RESUMO
BACKGROUND: Leptospirosis is a zoonotic disease that is found globally and affects most mammalian species. Vaccination of dogs against leptospirosis is an important approach to preventing clinical disease, or reducing disease severity, as well as reducing transmission of the infection to humans. Although it is generally considered to be a 'core' vaccine, there is limited information on the level of leptospirosis vaccine usage and factors associated with its usage in dogs in the UK. The study aimed to report the uptake of leptospirosis vaccination and factors associated with its usage in a cohort of dogs under primary veterinary care during a 12-month period. RESULTS: From a population of 905,543 dogs, 49% (95%CI 48.9-49.1%) had at least one leptospirosis vaccine administered during the 12 months of study. Adult dogs had reduced odds of receiving a leptospirosis vaccine when compared to dogs < 1 year old, with dogs > 8 years old having a greater than ten-fold reduction in odds (OR = 0.08, 95%CI 0.07-0.09). Odds of receiving a leptospirosis vaccine was increased in insured dogs when compared to uninsured dogs (OR = 1.22, 95%CI = 1.17-1.28). Neutered dogs had reduced odds of receiving a leptospirosis vaccine (OR = 0.87, 95%CI 0.83-0.91). Breed associations with receiving a leptospirosis vaccine varied. Several breeds were associated with increased odds of receiving a leptospirosis vaccine when compared to crossbreed dogs, including Border Terriers (OR = 1.49, 95%CI 1.42-1.57), Golden Retrievers (OR = 1.30, 95%CI = 1.24-1.37), Cocker Spaniels (OR = 1.27, 95%CI 1.23-1.31) and West Highland White Terriers (OR = 1.27, 95%CI 1.22-1.31). French Bulldogs (OR = 0.64, 95%CI = 0.62-0.67), Staffordshire Bull Terriers (OR = 0.79, 95%CI 0.78-0.82) and Pugs (OR = 0.91, 95%CI =0.88-0.95) had significantly reduced odds of receiving a leptospirosis vaccination during the study. CONCLUSION: This work identified that almost half of the UK primary care attending population received a leptospirosis vaccine during the year. Several demographic variables were associated with leptospirosis vaccine administration, with age being particularly important. Both the proportion of uptake and factors associated with leptospirosis vaccine usage can be used as a benchmark for comparisons in the future. Additionally, an understanding of which populations have reduced odds of receiving a leptospirosis vaccine can potentially be used for initiatives to encourage owner vaccination uptake in these groups.
Assuntos
Doenças do Cão , Leptospirose , Vacinas , Animais , Doenças do Cão/epidemiologia , Cães , Humanos , Leptospirose/epidemiologia , Leptospirose/prevenção & controle , Leptospirose/veterinária , Mamíferos , Atenção Primária à Saúde , Fatores de Risco , Reino Unido/epidemiologia , Vacinação/veterináriaRESUMO
Students entering the final year of the veterinary curriculum need to integrate information and problem solve. Assessments used to document competency prior to entry to the clinical environment should ideally provide a reliable measurement of these essential skills. In this study, five internal medicine specialists evaluated the cognitive grade (CG) and structural integrity of 100 multiple-choice questions (MCQs) used to assess learning by third-year students at a United States (US) veterinary school. Questions in CG 1 tested factual recall and simple understanding; those in CG 2 required interpretation and analysis; CG 3 MCQs tested problem solving. The majority (53%) of questions could be answered correctly using only recall or simple understanding (CG 1); 12% of MCQs required problem solving (CG 3). Less than half of the questions (43%) were structurally sound. Overall student performance for the 3 CGs differed significantly (92% for CG 1 vs. 84% for CG 3; p = .03. Structural integrity did not appear to impact overall performance, with a median pass rate of 90% for flawless questions versus 86% for those with poor structural integrity (p = .314). There was a moderate positive correlation between individual student outcomes for flawless CG 1 versus CG 3 questions (rs = 0.471; p = < .001), although 13% of students failed to achieve an aggregate passing score (65%) on the CG 3 questions. These findings suggest that MCQ-based assessments may not adequately evaluate intended learning outcomes and that instructors may benefit from guidance and training for this issue.
