Comparative Medicine
Volume 59, Number 2, April 2009
OVERVIEW
Emerson et al. Enhancing the Ability of Experimental Autoimmune Encephalomyelitis to Serve as a More Rigorous Model of Multiple Sclerosis through Refinement of the Experimental Design, pp. 112-128
ACLAM Task Designation: Tasks 9 and 10
Species: multiple, including mice, rats, and nonhuman primates
SUMMARY: This paper reviews the mouse EAE model for multiple sclerosis (MS). It describes why the model is useful, including the similarity of clinical and pathologic signs between mice and humans. Additionally, the authors describe flaws in the EAE model, including extravasation of RBCs into the CNS of SJL mice and Lewis rats, but not in humans with MS. The paper also suggests ways in which investigators can improve experimental design to obtain more consistent and accurate study results. The authors focus on careful selection and preparation, and administration of drug and compare methods by which to induce EAE in animals. A significant portion of the paper is devoted to the scoring system used to evaluate EAE study animals, and ways in which the scoring system can be tailored to individual EAE models.
QUESTIONS:
1. All but which of the following are involved in the induction of EAE in animals?
a. Complete Freund Adjuvant (CFA)
b. Incomplete Freund Adjuvant (IFA)
c. E. coli
d. M. tuberculosis
e. Pertussus toxin
f. Myelin oligodendrocyte glycoprotein (MOG)
g. UV light
2. What does the acronym EAE stand for?
3. T/F: Scorers should be blinded to grouping of experimental animals and two scorers should ideally evaluate animals at each time point?
4. Which method is not useful in evaluating disease in EAE animals?
a. MRI
b. Behavioral testing
c. CBC
d. Observational scoring systems
ANSWERS:
1. C. All other drugs listed may be used in to induce EAE.
2. Experimental Autoimmune Encephalomyelitis or Experimental Allergic Encephalomyelitis
3. True. Scores from individual scorers can be averaged to determine an overall score for each time point. Studies where scorers were not blinded showed more positive results (significant differences between groups) than studies where scorers were blinded.
4. C. All other choices are used to evaluate disease in EAE models of MS.
ORIGINAL RESEARCH
Mouse Models
Marques et al. The Spatial Learning Phenotype of Heterozygous Leaner Mice is Robust to Systematic Variation of the Housing Environment, pp. 129-138
Primary Species: Mouse
Introduction: Calcium is well recognized as an important intracellular messenger. The mouse being tested in this study is the leaner mouse which has a mutation in the Cacna1a gene that encodes a subunit of a brain-specific voltage-gated Ca 2.1 calcium channel. Leaner mice have small cerebellums with loss of Purkinje and granule cells. In the homozygous mouse clinical cells include seizures and cerebellar ataxia leading to death by 3 weeks of age. Historically the heterozygous mice have been described as normal. The current study has hypothesized that the heterozygous mice would not be normal compared to wild type mice if they were examined more closely over time and that this model would be more typical of the human condition. The authors go on to affirm that "for a behavioral difference to be relevant, it should be reasonably robust over environments". Having varying degrees of environmental enrichment (EE) in the cage would also allow observation as to whether EE could affect the onset and progression of this neurodegenerative disorder. Three different housing systems were used, a barren cage, a cage with nesting material and a cardboard tube, and thirdly a cage that is larger with nesting material, cardboard tube, nestbox and a running wheel. A Morris water maze was used to evaluate learning and memory. A ceiling camera placed above the water maze recorded animal movement. Mice were tested at 6, 12, and 20 months of age.
Results: WT mice were heavier than the heterozygous (tg la/+) mice. At later time points mice from furnished cages were heavier then mice from less furnished cages. The tg la/+ mice showed cognitive and memory impairment at all time points examined compared with WT mice. For both WT and tg la/+ mice from furnished cages outperformed mice from less furnished cages.
Discussion: The authors were able to show that the heterozygous leaner mouse does have cognitive impairment that is robust across the microenvironments evaluated. The results also demonstrated that physical and behavioral stimulation helped to rescue cognitive deficits in this mouse model.
QUESTIONS:
1. What structural anomalies are associated with the leaner mouse?
2. T/F The complexity of the microenvironment had no effect on cognition and learning in the tg la/+
3. In humans the heterozygous condition is a mutation in the CACNA1A gene. What is the mode of inheritance?
ANSWERS:
1. Small cerebellums with loss of Purkinje and granule cells.
2. F
3. Autosomal dominant
Gografe et al. Novel Pathologic Findings Associated with Urinary Retention in a Mouse Model of Mucopolysaccharidosis Type IIIB, pp. 139-146
Domain 3; Tasks: T3; TT 3.1, 3.3, 3.6
Primary Species: Mouse
SUMMARY: Mucopolysaccharidosis type IIIB, also called Sanfilippo syndrome type B, is a metabolic disorder with devastating clinical characteristics in humans. Onset of disease is between 2 and 4 years, and clinical symptoms include hyperactivity, aggressive behavior, hearing and vision defects, mental retardation, and mild somatic changes. These symptoms usually progress rapidly, and are followed by premature death. MPS IIIB is a lysosomal storage disorder, and is elicited by mutations in the Naglu gene, which is located on chromosome 17q21.
