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Mutations of p53 gene in canine sweat gland carcinomas probably associated with UV radiation
2021
Jasik, Agnieszka | Kycko, Anna | Olech, Monika | Wyrostek, Krzysztof | Smiech, Anna | Łopuszyński, Wojciech | Otrocka-Domagała, Iwona | Mikiewicz, Mateusz | Dolka, Izabella
Apocrine sweat gland carcinomas (ASGCs) are rare malignant skin tumours in dogs and humans. The literature published so far focuses mostly on the clinico-epidemiological aspect of these tumours, but little is known about their pathogenesis. In this study we aimed to determine whether the p53 gene is involved in the carcinogenesis of the apocrine sweat gland in dogs and whether ultraviolet radiation (UV) is related to it. Forty canine ASGCs were submitted to laser capture microdissection to isolate neoplastic cells, from which DNA was subsequently extracted. PCR amplification and sequencing of p53 exons 2–8 was then performed, followed by computer analysis of the obtained sequences. Sixteen mutations within the p53 gene were found in 13 tumours. The mutations involved C → T, T → C, G → A, and CC → TT transitions, C → G transversion and adenine deletion, which are gene alteration types known to be related to UV radiation in the process of skin carcinogenesis in humans. Six of the thirteen tumour cases displayed the C → T transitions in the same location in exon 4 and three of the thirteen cases displayed T → C in the same location in exon 5. The results of the present study indicate both the participation of the p53 gene and the influence of UV radiation in the formation of ASGCs in dogs.
Show more [+] Less [-]Mutation responsible for congenital photosensitivity and hyperbilirubinemia in Southdown sheep
2018
Posbergh, Christian J. | Kalla, Sara E. | Sutter, Nathan B. | Tennant, Bud C. | Huson, Heather J.
OBJECTIVE To identify the genetic cause for congenital photosensitivity and hyperbilirubinemia (CPH) in Southdown sheep. ANIMALS 73 Southdown sheep from a CPH research flock and 48 sheep of various breeds from commercial flocks without CPH. PROCEDURES Whole-genome sequencing was performed for a phenotypically normal Southdown sheep heterozygous for CPH. Heterozygous variants within Slco1b3 coding exons were identified, and exons that contained candidate mutations were amplified by PCR assay methods for Sanger sequencing. Blood samples from the other 72 Southdown sheep of the CPH research flock were used to determine plasma direct and indirect bilirubin concentrations. Southdown sheep with a plasma total bilirubin concentration < 0.3 mg/dL were classified as controls, and those with a total bilirubin concentration ≥ 0.3 mg/dL and signs of photosensitivity were classified as mutants. Sanger sequencing was used to determine the Slco1b3 genotype for all sheep. Genotypes were compared between mutants and controls of the CPH research flock and among all sheep. Protein homology was measured across 8 species to detect evolutionary conservation of Slco1b. RESULTS A nonsynonymous mutation at ovine Chr3:193,691,195, which generated a glycine-to-arginine amino acid change within the predicted Slco1b3 protein, was significantly associated with hyperbilirubinemia and predicted to be deleterious. That amino acid was conserved across 7 other mammalian species. CONCLUSIONS AND CLINICAL RELEVANCE Results suggested a nonsynonymous mutation in Slco1b3 causes CPH in Southdown sheep. This disease appears to be similar to Rotor syndrome in humans. Sheep with CPH might be useful animals for Rotor syndrome research.
Show more [+] Less [-]ACTN3 gene variants as potential phenotype and performance biomarkers in Brazilian sport horses training for eventing in a tropical climate
2018
Padilha, Felipe Gomes Ferreira | El-Jaick, Kênia Balbi | Castro, Liane de | Santos Moreira, A dos | Ferreira, Ana Maria Reis | Almeida, Fernando Queiroz de
The aim of this study was to look for mutations in the equine ACTN3 gene and to identify sequence variants that might be associated with the phenotype and performance of Brazilian sport horses training for events in a tropical climate. Among 17 such horses direct DNA sequencing and mutation analysis of the exon 15 and the intron-exon boundaries of ACTN3 revealed 2 new sequence variants in the ACTN3 intron 14-15, designated c.1681-86G > A and c.1681-129delA. Wild-type/deletion heterozygotes (A/del) had a lower mean subcutaneous fat layer in the region of the gluteus medius, as measured by ultrasonography, than the del/del homozygotes; the correlation was significant (P = 0.017). This single base-pair deletion in ACTN3 intron 14-15 may have resulted in metabolic changes that led to increased deposition of body fat in the homozygous state. However, neither sequence variant was correlated with the time to fatigue in a test on a high-speed treadmill with an incremental-speed protocol.