Assuntos
Educação em Veterinária , Estudantes de Medicina , Animais , Cognição , Avaliação Educacional , Humanos , Reprodutibilidade dos TestesRESUMO
The clinical application of cardiosphere-derived cells (CDCs) to treat cardiac disease has gained increasing interest over the past decade. Recent clinical trials confirm their regenerative capabilities, although much remains to be elucidated about their basic biology. To develop this new treatment modality, in a cost effective and standardized workflow, necessitates the creation of cryopreserved cell lines to facilitate access for cardiac patients requiring urgent therapy. Cryopreservation may however lead to alterations in cell behavior and potency. The aim of this study was to investigate the effect of cryopreservation on canine CDCs. CDCs and mesenchymal stem cells (MSCs) isolated from five dogs were characterized. CDCs demonstrated a population doubling time that was unchanged by cryopreservation (fresh vs. cryopreserved; 57.13 ± 5.27 h vs. 48.94 ± 9.55 h, P = 0.71). This was slower than for MSCs (30.46 h, P < 0.05). The ability to form clones, self-renew, and commit to multiple lineages was unaffected by cryopreservation. Cryopreserved CDCs formed larger cardiospheres compared to fresh cells (P < 0.0001). Fresh CDCs showed a high proportion of CD105+ (89.0% ± 4.98) and CD44+ (99.68% ± 0.13) cells with varying proportions of CD90+ (23.36% ± 9.78), CD34+ (7.18% ± 4.03) and c-Kit+ (13.17% ± 8.67) cells. CD45+ (0.015% ± 0.005) and CD29+ (2.92% ± 2.46) populations were negligible. Increasing passage number of fresh CDCs correlated with an increase in the proportion of CD34+ and a decrease in CD90+ cells (P = 0.003 and 0.03, respectively). Cryopreserved CDCs displayed increased CD34+ (P < 0.001) and decreased CD90+ cells (P = 0.042) when compared to fresh cells. Overall, our study shows that cryopreservation of canine CDCs is feasible without altering their stem characteristics, thereby facilitating their utilization for clinical trials. © 2017 International Society for Advancement of Cytometry.
Assuntos
Células-Tronco Adultas/citologia , Criopreservação/veterinária , Mioblastos Cardíacos/citologia , Animais , Antígenos CD34/metabolismo , Cardiomiopatia Dilatada/terapia , Cardiomiopatia Dilatada/veterinária , Diferenciação Celular/imunologia , Linhagem da Célula , Proliferação de Células , Separação Celular , Células Cultivadas , Criopreservação/métodos , Doenças do Cão/terapia , Cães , Átrios do Coração/citologia , Células-Tronco Mesenquimais/citologia , Miócitos Cardíacos/citologia , Transplante de Células-Tronco/métodos , Transplante de Células-Tronco/veterinária , Antígenos Thy-1/metabolismoRESUMO
Canine anal furunculosis (AF) is characterised by ulceration and fistulation of perianal tissue and is a disease that particularly affects German shepherd dogs (GSDs). There are some similarities between AF and perianal Crohn's disease (CD) in man. An immune-mediated aetiopathogenesis for AF has been suggested due to tissue pathology, a major histocompatibility complex (MHC) association and clinical response to ciclosporin. Genome-wide association studies (GWAS) can be conducted in dogs with fewer markers and individuals than would be required in a human study. A discovery GWAS was performed on 21 affected and 25 control GSDs from the UK. No SNPs reached genome-wide significance levels at this stage. However, 127 nominally associated SNPs were genotyped in further 76 cases and 191 controls from the UK and Finland. Sequencing of these regions was undertaken to discover novel genetic variation. Association testing of these variants in the UK and Finnish cohorts revealed nine significantly associated SNPs, six of which cause non-synonymous changes in protein sequence. The ADAMTS16 and CTNND2 gene regions were most significantly associated with disease. Members of the butyrophilin protein family, important in intestinal inflammatory regulation, were also associated with disease, but their independence from the MHC region remains to be established. The CTNND2 gene region is also interesting as this locus was implicated in human ulcerative colitis and CD, albeit at a different candidate gene: DAP. We suggest that this represents a common association between inflammatory bowel disease-related conditions in both species and believe that future studies will strengthen this link.
Assuntos
Doenças do Ânus/veterinária , Doenças do Cão/genética , Furunculose/veterinária , Animais , Estudos de Casos e Controles , Mapeamento Cromossômico , Cães , Finlândia , Estudo de Associação Genômica Ampla , Razão de Chances , Polimorfismo de Nucleotídeo Único , Reprodutibilidade dos Testes , Análise de Sequência de DNA , Reino UnidoRESUMO
Canine hypoadrenocorticism is an endocrine disorder characterised by inadequate secretion of steroid hormones from the adrenal glands. Pathology results from immune-mediated destruction of the adrenal cortex, which is similar to that seen in the human Addison's disease. Both the canine and human diseases have similar clinical presentation, with the diagnosis based on performing a dynamic adrenocorticotropic hormone stimulation test. MHC class II has previously been associated with the human and canine diseases. In the current study, we conducted an MHC class II association study in eight breeds of dog with diagnoses of hypoadrenocorticism. We demonstrated significant differences in dog leukocyte antigen (DLA) haplotype frequencies in six of these breeds: Cocker spaniel, Springer spaniel, Labrador, West Highland white terrier (WHWT), Bearded collie, and Standard poodle. In the Springer spaniel, the DLA-DRB1*015:01--DQA1*006:01--DQB1*023:01 haplotype was significantly associated with disease risk (p = 0.014, odds ratio (OR) = 5.14) and showed a similar trend in the Cocker spaniel. This haplotype is related to one associated with hypoadrenocorticism in the Nova Scotia duck tolling retriever. Similar haplotypes shared between breeds were demonstrated, with DLA-DRB1*001:01--DQA1*001:01--DQB1*002:01 more prevalent in both affected Labrador (p = 0.0002, OR = 3.06) and WHWT (p = 0.01, OR = 2.11). Other haplotypes that have not previously been associated with the disease were identified. The inter-breed differences in DLA haplotypes associated with susceptibility to canine hypoadrenocorticism could represent divergent aetiologies. This could have implications for clinical diagnosis and future comparative studies. Alternatively, it may suggest that the gene of interest is closely linked to the MHC.