The knockout B6.129S6- Naglutm1Efn/J strain was developed through targeted mutation, and has become invaluable to continued investigations of the pathogenesis of this disease. However, has exhibited several objectionable phenotypic features such as, urinary retention, leading to a grossly enlarged urinary bladder, potential hydronephrosis, and uremia. These abnormalities are not typically seen in human MPS IIIB, and have also become an animal welfare issue for the mice, often leading to the early termination of experiments. The aim of this article was to investigate clinical and pathologic correlations underlying anomalous bladder enlargement in this mouse model.
Necropsies were performed on 9-mo-old mice; urinalysis, hematology and blood chemistry parameters were evaluated, and urogenital specimens were microscopically examined. All mutant mice examined showed significantly distended bladders, with most containing dark yellow to brown colored urine. The most striking histological finding was that of a mononuclear cell infiltrate in the lower urinary tract of the mutant mice. The authors were unable to confirm a causative relationship between the infiltrate and the urinary retention.
QUESTIONS:
1. What is the normal daily amount of urine produced by a mouse?
a. 0.2 – 0.4mL
b. 0.5 – 1.0mL
c. 1.0 – 1.3mL
2. T or F; Urinary retention, leading to grossly enlarged bladders is typical of human MPS IIIB.
ANSWERS:
1. b. 0.5 – 1.0 mL
2. False, this is why B6.129S6-Naglu tm1Efn/J mouse strain is not the best model for this disease.
Kolappaswamy et al. Effect of Inositol Hexaphosphate on the Development of UVB-Induced Skin Tumors in SKH1 Hairless Mice, pp. 147-152
Domain 3
Primary Species: Mouse
SUMMARY: This study evaluated the effect of Inositol Hexaphosphate (IP6) on UVB-induced skin tumors in hairless mice. IP6 is a compound found in most plants and is important in mammals for the regulation of cellular functions and the signal transduction system. IP6 has been shown to have antineoplastic properties. Two groups of 15 female SKH1 hairless mice were exposed to UVB radiation 3 times per week until tumors began appearing by week 12. One group concurrently received 2% IP6 in the drinking water beginning 3 days before irradiation. Tumor formation was monitored until week 31. Mice receiving IP6 in the drinking water had a significantly decreased incidence of tumor formation. Additionally, IP6 decreased NFκB expression in the skin tumors.
QUESTIONS:
1. Which of the following is NOT true?
a. IP6 has antineoplastic, antioxidative, and antiangiogenic properties
b. IP6 is found in most plants
c. IP6 may cause severe liver damage at the dose administered
d. IP6 and its lower phosphorylated structures occur in almost all cell types
2. Stronger tumor inhibition may occur with IP6 administration in the drinking water as opposed to in the food because:
a. IP6 in food tastes very bad
b. IP6 is more readily absorbed when in water
c. IP6 causes food to clump, making it inedible
d. A higher concentration of IP6 can be made in drinking water than in food
ASWERS:
1. c
2. b
Canine Models
Jergens et al. Intestinal Cytokine mRNA Expression in Canine Inflammatory Bowel Disease: A Meta-Analysis with Critical Appraisal, pp. 153-162
Primary Species: Dog
SUMMARY: Idiopathic inflammatory bowel disease (IBD) in dogs is thought to result from an altered interaction between resident micro flora and mucosal immunological defenses in a susceptible host. Cytokines are thought to play a key role in the modulation of the mucosal immune system as in humans, in which disturbances in the balance of proinflammatory (Th1/Th17-derived) and immunoregulatory (Th2/Tr1- derived) cytokines occur.
The objectives of this study were to 1)assess cytokine mRNA expression in biopsies of the intestinal mucosa collected prospectively from dogs diagnosed with small and large intestinal IBD by using semi quantitative RT-PCR technique and 2) to compare these data with previously published data to determine the putative role of cytokine expression in the pathogenesis of canine IBD and other forms of chronic enteropathy through meta-analysis of combined findings. The prospective study revealed that the intestines of both health and IBD dogs express various cytokines and a proinflammatory profile (Th-1) is not a feature of IBD. Meta-analysis shows that dogs with IBD fail to express a predominant Th1- or Th2 cytokine bias in the inflamed mucosa, and a future studies will require greater standardization of stage of disease and demographic factors of the study population.