Show more [+] Less [-]Evaluation of vincristine-associated myelosuppression in Border Collies
2013
Lind, Denise L. | Fidel, Janean (Janean L) | Gay, John M. | Mealey, Katrina L.
Objective: To determine whether Border Collies (ATP binding cassette subfamily B1 gene [ABCB1] wildtype) were more likely than other breeds to develop vincristine-associated myelosuppression (VAM) and, if so, whether this was caused by a mutation in ABCB1 distinct from ABCB1-1Δ. Animals: Phase 1 comprised 36 dogs with the ABCB1 wildtype, including 26 dogs with lymphoma (5 Border Collies and 21 dogs representing 13 other breeds) treated with vincristine in a previous study; phase 2 comprised 10 additional Border Collies, including 3 that developed VAM and 7 with an unknown phenotype. Procedures: For phase 1, the prevalence of VAM in ABCB1-wildtype Border Collies was compared with that for ABCB1-wildtype dogs of other breeds with data from a previous study. For phase 2, additional Border Collies were included. Hematologic adverse reactions were graded with Veterinary Co-operative Oncology Group criteria. Genomic DNA was used to amplify and sequence all 27 exons of the canine ABCB1. Sequences from affected dogs were compared with those of unaffected dogs and dogs of unknown phenotype. Results: 3 of 5 Border Collies with the ABCB1 wildtype developed VAM; this was significantly higher than the proportion of other dogs that developed VAM (0/21). A causative mutation for VAM in Border Collies was not identified, although 8 single nucleotide polymorphisms in ABCB1 were detected. Conclusions and Clinical Relevance: Breed-associated sensitivity to vincristine unrelated to ABCB1 was detected in Border Collies. Veterinarians should be aware of this breed predisposition to VAM. Causes for this apparent breed-associated sensitivity should be explored.
Show more [+] Less [-]Evaluation of dysregulation of the receptor tyrosine kinases Kit, Flt3, and Met in histiocytic sarcomas of dogs
2006
Zavodovskaya, R. | Liao, A.T. | Jones, C.L.R. | Yip, B. | Chien, M.B. | Moore, P.F. | London, C.A.
Objective-To evaluate canine histiocytic sarcoma cell lines and tumor samples for dysregulation of the Kit/stem-cell factor (SCF), Flt3/Flt3 ligand (Flt3L), and Met/hepatocyte growth factor (HGF) receptor tyrosine kinase signaling pathways, as these are known to contribute to the differentiation and survival of normal dendritic cells as well as malignant transformation of dendritic cells in mouse models. Sample Population-4 histiocytic sarcoma tumor cell lines and 35 formalin-fixed histiocytic sarcoma specimens obtained from dogs. Procedure-Histiocytic sarcoma cell lines were evaluated for expression of Kit/SCF, Flt3/Flt3L, and Met/HGF by use of reverse transcriptase-PCR procedures. Histiocytic sarcoma cell lines and tumor samples were evaluated for mutations in Kit, Flt3, and Met by use of PCR analysis of genomic DNA, followed by both sequencing and fluorescent PAGE for deletions or internal tandem duplications. The ability of the multitargeted split-kinase inhibitor SU11654 to block proliferation and induce apoptosis of histiocytic sarcoma cell lines was also evaluated. Results-No mutations in Kit, Flt3, and Met were identified in any of the cell lines or tumor samples evaluated. Furthermore, SU11654 did not induce cellcycle arrest or apoptosis of histiocytic sarcoma lines, even at supratherapeutic doses. Conclusions and Clinical Relevance-These data suggest that dysregulation of Kit/SCF, Flt3/Flt3L, and Met/HGF signaling pathways is unlikely to occur in histiocytic sarcomas of dogs and that inhibitors of the Kit, Flt3, and Met pathways are unlikely to provide clinical benefit to dogs with histiocytic sarcomas.