Assuntos
Insuficiência Adrenal/veterinária , Doenças do Cão/genética , Genes MHC da Classe II , Predisposição Genética para Doença , Insuficiência Adrenal/genética , Sequência de Aminoácidos , Animais , Cães , Cadeias beta de HLA-DQ/química , Cadeias beta de HLA-DQ/genética , Cadeias HLA-DRB1/química , Cadeias HLA-DRB1/genética , Haplótipos , Homozigoto , Dados de Sequência Molecular , Alinhamento de SequênciaRESUMO
Canine hypoadrenocorticism is believed to be an immune-related condition. It is rare in the overall dog population but shows a breed-related predisposition with Standard poodles and Portuguese water dogs having a greater prevalence of the condition. It shares many similarities with human primary adrenal insufficiency and is believed to be a naturally occurring, spontaneous model for the human condition. Short haplotype blocks and low levels of linkage disequilibrium in the human genome mean that the identification of genetic contributors to the condition requires large sample numbers. Pedigree dogs have high linkage disequilibrium and long haplotypes within a breed, increasing the potential of identifying novel genes that contribute to canine genetic disease. We investigated 222 SNPs from 42 genes that have been associated or may be implicated in human Addison's disease. We conducted case-control analyses in 3 pedigree dog breeds (Labrador retriever: affected n = 30, unaffected = 76; Cocker Spaniel: affected n = 19, unaffected = 53; Springer spaniel: affected n = 26, unaffected = 46) and identified 8 associated alleles in genes COL4A4, OSBPL9, CTLA4, PTPN22, and STXBP5 in 3 pedigree breeds. Association with immune response genes PTPN22 and CTLA4 in certain breeds suggests an underlying immunopathogenesis of the disease. These results suggest that canine hypoadrenocorticism could be a useful model for studying comparative genetics relevant to human Addison's disease.
Assuntos
Insuficiência Adrenal/veterinária , Doenças do Cão/genética , Estudos de Associação Genética , Alelos , Animais , Sequência de Bases , Cruzamento , Mapeamento Cromossômico , Doenças do Cão/imunologia , Cães , Frequência do Gene , Genótipo , Haplótipos , Anotação de Sequência Molecular , Polimorfismo de Nucleotídeo ÚnicoRESUMO
OBJECTIVES: To estimate the annual incidence risk of leptospirosis diagnosis in practice-attending dogs in the UK during 2016 and identify risk factors for diagnosis. METHODS: Incidence of leptospirosis diagnosis in dogs during 2016 was estimated from dogs in primary-care practices from the VetCompass Programme (n = 905,543). A case-control study of laboratory cases (n = 362) versus VetCompass controls explored factors (age, sex, neutering, breed, Kennel Club group, urban-rural location, indices of deprivation) associated with leptospirosis diagnosis through multivariable logistic regression. RESULTS: Annual incidence risk of leptospirosis in the VetCompass population was 0.8 cases per 100,000 dogs (0.0008%, 95% CI 9.1 × 10-8 -5.2 × 10-5 ). Adult dogs, especially 1- < 5 years olds (odds ratio [OR] = 0.38, 95% CI 0.27-0.54), and dogs attending urban clinics (OR = 0.26, 95% CI 0.19-0.35) had reduced odds of leptospirosis versus dogs < 1 year old and rural dogs, respectively. Dogs attending clinics in less deprived areas had increased odds of diagnosis (OR = 3.63, 95% CI 2.28-5.78) compared to crossbreds, Cocker Spaniels (OR = 4.25, 95% CI 2.65-6.84), Collies (OR = 3.53, 95% CI 2.22-5.62) and Lurchers (OR = 3.49, 95% CI 1.50-8.11) had increased odds of diagnosis. DISCUSSION: Leptospirosis is rarely diagnosed in clinical practice, suggesting that many true cases may be missed. Demographic risk factors identified here may inform the index of suspicion and encourage increased use of confirmatory diagnostic testing.