QUESTIONS:
1. Th1 and Th2 cytokines are primarily produced by subsets of :
A. CD8 T lymphocytes
B. CD4 T lymphocytes
C. B Plasma Cells
D. NKT Cells
2. T/F. Increases in exclusively Th1- derived cytokines are primarily pro-inflammatory in the gastrointestinal mucosa.
3. One breed of dog that was overrepresented in the Meta-analysis of several studies of IBD
A. Dachshund
B. Yorkshire Terrier
C. Doberman Pincher
D. German Shepherd
ANSWERS:
1. B
2. T
3. D
Kuribayashi et al. Bovine Colostral Antibody Against Verotoxin 2 Derived from Escherichia coli O157:H7: Resistance to Proteases and Effects in Beagle Dogs, pp. 163-167
Primary Species: Canine
RDD: Task 3
SUMMARY: A bovine colostral antibody against verotoxin (VT)2 of E. Coli 0157:H7 was administered orally to beagle dogs. The antibody remained in the dogs’ small intestine for at least 2 h, whereas little serum antibody remained 1.5 h after administration. The antibody activity of secretory IgA did not change until 2 h after administration; however, the activity of IgG and IgM antibodies decreased by approximately 60% and 40% at 2 h after administration , respectively. Seven beagle dogs inoculated with E. coli 0157:H7 producing VT2 were administered bovine colostral antibody or bovine colostral whey without antibody. With administration of bovine colostral whey without antibody, the amount of VT2 in feces decreased gradually after administration and increased again at 5 d after inoculation, whereas bovine colostral antibody significantly reduced the amount of VT2 in feces on the day after administration. In addition 9 beagle dogs were given bovine colostral antibody, bovine plasma antibody, or saline. The amount of VT2 in feces again decreased significantly more rapidly after administration of bovine colostral antibody than after administration of bovine plasma antibody or saline.
QUESTIONS:
1. T/F Colostral antibody (which includes secretory IgA and IgG components) was resistant to protease degradation and showed sufficient neutralization efficacy against VT2 in the intestine.
2. Why was an antibiotic administered to pre-treat the beagles in the study prior to E. coli administration?
3. What antibiotic was used to pre-treat the beagles prior to the study?
ANSWERS:
1. T
2. The antibiotic was used because without pretreatment the healthy dogs with normal enterobacterial flora may not be sensitive to E. coli 0157:H7. The antibiotic served to alter the dogs’ indigenous flora symptoms of diarrhea.
3. Fradiomycin sulfate
Nonhuman Primate Models
MacGuire et al. Serologic Evaluation of Clinical and Subclinical Secondary Hepatic Amyloidosis in Rhesus Macaques (Macaca mulatta), pp. 168-173
Primary Species: Monkey
Domain 1: Management of spontaneous and experimentally induced diseases and conditions; Task3: Diagnose disease or condition as appropriate
SUMMARY: This is a retrospective study based on 26 histologically diagnosed amyloidotic macaques evaluated at 2 stages of the disease; clinical and subclinical, and 19 additional histologically diagnosed amyloidotic macaques with 19 age-matched controls. The purpose was to establish serologic parameters to identify macaques in the subclinical stage of the disease.
Clinically amyloidotic animals displayed increases in alkaline phosphatase, aspartate amino transferase, lactate dehydrogenase, and gamma glutamyltranspeptidase and decreased concentrations of albumen and total protein. Subclinical animals showed increased quantities of alkaline phosphatase, aspartate amino transferase and lactate dehydrogenase but decreased albumin and cholesterol. All other serum biomarkers were not significantly different from colony references.
ELISA for serum amyloid A (SAA) and macrophage stimulating factor (mCSF) used in combination are predictive of amyloidosis in the subclinical stage and mCSF in the clinical stage.
QUESTIONS:
1. Amyloidosis is classified into three categories: primary, secondary and hereditary. Which is the most common in animals and what is associated with?
2. T/F: Clinical signs of amyloidosis in macaques include: cachexia, muscle wasting, recurrent diarrhea, hepatomegaly, anemia, hypoproteinemia, elevated liver enzymes or arthralgia. Therefore sedation of these animals for diagnostics should be routine.
3. Which category of amyloidosis is rare?
ANSWERS:
1. Secondary is most common; associated with chronic inflammation.
2. false of course - prognosis is grave
3. hereditary syndromesare rare and include Mediterranean fever, Muckle-Wells syndrome, and familial amyloid cardiomyopathy.
Kramer et al. Treatment of Giardiasis in Common Marmosets (Callithrix jacchus) with Tinidazole, pp. 174-179