ABSTRACT
Tumours shows aberrant DNA methylation patterns, being hypermethylated or hypomethylated compared with normal tissues. In human acute myeloid leukaemia (hAML) mutations in DNA methyltransferase (DNMT3A) are associated to a more aggressive tumour behaviour. As AML is lethal in dogs, we defined global DNA methylation content, and screened the C-terminal domain of DNMT3 family of genes for sequence variants in 39 canine acute myeloid leukaemia (cAML) cases. A heterogeneous pattern of DNA methylation was found among cAML samples, with subsets of cases being hypermethylated or hypomethylated compared with healthy controls; four recurrent single nucleotide variations (SNVs) were found in DNMT3L gene. Although SNVs were not directly correlated to whole genome DNA methylation levels, all hypomethylated cAML cases were homozygous for the deleterious mutation at p.Arg222Trp. This study contributes to understand genetic modifications of cAML, leading up to studies that will elucidate the role of methylome alterations in the pathogenesis of AML in dogs.
Subject(s)
DNA Methylation/genetics , DNA Modification Methylases/genetics , Dog Diseases/genetics , Leukemia, Myeloid, Acute/veterinary , Animals , Case-Control Studies , Disease Models, Animal , Dogs , Female , Flow Cytometry/veterinary , Genetic Predisposition to Disease/genetics , Leukemia, Myeloid, Acute/genetics , MaleABSTRACT
OBJECTIVE: The prevalence of osteoarthritis (OA) varies between joints. Cartilage in eight different joints was evaluated to elucidate the disparate susceptibilities between joints to post-traumatic OA (PTOA) and provide evidence for joint-specific clinical treatments. The hypothesis was that cartilage in different joints would have varying cell death and anabolic gene expression profiles after injury. METHODS: Adult equine cartilage explants were harvested from shoulder (SH), elbow (EL), carpal (CA), metacarpophalangeal (MC), patellofemoral (FP), tarsal (TA), metatarsophalangeal (MT), and proximal interphalangeal (PP) joints, and injured by loading with 30 MPa within 1 s. Fractional dissipated energy, cell density, cell death, and gene expression were quantified. RESULTS: PP had the highest fractional dissipated energy (94%, 95% confidence interval [CI] 88 to 101%). Cell density was highest in the superficial zone in all samples, with MC and MT having the highest peak density. Injured samples had significantly increased cell death (13.5%, 95% CI 9.1 to 17.9%) than non-injured samples (6.8%, 95% CI 2.5 to 11.1%, P = 0.016); however, cell death after injury was not significantly different between joints. Gene expression was significantly different between joints. CD-RAP expression in normal cartilage was lowest in FP (Cp = 21, 95% CI -80 to 122). After injury, the change in CD-RAP expression increased and was highest in FP (147% relative increase after injury, 95% CI 64 to 213). CONCLUSION: Different joints have different baseline characteristics, including cell density and gene expression, and responses to injury, including energy dissipation and gene expression. These unique characteristics may explain differences in OA prevalence and suggest differences in susceptibility to PTOA. CLINICAL RELEVANCE: Understanding differences in the response to injury and potential susceptibility to OA can lead to the development of preventative or treatment strategies. KEY TERMS: Gene expression, cartilage injury, chondrocyte, multiphoton microscopy, cartilage biomechanical properties, PTOA. WHAT IS KNOWN ABOUT THE SUBJECT: The prevalence of OA is variable among joints; however, most laboratory studies are performed on a single joint - most commonly the knee, and extrapolated to other joints such as the ankle or shoulder. A small number of studies have compared knee and ankle cartilage and reported differences in mechanical properties and gene expression. WHAT THIS STUDY ADDS TO EXISTING KNOWLEDGE: There are differences in baseline cell density and gene expression, and differences in response to injury, including gene expression and cell death. This suggests that there are inherent differences leading to varying susceptibilities in OA prevalence among joints. Joint-specific treatments may improve OA therapies.
Subject(s)
Cartilage, Articular/injuries , Horse Diseases/physiopathology , Osteoarthritis/veterinary , Animals , Arthritis, Experimental/etiology , Arthritis, Experimental/pathology , Arthritis, Experimental/physiopathology , Cartilage, Articular/pathology , Cartilage, Articular/physiopathology , Cell Death , Chondrocytes/pathology , Gene Expression , Gene Expression Profiling/methods , Horse Diseases/etiology , Horse Diseases/pathology , Horses , Osteoarthritis/etiology , Osteoarthritis/pathology , Osteoarthritis/physiopathology , RNA, Messenger/genetics , Stress, MechanicalSubject(s)
Biomarkers, Tumor/genetics , Gene Deletion , Ikaros Transcription Factor/genetics , Mutation/genetics , Philadelphia Chromosome , Polymorphism, Single Nucleotide/genetics , Precursor Cell Lymphoblastic Leukemia-Lymphoma/genetics , Child , Cohort Studies , Humans , Precursor Cell Lymphoblastic Leukemia-Lymphoma/pathology , Prognosis , Sequence Analysis, DNAABSTRACT
Cell-based therapies, such as the use of mesenchymal stromal cells (MSCs), are becoming popular in veterinary medicine. When MSCs are not cryopreserved, they are shipped in suspension, but no previous studies have analyzed MSC viability during delivery. Here, the impact of several experimental shipping conditions on the number of equine blood-derived (ePB-MSC) and canine adipose-derived (cA-MSC) MSCs were evaluated. Among the different parameters tested, only time and temperature influenced MSC number during the experimental shipping conditions. Cells were monitored over different time intervals for gene expression of typical MSC markers and to evaluate acquired resistance to apoptosis and beta-galactosidase activity. Overall, these results indicate that ePB-MSC and cA-MSC should be delivered in phosphate buffered saline at room temperature and within 9-12 h.
Subject(s)
Culture Media/chemistry , Mesenchymal Stem Cells/physiology , Specimen Handling/veterinary , Animals , Cell Survival , Culture Media/pharmacology , Dogs , Gene Expression Regulation , Horses , Mesenchymal Stem Cells/drug effects , Real-Time Polymerase Chain Reaction , Specimen Handling/methods , Temperature , Time FactorsABSTRACT
Adult stem cells are nowadays used for treating several pathologies. A putative stem cell population was found in the adipose tissue of mammals and canine adipose tissue-derived-mesenchymal stem cells (cA-MSC) have been shown to possess the capacity to differentiate into several lineages. The main goal of our research was to fully characterize cA-MSC and examine the effects of cryopreservation on their stemness features. Each sample of cA-MSC was analyzed immediately and then again after being frozen in liquid nitrogen for one year. After the cryopreservation period cells conserved their fibroblast-like morphology, alkaline phosphatase positivity and CD expression but showed a lower proliferation ratio and a lower telomerase activity in comparison with fresh cells. Finally, the cryopreservation protocol did not change the cA-MSC adipogenic, osteogenic and myogenic differentiative potential. Our data demonstrate that stored cA-MSC might represent a promising type of progenitor cell for autologous cellular-based therapies in veterinary medicine.