Assuntos
Doenças do Cão , Leptospirose , Animais , Estudos de Casos e Controles , Doenças do Cão/epidemiologia , Cães , Incidência , Leptospirose/epidemiologia , Leptospirose/veterinária , Fatores de Risco , Reino Unido/epidemiologiaRESUMO
In-vivo models of Actinobacillus pleuropneumoniae (App) infection in pigs are required for the development of vaccines and investigations of pathogenicity. Existing models cause severe respiratory disease with pulmonary oedema, dyspnoea and severe thoracic pain, and careful monitoring and early intervention with euthanasia is, therefore, needed to avoid unnecessary suffering in experimental animals. As a potential replacement for the existing respiratory infection model, an in-vivo protocol was evaluated using intradermal or subcutaneous injection of different App strains and Apx toxins into the abdominal skin of pigs. High concentrations of serovar 1 and serovar 10 App induced diffuse visible dermal oedema and inflammation. Injection of Apx toxins alone did not adequately produce macroscopic lesions, although an influx of inflammatory cells was seen on histopathology. ApxI-producing strains of App induced more inflammation than ApxII- and ApxIII-producing strains. Induction of skin lesions by injection of App or Apx toxins was not sufficiently repeatable or discrete for a robust experimental model that could be used for assessment of novel interventions.
Assuntos
Infecções por Actinobacillus , Actinobacillus pleuropneumoniae , Toxinas Bacterianas , Doenças dos Suínos , Infecções por Actinobacillus/prevenção & controle , Infecções por Actinobacillus/veterinária , Animais , Proteínas de Bactérias , Edema/veterinária , Proteínas Hemolisinas , Inflamação/veterinária , Modelos Teóricos , SuínosRESUMO
This two-part article discusses the mechanisms by which genetic variation can influence the risk of complex diseases, with a focus on canine diabetes mellitus. In Part 1, presented here, the importance of accurate methods for classifying different types of diabetes will be discussed, since this underpins the selection of cases and controls for genetic studies. Part 2 will focus on our current understanding of the genes involved in diabetes risk, and the way in which new genome sequencing technologies are poised to reveal new diabetes genes in veterinary species.
Assuntos
Diabetes Mellitus/veterinária , Doenças do Cão/genética , Predisposição Genética para Doença , Fenótipo , Animais , Diabetes Mellitus/classificação , Diabetes Mellitus/genética , Doenças do Cão/imunologia , Cães , Variação Genética , Sequenciamento de Nucleotídeos em Larga Escala/veterinária , Humanos , Resistência à Insulina , Células Secretoras de Insulina/imunologia , Obesidade/veterinária , Especificidade da EspécieRESUMO
Part 1 of this 2-part review outlined the importance of disease classification in diabetes genetic studies, as well as the ways in which genetic variants may contribute to risk of a complex disease within an individual, or within a particular group of individuals. Part 2, presented here, describes in more detail our current understanding of the genetics of canine diabetes mellitus compared to our knowledge of the human disease. Ongoing work to improve our knowledge, using new technologies, is also introduced.
Assuntos
Diabetes Mellitus/veterinária , Doenças do Cão/genética , Predisposição Genética para Doença/genética , Animais , Diabetes Mellitus/classificação , Diabetes Mellitus/genética , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/imunologia , Doenças do Cão/classificação , Doenças do Cão/imunologia , Cães , Antígenos HLA/genética , Antígenos HLA/imunologia , Sequenciamento de Nucleotídeos em Larga Escala/veterinária , Humanos , Imunidade/genética , Células Secretoras de Insulina/imunologia , Complexo Principal de Histocompatibilidade/genética , MutaçãoRESUMO
Diabetes mellitus (DM) is a common feline endocrinopathy, which is similar to human type 2 diabetes (T2DM) in terms of its pathophysiology. T2DM occurs due to peripheral insulin resistance and/or ß-cell dysfunction. Several studies have identified genetic and environmental factors that contribute to susceptibility to human T2DM. In cats, environmental factors such as obesity and physical inactivity have been linked with DM, although to date, the only genetic association that has been demonstrated is with a polymorphism in the feline MC4R gene. The aim of this study was to perform a genome-wide association study (GWAS) to identify polymorphisms associated with feline DM. Illumina Infinium 63k iSelect DNA arrays were used to analyse genomic DNA samples from 200 diabetic domestic shorthair cats and 399 non-diabetic control cats. Data was analysed using PLINK whole genome data analysis toolset. A linear model analysis, EMMAX, was done to test for population structure and HAPLOVIEW was used to identify haplotype blocks surrounding the significant SNPs to assist with candidate gene nomination. A total of 47,497 SNPs were available for analysis. Four SNPs were identified with genome-wide significance: chrA2.4150731 (praw = 9.94 x10-8); chrUn17.115508 (praw = 6.51 x10-8); chrUn17.394136 (praw = 2.53 x10-8); chrUn17.314128 (praw = 2.53 x10-8) as being associated with DM. The first SNP is located within chromosome A2, less than 4kb upstream of the dipeptidyl-peptidase-9 (DPP9) gene, a peptidase involved in incretin inactivation. The remaining three SNPs are located within a haplotype block towards the end of chromosome A3; within this region, genes of interest include TMEM18 and ACP1, both previously associated with T2DM. This study indicates a polygenic component to susceptibility to DM in cats and has highlighted several loci and candidate genes worthy of further investigation.
