008 and February 2009. Plague antibody detection was assessed using anti-F1 ELISA. Our study showed a significant difference in dog prevalence rates between plague-endemic and plague-free areas, with no seropositive dogs detected in the plague free area. No correlation was found between rodents and dog prevalence rates, with an absence of seropositive rodents in some area where plague circulation was indicated by seropositive dogs. This is consistent with high mortality rates in rodents following infection. Re-sampling dogs identified individuals seropositive on both occasions, indicating high rates of re-exposure and/or persistence of plague antibodies for at least 9 months. Seroconversion or seropositive juvenile dogs indicated recent local plague circulation. In Madagascar, dog surveillance for plague antibody could be useful to identify plague circulation in new areas or quiescent areas within endemic zones. Within active endemic areas, monitoring of dog populations for seroconversion (negative to positive) or seropositive juvenile dogs could be useful for identifying areas at greatest risk of human outbreaks.     

Field Epidemiology and Laboratory Training Programs in West Africa as a model for sustainable partnerships in animal and human health

The concept of animal and human health experts working together toward a healthier world has been endorsed, but challenges remain in identifying concrete actions to move this one health concept from vision to action. In 2008, as a result of avian influenza outbreaks in West Africa, international donor support led to a unique opportunity to invest in Field Epidemiology and Laboratory Training Programs (FELTPs) in the region that engaged the animal and human health sectors to strengthen the capacity for prevention and control of zoonotic diseases. The FELTPs mixed 25% to 35% classroom and 65% to 75% field-based training and service for cohorts of physicians, veterinarians, and laboratory scientists. They typically consisted of a 2-year course leading to a master's degree in field epidemiology and public health laboratory management for midlevel public health leaders and competency-based short courses for frontline public health surveillance workers. Trainees and graduates work in multidisciplinary teams to conduct surveillance, outbreak investigations, and epidemiological studies for disease control locally and across borders. Critical outcomes of these programs include development of a cadre of public health leaders with core skills in integrated disease surveillance, outbreak investigation, vaccination campaigns, laboratory diagnostic testing, and epidemiological studies that address priority public health problems. A key challenge exists in identifying ways to successfully scale up and transform this innovative donor-driven program into a sustainable multisectoral one health workforce capacity development model.     

The pentobarbital anesthetized dog: an animal model for assessing chemically induced changes in renal function and ultrastructure

An experimental procedure was devised using the pentobarbital-anesthetized dog that could be used for the comprehensive evaluation of the renal effects of chemicals. After IV or renal arterial administration of 0.9% saline solution (vehicle), 12 renal function determinants were continuously monitored for periods of 2 and 6 hours. At the completion of the 2 or 6 hours of study, the kidneys of a number of dogs (usually between 1 and 7) in each vehicle-treated group were subjected to a modification of the intravascular perfusion-of-fixative technique to evaluate the ultrastructural status of the outer cortical, inner cortical, and outer medullary tissue. The remaining dogs (at least 3) in each vehicle-treated group were given a nonnephrotoxic, but maximally effective, diuretic dose of ethacrynic acid, which enabled an assessment of the functional integrity of the thick ascending limb of Henle's loop. Renal function and glomerular and tubular ultrastructure remained stable in the pentobarbital-anesthetized dog for up to 6 hours after administration of vehicle. Sustained infusion of inulin (included in the procedure to estimate glomerular filtration rate) throughout the duration of the experiments, and pentobarbital anesthesia of various durations did not alter the morphologic status of the canine nephron. The procedure used for the renal perfusion of fixative circumvented any manipulation of the kidneys before fixation and allowed for the acquisition of normal (unaltered) appearing tissue from all areas of the kidneys. The responses of pentobarbital-anesthetized dogs to ethacrynic acid administration were similar when given 2 and 6 hours after the vehicle administration.(ABSTRACT TRUNCATED AT 250 WORDS)     

Comparative phenotypic and molecular characterization of porcine mesenchymal stem cells from different sources for translational studies in a large animal model

Mesenchymal stem cells have demonstrated their potentiality for therapeutic use in treating diseases or repairing damaged tissues. However, in some cases, the results of clinical trials have been disappointing or have not worked out as well as hoped. These disappointing results can be attributed to an inadequate or insufficient preclinical study. For medical and surgical purposes, the similarities between the anatomy of pig and human make this animal an attractive preclinical model. In this sense, for mesenchymal stem cell-based therapy, it is strongly necessary to have well characterized animal-derived mesenchymal stem cell lines to validate preclinical effectiveness of these cells. In this work, porcine mesenchymal stem cells (pMSCs) were isolated from bone marrow, adipose tissue and peripheral blood and compared in terms of differentiation potential, cell surface markers and gene expression. Our results demonstrated that the isolation and in vitro expansion protocols were feasible and effective. The data presented in this work are relevant because they provide an extensive phenotypic characterization; genetic study and differentiation behavior of the most commonly used stem cell lines for clinical practices. These pMSCs are widely available to scientists and could be a valuable tool to evaluate the safety and efficacy of adoptively transferred cells.     

Proliferative glomerulonephritis and chronic active hepatitis with cirrhosis associated with Corynebacterium parvum immunotherapy in a dog

Ivermectin, a broad-spectrum antiparasitic agent, was believed responsible for signs of CNS dysfunction in a dog. Within 2 hours of oral administration of ivermectin, the dog had hind limb ataxia. Neurologic signs progressed rapidly until the dog was in a semicomatose state at admission 20 hours later. The dog improved gradually under supportive care. The ivermectin had been given for suspected endoparasitism. Clinical signs were similar to those reported in dogs with clinical and experimental exposure to ivermectin.