Show more [+] Less [-]Prognostic value of CD44 variant isoform expression in dogs with multicentric high-grade B-cell lymphoma
2018
Motegi, Tomoki | Tomiyasu, Hirotaka | Goto-Koshino, Yuko | Takahashi, Masashi | Hiyoshi-Kanemoto, Saaya | Fujino, Yasuhito | Ohno, Koichi | Tsuimoto, Hajime
OBJECTIVE To determine the prognostic value of CD44 variant isoform expression in dogs with multicentric high-grade B-cell lymphoma (BCL). ANIMALS 45 dogs with multicentric BCL and 10 healthy control Beagles. PROCEDURES The medical record database of a veterinary teaching hospital was searched to identify dogs with BCL that were treated between November 2005 and April 2015. Information regarding overall response to chemotherapy, progression-free survival (PFS) time, and overall survival time was extracted from each record. Archived lymph node aspirate specimens from dogs with BCL and lymph node aspirate specimens from the 10 control dogs underwent real-time PCR analysis to determine mRNA expression of CD44 variant isoforms of exons 3, 6, and 7 and the CD44 whole isoform. For each isoform, mRNA expression was compared between dogs with BCL and control dogs. The mean relative expression of each isoform was used to classify dogs with BCL into either a high- or low-expression group, and overall response rate, PFS time, and overall survival time (ie, indices of prognosis) were compared between the 2 groups. RESULTS For all isoforms evaluated, mean relative mRNA expression for dogs with BCL was numerically lower than that for control dogs. Dogs with BCL and high CD44 isoform expression had a lower overall response rate, median PFS time, and median overall survival time, compared with dogs with BCL and low CD44 isoform expression. CONCLUSIONS AND CLINICAL RELEVANCE Results indicated that, for dogs with BCL, high expression of exons 3, 6, and 7 was associated with a poor prognosis.
Show more [+] Less [-]Evaluation of ADAMTS17 in Chinese Shar-Pei with primary open-angle glaucoma, primary lens luxation, or both
2018
Oliver, James A. C. | Rustidge, Sophie | Pettitt, Louise | Jenkins, Christopher A. | Farias, Fabiana H. G. | Giuliano, Elizabeth A. | Mellersh, Cathryn S.
OBJECTIVE To evaluate the coding regions of ADAMTS17 for potential mutations in Chinese Shar-Pei with a diagnosis of primary open-angle glaucoma (POAG), primary lens luxation (PLL), or both. ANIMALS 63 Shar-Pei and 96 dogs of other breeds. PROCEDURES ADAMTS17 exon resequencing was performed on buccal mucosal DNA from 10 Shar-Pei with a diagnosis of POAG, PLL, or both (affected dogs). A candidate causal variant sequence was identified, and additional dogs (53 Shar-Pei [11 affected and 42 unaffected] and 95 dogs of other breeds) were genotyped for the variant sequence by amplified fragment length polymorphism analysis. Total RNA was extracted from ocular tissues of 1 affected Shar-Pei and 1 ophthalmologically normal Golden Retriever; ADAMTS17 cDNA was reverse transcribed and sequenced, and ADAMTS17 expression was evaluated by quantitative reverse-transcription PCR assay. RESULTS All affected Shar-Pei were homozygous for a 6-bp deletion in exon 22 of ADAMTS17 predicted to affect the resultant protein. All unaffected Shar-Pei were heterozygous or homozygous for the wild-type allele. The variant sequence was significantly associated with affected status (diagnosis of POAG, PLL, or both). All dogs of other breeds were homozygous for the wild-type allele. The cDNA sequencing confirmed presence of the expected variant mRNA sequence in ocular tissue from the affected dog only. Gene expression analysis revealed a 4.24-fold decrease in the expression of ADAMTS17 in ocular tissue from the affected dog. CONCLUSIONS AND CLINICAL RELEVANCE Results supported that the phenotype (diagnosis of POAG, PLL, or both) is an autosomal recessive trait in Shar-Pei significantly associated with the identified mutation in ADAMTS17.
Show more [+] Less [-]Evaluation of the dynactin 1 gene in Leonbergers and Labrador Retrievers with laryngeal paralysis
2016
Holt, David E. | Brown, Dorothy C. | Henthorn, Paula S.