Assuntos
Doenças do Gato/genética , Diabetes Mellitus/veterinária , Estudo de Associação Genômica Ampla/veterinária , Polimorfismo de Nucleotídeo Único , Animais , Estudos de Casos e Controles , Gatos , Mapeamento Cromossômico , Diabetes Mellitus/genética , Dipeptidil Peptidases e Tripeptidil Peptidases/genética , Feminino , Predisposição Genética para Doença , Masculino , Proteínas de Membrana/genética , Proteínas Tirosina Fosfatases/genéticaRESUMO
BACKGROUND: Canine diabetes mellitus has mostly been studied in northern European, Australian and American populations, whereas other regions have received less attention. OBJECTIVES: We evaluated the epidemiological, clinical and histopathological features of diabetic dogs in Gran Canaria, Spain. METHODS: Prevalence and incidence were estimated. Clinical features were analysed, and serum and genomic DNA were obtained. Dogs with presumed idiopathic or immune-mediated diabetes, were DLA-typed and antibodies against GAD65 and IA-2 were assessed. Pancreases from ten diabetic dogs were examined and compared with pancreases from non-diabetic dogs. RESULTS AND CONCLUSIONS: Twenty-nine diabetic dogs were identified in a population of 5,213 (prevalence: 0.56%; incidence: 0.37%). Most were female (79%) and sexually intact (87% of females, 83% of males). Diabetes secondary to dioestrus (55.2%) and insulin-deficient diabetes (20.7%) were the most frequent types. Antibodies against GAD65 and IA-2 were identified in two out of five cases and DLA-genotyping revealed novel haplotypes. Breed distribution differed between diabetic and non-diabetic dogs. Reduced number of pancreatic islets and ß-cell mass were observed, with vacuolation of islet cells and ductal epithelium. In this population, where neutering is not standard practice, diabetes secondary to dioestrus is the most frequent diabetes subtype. Genetic susceptibility also differed from previous studies. These results support the heterogeneous pathogenesis of canine diabetes.
Assuntos
Diabetes Mellitus/veterinária , Doenças do Cão/epidemiologia , Animais , Diabetes Mellitus/epidemiologia , Diabetes Mellitus/etiologia , Doenças do Cão/etiologia , Cães , Feminino , Incidência , Ilhas/epidemiologia , Masculino , Prevalência , Espanha/epidemiologiaRESUMO
BACKGROUND: Canine hypoadrenocorticism is an immune-mediated endocrinopathy that shares both clinical and pathophysiological similarities with Addison's disease in humans. Several dog breeds are overrepresented in the disease population, suggesting that a genetic component is involved, although this is likely to be polygenic. Previous research has implicated CTLA4 as a potential susceptibility gene. CTLA4 is an important regulator of T cell function and polymorphisms/mutations in CTLA4 have been associated with a number of autoimmune phenotypes in both humans and rodent models of autoimmunity. The aim of the current study was to undertake a case:control association study of CTLA4 promotor polymorphisms in three dog breeds, cocker spaniels, springer spaniels and West Highland white terriers (WHWT). RESULTS: Polymorphisms in the CTLA4 promoter were determined by PCR and sequence-based typing. There were significant associations with three promoter haplotypes in cocker spaniels (p = 0.003). A series of SNPs were also associated with hypoadrenocorticism in cocker spaniels and springer spaniels, including polymorphisms in predicted NFAT and SP1 transcription factor binding sites. CONCLUSIONS: This study provides further evidence that CTLA4 promotor polymorphisms are associated with this complex genetic disease and supports an immune mediated aetiopathogenesis of canine hypoadrenocorticism.
