Safety of Intraperitoneal Injection of Adipose Tissue-Derived Autologous Mesenchymal Stem Cells in Cats

BACKGROUND: Chronic inflammatory diseases are common in cats and mesenchymal stem cells (MSC) are a promising therapeutic approach for management of these disorders. The purpose of this study was to evaluate the safety of intraperitoneal injection of MSC in cats. HYPOTHESIS: Intrapertioneal injection of autologous MSC in cats is safe. ANIMALS: Ten healthy adult purpose‐bred cats. METHODS: Mesenchymal stem cells were isolated from subcutaneous adipose tissue collected during ovariohysterectomy and characterized for expression of CD90, CD105 and CD44 and trilineage differentiation. Three weeks postoperatively a complete blood count, serum chemistry profile, urinalysis, and abdominal ultrasound were performed. Five cats then received 1 × 10(6) of autologous MSC/kg of body weight intraperitoneally with ultrasound guidance; 5 additional cats were sham injected. Cats were monitored for 6 weeks with daily physical examinations and weekly clinicopathological evaluations. Abdominal ultrasonography was repeated at weeks 1 and 5 after injection. RESULTS: Serious adverse effects were not observed in any MSC‐injected cat. Two animals developed transient lethargy and decreased activity. Jejunal lymph node size was increased in MSC‐injected cats compared to controls at weeks 1 (1.38 ± 0.25 versus 0.88 ± 0.25 cm(2); P = .036) and 5 (1.75 ± 0.82 versus 0.79 ± 0.12 cm(2); P = .047). A hyperechoic renal segmental cortical lesion was observed in 1 MSC‐injected cat. CONCLUSIONS AND CLINICAL RELEVANCE: Intraperitoneal MSC injection was well tolerated with only mild, self‐limiting adverse effects being observed in 2 cats. This route provides a safe means of administration for cell‐based treatment in cats

Comparative effectiveness of common therapies for Wilson disease: A systematic review and meta-analysis of controlled studies

BACKGROUND & AIMS: Wilson disease (WD) is a rare disorder of copper metabolism. The objective of this systematic review is to determine the comparative effectiveness and safety of common treatments of WD. METHODS: We included WD patients of any age or stage and the study drugs D-penicillamine, zinc salts, trientine, and tetrathiomolybdate. The control could be placebo, no treatment, or any other treatment. We included prospective, retrospective, randomized, and non-randomized studies. We searched Medline and Embase via Ovid, the Cochrane Central Register of Controlled Trials, and screened reference lists of included articles. Where possible, we applied random-effects meta-analyses. RESULTS: The 23 included studies reported on 2055 patients and mostly compared D-penicillamine to no treatment, zinc, trientine, or succimer. One study compared tetrathiomolybdate and trientine. Post-decoppering maintenance therapy was addressed in one study only. Eleven of 23 studies were of low quality. When compared to no treatment, D-penicillamine was associated with a lower mortality (odds ratio 0.013; 95% CI 0.0010 to 0.17). When compared to zinc, there was no association with mortality (odds ratio 0.73; 95% CI 0.16 to 3.40) and prevention or amelioration of clinical symptoms (odds ratio 0.84; 95% CI 0.48 to 1.48). Conversely, D-penicillamine may have a greater impact on side effects and treatment discontinuations than zinc. CONCLUSIONS: There are some indications that zinc is safer than D-penicillamine therapy while being similarly effective in preventing or reducing hepatic or neurologic WD symptoms. Study quality was low warranting cautious interpretation of our findings

Proposed Diagnostic Criteria and Classification of Canine Mast Cell Neoplasms: A Consensus Proposal

Mast cell neoplasms are one of the most frequently diagnosed malignancies in dogs. The clinical picture, course, and prognosis vary substantially among patients, depending on the anatomic site, grade and stage of the disease. The most frequently involved organ is the skin, followed by hematopoietic organs (lymph nodes, spleen, liver, and bone marrow) and mucosal sites of the oral cavity and the gastrointestinal tract. In cutaneous mast cell tumors, several grading and staging systems have been introduced. However, no comprehensive classification and no widely accepted diagnostic criteria have been proposed to date. To address these open issues and points we organized a Working Conference on canine mast cell neoplasms in Vienna in 2019. The outcomes of this meeting are summarized in this article. The proposed classification includes cutaneous mast cell tumors and their sub-variants defined by grading- and staging results, mucosal mast cell tumors, extracutaneous/extramucosal mast cell tumors without skin involvement, and mast cell leukemia (MCL). For each of these entities, diagnostic criteria are proposed. Moreover, we have refined grading and staging criteria for mast cell neoplasms in dogs based on consensus discussion. The criteria and classification proposed in this article should greatly facilitate diagnostic evaluation and prognostication in dogs with mast cell neoplasms and should thereby support management of these patients in daily practice and the conduct of clinical trials