OBJECTIVE To sequence exons and splice consensus sites of the dynactin subunit 1 (DCTN1) gene in Leonbergers and Labrador Retrievers with clinical laryngeal paralysis. ANIMALS 5 unrelated Leonbergers with laryngeal paralysis, 2 clinically normal Leonbergers, 7 unrelated Labrador Retrievers with laryngeal paralysis, and 2 clinically normal Labrador Retrievers. PROCEDURES Primers were designed for the entire coding regions of the DCTN1 gene, a noncoding exon at the 5´ end of the gene, and a 900-bp single-nucleotide polymorphism (SNP)-rich region located 17 kb upstream of the DCTN1 gene by use of the CanFam3 assembly of the canine genome sequence. Sequences were generated and compared between clinically normal and affected dogs. The SNPs flanking the DCTN1 gene as well as a previously identified nonsynonymous SNP in exon 32 were genotyped in affected and clinically normal Leonbergers and Labrador Retrievers. RESULTS None of the affected dogs were homozygous for any mutation affecting coding regions or splicing consensus sequences. Of the 16 dogs tested for the missense SNP in exon 32, all were homozygous for the reference allele, except for 2 affected and 1 clinically normal Labrador Retriever and 1 clinically normal Leonberger. The DCTN1 gene sequences (5 dogs) and haplotypes of polymorphic markers surrounding the DCTN1 gene (all dogs) were not consistent with the hypothesis that laryngeal paralysis was associated with inheritance of the same DCTN1 disease-causing allele within all Labrador Retrievers or Leonbergers evaluated. CONCLUSIONS AND CLINICAL RELEVANCE Mutations in the DCTN1 gene did not appear to cause laryngeal paralysis in Leonbergers or Labrador Retrievers.
Show more [+] Less [-]Characterization of the cDNA and genomic DNA sequence encoding for the platelet integrin alpha IIB and beta III in a horse with Glanzmann thrombasthenia
2011
Macieira, Susana | Lussier, Jacques | Bédard, Christian
Glanzmann thrombasthenia (GT) is characterized by a defect of platelet aggregation. This autosomal recessive genetic disorder is caused by an abnormality of the platelet glycoprotein receptors alpha IIb or beta III. Recently, we identified a horse with clinical and pathological features of GT. The aim of this study was to describe this case of GT at the molecular level. A point mutation from G to C in exon 2 of ITGA2B causing a substitution of the expected amino acid arginine 72 (Arg72) by a proline (Pro72) was encountered. This amino acid change may result in abnormal structural conformations that yield an inactive alpha IIb subunit. The genomic DNA analysis showed that this horse was homozygous for the missense mutation.
Show more [+] Less [-]Prevalence and importance of internal tandem duplications in exons 11 and 12 of c- kit in mast cell tumors of dogs
2002
Downing, Sue | Chien, May B. | Kass, Philip H. | Moore, Peter F. | London, Cheryl A.
Objective-To determine the prevalence of activating internal tandem duplications (ITDs) in exons 11 and 12 of c-kit in mast cell tumors (MCTs) of dogs and to correlate these mutations with prognosis. Sample Population-157 formalin-fixed, paraffinembedded MCTs from dogs in the pathology database of the Veterinary Medical Teaching Hospital at the University of California, Davis. Procedure-Genomic DNA was isolated from tumor specimens and a polymerase chain reaction procedure was performed to determine whether there were ITDs in exons 11 and 12. Results-We identified ITDs in 1 of 12 (8%) grade-I, 42 of 119 (35%) grade-II, and 9 of 26 (35%) grade-III tumors (overall prevalence, 52 of 157 [33%]). Logistic regression analysis revealed that the odds of grade-II and -III tumors possessing an ITD were approximately 5 times greater than that for grade-I tumors, although these odds did not differ significantly. Although MCTs possessing an ITD were twice as likely to recur after excision and twice as likely to result in metastasis as those without an ITD, these values also did not differ significantly. Conclusions and Clinical Relevance-These results provide evidence that ITDs in c-kit occur frequently in MCTs of dogs. The high prevalence of c-kit activating mutations in MCTs of dogs combined with the relative abundance of mast cell disease in dogs provide an ideal naturally developing tumor in which to test the safety and efficacy of novel small-molecule kinase inhibitors such as imatinib mesylate.
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