RESUMO
Insulin-deficiency diabetes in dogs shares some similarities with human latent autoimmune diabetes of adults (LADA). Canine diabetes is likely to have a complex pathogenesis with multiple genes contributing to overall susceptibility and/or disease progression. An association has previously been shown between canine diabetes and MHC class II genes, although other genes are also likely to contribute to the genetic risk. Potential diabetes susceptibility genes include immuno-regulatory TH1/TH2 cytokines such as IFNgamma, IL-12, IL-4 and IL-10. We screened these candidate genes for single nucleotide polymorphisms (SNPs) in a range of different dog breeds using dHPLC analysis and DNA sequencing. Thirty-eight of the SNPs were genotyped in crossbreed dogs and seven other breed groups (Labrador Retriever, West Highland White Terrier, Collie, Schnauzer, Cairn Terrier, Samoyed and Cavalier King Charles Spaniel), which demonstrated substantial intra-breed differences in allele frequencies. When SNPs were examined for an association with diabetes by case:control analysis significant associations were observed for IL-4 in three breeds, the Collie, Cairn Terrier and Schnauzer and for IL-10 in the Cavalier King Charles Spaniel. These results suggest that canine cytokine genes regulating the TH1/TH2 immune balance might play a contributory role in determining susceptibility to diabetes in some breeds.
Assuntos
Citocinas/genética , Diabetes Mellitus/veterinária , Doenças do Cão/genética , Polimorfismo de Nucleotídeo Único , Linfócitos T/imunologia , Animais , Estudos de Casos e Controles , Citocinas/metabolismo , Diabetes Mellitus/genética , Cães , Frequência do Gene , Predisposição Genética para Doença , GenótipoRESUMO
OBJECTIVE-To characterize variability in melanoma-associated antigen (MAA) genes and gene expression in melanomas of dogs. ANIMALS-18 dogs with malignant melanomas and 8 healthy control dogs. PROCEDURES-cDNA was prepared from malignant melanoma biopsy specimens and from pigmented oral mucocutaneous tissues of healthy control dogs. Genomic DNA was extracted from poorly pigmented melanomas. A PCR assay was performed by use of Melan-A, SILV, or tyrosinase-specific primers. RESULTS-Splice variants of Melan-A and SILV were identified in malignant melanomas and also in healthy pigmented tissues, whereas a tyrosinase splice variant was detected in melanoma tissues only. A short interspersed nuclear element (SINE) insertion mutation was identified in the SILV gene in 1 of 10 poorly pigmented melanomas. Six novel exonic single nucleotide polymorphisms (SNPs; 3 synonymous and 3 nonsynonymous) were detected in the tyrosinase gene, and 1 nonsynonymous exonic SNP was detected in the SILV gene. CONCLUSIONS AND CLINICAL RELEVANCE-Variants of MAA mRNA were detected in malignant melanoma tissues of dogs. The importance of MAA alternative transcripts expressed in melanomas and normal pigmented tissues was unclear, but they may have represented a means of regulating melanin synthesis. The tyrosinase splice variant was detected only in melanomas and could potentially be a tumor-specific target for immunotherapy. A SILV SINE insertion mutation was identified in a melanoma from a Great Dane, a breed known to carry this mutation (associated with merle coat color). The nonsynonymous SNPs detected in tyrosinase and SILV transcripts did not appear to affect tumor pigmentation.
Assuntos
Antígenos de Neoplasias/genética , Doenças do Cão/metabolismo , Regulação Neoplásica da Expressão Gênica/fisiologia , Melanoma/veterinária , RNA Mensageiro/metabolismo , Animais , Antígenos de Neoplasias/metabolismo , Sequência de Bases , Doenças do Cão/genética , Cães , Variação Genética , Melanoma/metabolismo , Dados de Sequência Molecular , Isoformas de Proteínas , RNA Mensageiro/genéticaRESUMO
The outcome of infection with Leishmania infantum in dogs is variable, which is thought to be due to the nature of the immune response mounted by the host. As a consequence, the clinical signs and severity of canine leishmaniosis vary between individual dogs. Host immunogenetic factors might play an important role in determining the outcome of infection. The aim of this study was to examine polymorphisms in innate and adaptive immune response genes, to determine whether any of these were associated with susceptibility or resistance to L. infantum infection. Genomic DNA was obtained from two groups: pet dogs in endemic regions of Europe and a group of Beagles exposed to sand fly infection as part of a vaccine study. Genotyping was performed using a SNP (single nucleotide polymorphism) array for selected immune response genes. The first part of the study compared 62 clinical cases with 101 clinically unaffected dogs that were seronegative for Leishmania antibodies. One SNP in the CIITA gene demonstrated a significantly higher minor allele frequency in the case group, compared with the control group at the individual SNP level after permutation, but was not significant after correction for multiple testing. The second part of the study examined 48 Beagle dogs exposed to L. infantum over two transmission seasons. Twenty-seven dogs with a resistant phenotype (no evidence of clinical disease, seronegative at the end of the study period, negative on lymph node culture and only transiently PCR positive in bone marrow) were compared with 21 dogs demonstrating a susceptible phenotype (clinical disease, seropositive, positive lymph node culture and consistently PCR positive in bone marrow). Three SNPs in TLR3, two SNPs in PTPN22 and one SNP in TLR4 and IL1A were associated with the susceptible phenotype in the Beagle group at the individual SNP level after permutation analysis, but were not significant after correction for multiple testing. Further validation of these SNPs is required in a larger cohort of dogs, ideally with extreme phenotypes to confirm an association with the outcome of L. infantum infection.