A dual fluorescent multiprobe assay for prion protein genotyping in sheep

BACKGROUND: Scrapie and BSE belong to a group of fatal, transmissible, neurodegenerative diseases called TSE. In order to minimize the risk of natural scrapie and presumed natural BSE in sheep, breeding programmes towards TSE resistance are conducted in many countries based on resistance rendering PRNP polymorphisms at codons 136 (A/V), 154 (R/H) and 171 (R/H/Q). Therefore, a reliable, fast and cost-effective method for routine PRNP genotyping in sheep, applicable in standard equipped molecular genetic laboratories, will be a vital instrument to fulfill the need of genotyping hundreds or thousands of sheep. METHODS: A dual fluorescent multiprobe assay consisting of 2 closed tube PCR reactions containing respectively 4 and 3 dual-labelled fluorescent ASO probes for the detection in real-time of the 7 allelic variants of sheep PRNP mentioned above. RESULTS: The assay is succesfully performed using unpurified DNA as a template for PCR, without any post-PCR manipulations and with semi-automatic determination of the PRNP genotypes. The performance of the assay was confirmed via PCR-RFLP and sequencing in a cross-validation study with 50 sheep. CONCLUSIONS: We report the development and validation of a robust, reliable and reproducible method for PRNP genotyping of a few to many sheep samples in a fast, simple and cost-effective way, applicable in standard equipped molecular genetic laboratories. The described primer/probe design strategy can also be applied for the detection of other polymorphisms or disease causing mutations

MKLN1 splicing defect in dogs with lethal acrodermatitis

Lethal acrodermatitis (LAD) is a genodermatosis with monogenic autosomal recessive inheritance in Bull Terriers and Miniature Bull Terriers. The LAD phenotype is characterized by poor growth, immune deficiency, and skin lesions, especially at the paws. Utilizing a combination of genome wide association study and haplotype analysis, we mapped the LAD locus to a critical interval of similar to 1.11 Mb on chromosome 14. Whole genome sequencing of an LAD affected dog revealed a splice region variant in the MKLN1 gene that was not present in 191 control genomes (chr14:5,731,405T>G or MKLN/:c.400+3A>C). This variant showed perfect association in a larger combined Bull Terrier/Miniature Bull Terrier cohort of 46 cases and 294 controls. The variant was absent from 462 genetically diverse control dogs of 62 other dog breeds. RT-PCR analysis of skin RNA from an affected and a control dog demonstrated skipping of exon 4 in the MKLN1 transcripts of the LAD affected dog, which leads to a shift in the MKLN1 reading frame. MKLN1 encodes the widely expressed intracellular protein muskelin 1, for which diverse functions in cell adhesion, morphology, spreading, and intracellular transport processes are discussed. While the pathogenesis of LAD remains unclear, our data facilitate genetic testing of Bull Terriers and Miniature Bull Terriers to prevent the unintentional production of LAD affected dogs. This study may provide a starting point to further clarify the elusive physiological role of muskelin 1 in vivo.Peer reviewe

Global assessment of genomic variation in cattle by genome resequencing and high-throughput genotyping

<p>Abstract</p> <p>Background</p> <p>Integration of genomic variation with phenotypic information is an effective approach for uncovering genotype-phenotype associations. This requires an accurate identification of the different types of variation in individual genomes.</p> <p>Results</p> <p>We report the integration of the whole genome sequence of a single Holstein Friesian bull with data from single nucleotide polymorphism (SNP) and comparative genomic hybridization (CGH) array technologies to determine a comprehensive spectrum of genomic variation. The performance of resequencing SNP detection was assessed by combining SNPs that were identified to be either in identity by descent (IBD) or in copy number variation (CNV) with results from SNP array genotyping. Coding insertions and deletions (indels) were found to be enriched for size in multiples of 3 and were located near the N- and C-termini of proteins. For larger indels, a combination of split-read and read-pair approaches proved to be complementary in finding different signatures. CNVs were identified on the basis of the depth of sequenced reads, and by using SNP and CGH arrays.</p> <p>Conclusions</p> <p>Our results provide high resolution mapping of diverse classes of genomic variation in an individual bovine genome and demonstrate that structural variation surpasses sequence variation as the main component of genomic variability. Better accuracy of SNP detection was achieved with little loss of sensitivity when algorithms that implemented mapping quality were used. IBD regions were found to be instrumental for calculating resequencing SNP accuracy, while SNP detection within CNVs tended to be less reliable. CNV discovery was affected dramatically by platform resolution and coverage biases. The combined data for this study showed that at a moderate level of sequencing coverage, an ensemble of platforms and tools can be applied together to maximize the accurate detection of sequence and structural variants.</p

Canine models of copper toxicosis for understanding mammalian copper metabolism

Hereditary forms of copper toxicosis exist in man and dogs. In man, Wilson’s disease is the best studied disorder of copper overload, resulting from mutations in the gene coding for the copper transporter ATP7B. Forms of copper toxicosis for which no causal gene is known yet are recognized as well, often in young children. Although advances have been made in unraveling the genetic background of disorders of copper metabolism in man, many questions regarding disease mechanisms and copper homeostasis remain unanswered. Genetic studies in the Bedlington terrier, a dog breed affected with copper toxicosis, identified COMMD1, a gene that was previously unknown to be involved in copper metabolism. Besides the Bedlington terrier, a number of other dog breeds suffer from hereditary copper toxicosis and show similar phenotypes to humans with copper storage disorders. Unlike the heterogeneity of most human populations, the genetic structure within a purebred dog population is homogeneous, which is advantageous for unraveling the molecular genetics of complex diseases. This article reviews the work that has been done on the Bedlington terrier, summarizes what was learned from studies into COMMD1 function, describes hereditary copper toxicosis phenotypes in other dog breeds, and discusses the opportunities for genome-wide association studies on copper toxicosis in the dog to contribute to the understanding of mammalian copper metabolism and copper metabolism disorders in man