Assuntos
Resistência à Doença/genética , Doenças do Cão/imunologia , Leishmania infantum/imunologia , Leishmaniose/veterinária , Polimorfismo de Nucleotídeo Único/genética , Psychodidae/parasitologia , Imunidade Adaptativa/genética , Animais , Doenças do Cão/epidemiologia , Doenças do Cão/parasitologia , Cães , Doenças Endêmicas/veterinária , Europa (Continente)/epidemiologia , Imunidade Inata/genética , Leishmania infantum/genética , Leishmaniose/epidemiologia , Leishmaniose/imunologia , Leishmaniose/parasitologia , Reação em Cadeia da Polimerase/veterináriaRESUMO
Pattern-recognition receptors (PRRs) are important components of the innate immune system, enabling early detection of infection. Defective PRR function has been implicated in several infectious and immune-mediated diseases of human beings, including Crohn's disease (CD). Anal furunculosis (AF) is an immune-mediated disease which primarily occurs in German shepherd dogs (GSD) and could result from a similar type of PRR dysfunction. The aim of the current study was to investigate canine PRR responses in vitro and to test the hypothesis that these were altered in AF-affected GSD. The pattern-recognition receptors TLR1, TLR2, TLR4, TLR6, TLR9, NOD1 (nucleotide-binding oligomerisation domain) and NOD2 were evaluated in the DH82 canine monocyte/macrophage cell line. These cells were found to express mRNA for all the selected PRRs with TLR2 mRNA the most and TLR5 mRNA the least abundant. A similar pattern of expression was found in canine blood-derived monocyte/macrophages. Stimulation of DH82 cells and blood-derived monocyte/macrophages using specific PRR-ligands, resulted in expression of pro-inflammatory cytokine mRNA. Quantification of TNFalpha mRNA and protein secretion from stimulated cells demonstrated variable responses with lipopolysaccharide (TLR4 ligand) and PAM(3)CSK4 (TLR1/2 ligand) proving to be the most potent and CpG DNA (TLR9 ligand) the least potent. Comparing PRR responses in blood-derived monocyte/macrophages from healthy blood-donor dogs with those from AF-affected GSD showed a deficiency in the latter in response to LD-MDP (NOD2 ligand) at the mRNA level but not at the protein level. It is possible that dysfunctional NOD2 responses by cells of the monocyte/macrophage lineage are involved in the pathogenesis of AF.
Assuntos
Doenças do Ânus/veterinária , Doenças do Cão/imunologia , Furunculose/imunologia , Leucócitos Mononucleares/imunologia , Macrófagos/imunologia , Receptores de Reconhecimento de Padrão/genética , Animais , Doenças do Ânus/genética , Doenças do Ânus/imunologia , Doenças do Ânus/microbiologia , Linhagem Celular , Citocinas/genética , Citocinas/imunologia , Doenças do Cão/genética , Cães , Citometria de Fluxo/veterinária , Furunculose/genética , Furunculose/microbiologia , Imunidade Inata/imunologia , Ativação Linfocitária , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Receptores de Reconhecimento de Padrão/biossíntese , Receptores de Reconhecimento de Padrão/imunologia , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterináriaRESUMO
German Shepherd Dogs have an increased incidence of anal furunculosis (AF), which is a disease characterised by inflammation and ulceration of the perianal tissues. Ciclosporin, an immunosuppressive drug, has been successfully used to treat AF, suggesting that the pathogenesis of disease is likely to have an immune-mediated component. Previous research has shown that the cytokine mRNA profile in AF lesions is consistent with T cell-mediated inflammation. The aims of the current study were to quantify IL-2 and IFNgamma mRNA expression in AF biopsies taken before and after treatment with ciclosporin and to compare cytokine expression with lesion severity. Twenty-two dogs with AF were recruited into the study and lesional biopsies were taken prior to ciclosporin therapy. Lesion severity was graded using a visual analogue scale. All dogs were evaluated after 4 weeks of ciclosporin therapy and, in 10 dogs with persistent disease, residual lesions were resected. RNA was extracted from AF-lesional tissue and control perianal tissue samples (n=10), which was used as the template for RT-PCR. Analysis of IL-2 and IFNgamma mRNA expression was performed using real-time PCR. IL-2 and IFNgamma mRNA was consistently detected in pre-treatment AF biopsies and, when quantified, this was significantly increased compared to control tissue (P<0.05). However, no correlation was seen between lesion severity and pre-treatment cytokine mRNA expression. In the ten paired pre- and post-treatment samples, IL-2 mRNA expression was significantly reduced in residual disease tissue following ciclosporin therapy (P=0.013). Treatment with ciclosporin seemed to result in decreased expression of IFNgamma mRNA in AF lesions but this was not statistically significant. In six of the 10 dogs with persistent disease, T cell cytokine mRNA could still be detected in the tissues, suggesting that there was inadequate immunosuppression. The absence of a correlation between T cell cytokine expression and the severity of disease suggests that tissue destruction observed in AF might be a consequence of other inflammatory mediators or downstream effects of T cell activation.
Assuntos
Doenças do Ânus/veterinária , Ciclosporina/uso terapêutico , Doenças do Cão/imunologia , Furunculose/veterinária , Interferon gama/biossíntese , Interleucina-2/biossíntese , Animais , Doenças do Ânus/tratamento farmacológico , Doenças do Ânus/genética , Doenças do Ânus/imunologia , Biópsia/veterinária , Doenças do Cão/tratamento farmacológico , Doenças do Cão/genética , Cães , Feminino , Furunculose/tratamento farmacológico , Furunculose/genética , Furunculose/imunologia , Imunossupressores/uso terapêutico , Interferon gama/genética , Interleucina-2/genética , Masculino , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa/veterinária , Estatísticas não ParamétricasRESUMO
Interleukin-7 (IL-7) and its receptor (IL-7R) are essential for T cell development in the thymus, and changes in the IL-7/IL-7R pathway have been implicated in age-associated thymic involution which results in a reduction of naïve T cell output. The aim of this study was to investigate the relationship between IL7 and IL7R genetic variation and thymic output in dogs. No single nucleotide polymorphisms (SNPs) were identified in the canine IL7 gene, but a number were present in the canine IL7R gene. Polymorphisms in the IL7R exon 8 and 3'UTR were found to be associated with signal joint T cell receptor excision circle (sj-TREC) values (a biomarker of thymic output) in young and geriatric Labrador retrievers. Additionally, one of the SNPs in the IL7R 3'UTR (SNP 14 c.1371 + 446 A > C) was found to cause a change in the seed-binding site for microRNA 185 which, a luciferase reporter assay demonstrated, caused changes in post-transcriptional regulation, and therefore might be capable of influencing IL-7R expression. The research findings suggest a genetic link between IL7R genotype and thymic output in dogs, which might impact on immune function as these animals age and provide further evidence of the involvement of IL-7/IL-7R pathway in age-associated thymic involution.
Assuntos
Regiões 3' não Traduzidas/genética , Genótipo , Subunidade alfa de Receptor de Interleucina-7/genética , MicroRNAs/genética , Receptores de Antígenos de Linfócitos T/genética , Linfócitos T/fisiologia , Timo/fisiologia , Animais , Cruzamento , Diferenciação Celular , Seleção Clonal Mediada por Antígeno , Cães , Interleucina-7/metabolismo , Subunidade alfa de Receptor de Interleucina-7/metabolismo , Polimorfismo de Nucleotídeo Único , Transdução de SinaisRESUMO
Immunosenescence is the gradual deterioration in immune system function associated with ageing. This decline is partly due to involution of the thymus, which leads to a reduction in the output of naive T cells into the circulating lymphocyte pool. Expansion of existing naive and memory T cell populations, to compensate for the reduction in thymic output, can lead to reduced diversity in the T cell repertoire with increasing age, resulting in impairment of immune responses to novel antigenic challenges, such as during infection and vaccination. Since associations between T cell repertoire and age have only been examined in a limited number of species, to gain further insights into this relationship, we have investigated age-related changes in the canine T cell receptor (TCR) repertoire. Blood samples were obtained from Labrador retriever dogs of varying ages and variation in the complementary determining region 3 (CDR3) of the T cell receptor beta (TCRB) chain was investigated. CDR3 size spectratyping was employed to evaluate clonal expansion/deletion in the T cell repertoire, allowing identification of profiles within individual variable (V) region families that skewed away from a Gaussian distribution. Older dogs (10-13 years) were found to have an increased number of TCRB V gene spectratypes that demonstrated a skewed distribution, compared with young dogs (≤3 years). Additionally, there was a reduction in the number of clonal peaks present in the spectratypes of old dogs, compared with those of young dogs. The study findings suggest that there is an age-associated disturbance in the diversity of the T cell receptor repertoire in dogs.