RESUMO
Feline leukemia virus (FeLV) is an exogenous retrovirus that causes malignant hematopoietic disorders in domestic cats, and its virulence may be closely associated with viral sequences. FeLV is classified into several subgroups, including A, B, C, D, E, and T, based on viral receptor interference properties or receptor usage. However, the transmission manner and disease specificity of the recombinant viruses FeLV-D and FeLV-B remain unclear. The aim of this study was to understand recombination events between exogenous and endogenous retroviruses within a host and elucidate the emergence and transmission of recombinant viruses. We observed multiple recombination events involving endogenous retroviruses (ERVs) in FeLV from a family of domestic cats kept in one house; two of these cats (ON-T and ON-C) presented with lymphoma and leukemia, respectively. Clonal integration of FeLV-D was observed in the ON-T case, suggesting an association with FeLV-D pathogenesis. Notably, the receptor usage of FeLV-B observed in ON-T was mediated by feline Pit1 and feline Pit2, whereas only feline Pit1 was used in ON-C. Furthermore, XR-FeLV, a recombinant FeLV containing an unrelated sequence referred to the X-region, which is homologous to a portion of the 5'-leader sequence of Felis catus endogenous gammaretrovirus 4 (FcERV-gamma4), was isolated. Genetic analysis suggested that most recombinant viruses occurred de novo; however, the possibility of FeLV-B transmission was also recognized in the family. This study demonstrated the occurrence of multiple recombination events between exogenous and endogenous retroviruses in domestic cats, highlighting the contribution of ERVs to pathogenic recombinant viruses.IMPORTANCEFeline leukemia virus subgroup A (FeLV-A) is primarily transmitted among cats. During viral transmission, genetic changes in the viral genome lead to the emergence of novel FeLV subgroups or variants with altered virulence. We isolated three FeLV subgroups (A, B, and D) and XR-FeLV from two cats and identified multiple recombination events in feline endogenous retroviruses (ERVs), such as enFeLV, ERV-DC, and FcERV-gamma4, which are present in the cat genome. This study highlights the pathogenic contribution of ERVs in the emergence of FeLV-B, FeLV-D, and XR-FeLV in a feline population.
Assuntos
Retrovirus Endógenos , Vírus da Leucemia Felina , Leucemia Felina , Animais , Gatos , Retrovirus Endógenos/genética , Vírus da Leucemia Felina/genética , Vírus da Leucemia Felina/fisiologia , Leucemia Felina/transmissão , Leucemia Felina/virologia , Recombinação GenéticaRESUMO
Estrogen receptor α (ERα) is a transcription factor that induces cell proliferation and exhibits increased expression in a large subset of breast cancers. The molecular mechanisms underlying the up-regulation of ERα activity, however, remain poorly understood. We identified FK506-binding protein 52 (FKBP52) as a factor associated with poor prognosis of individuals with ERα-positive breast cancer. We found that FKBP52 interacts with breast cancer susceptibility gene 1 and stabilizes ERα, and is essential for breast cancer cell proliferation. FKBP52 depletion resulted in decreased ERα expression and proliferation in breast cancer cell lines, including MCF7-derived fulvestrant resistance (MFR) cells, suggesting that inhibiting FKBP52 may provide a therapeutic effect for endocrine therapyresistant breast cancer. In contrast, FKBP51, a closely related molecule to FKBP52, reduced the stability of ERα. Consistent with these findings, FKBP51 was more abundantly expressed in normal tissues than in cancer cells, suggesting that these FKBPs may function in the opposite direction. Collectively, our study shows that FKBP52 and FKBP51 regulate ERα stability in a reciprocal manner and reveals a regulatory mechanism by which the expression of ERα is controlled.
Assuntos
Neoplasias da Mama , Receptor alfa de Estrogênio , Proteínas de Ligação a Tacrolimo , Neoplasias da Mama/metabolismo , Linhagem Celular Tumoral , Receptor alfa de Estrogênio/genética , Receptor alfa de Estrogênio/metabolismo , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Células MCF-7 , Estabilidade Proteica , Proteínas de Ligação a Tacrolimo/metabolismoRESUMO
The endogenous regenerative capacity of the brain is quite weak; however, a regenerative reaction, the production of new neurons (neurogenesis), has been reported to occur in brain lesions. In addition, leukocytes are well known to infiltrate brain lesions. Therefore, leukocytes would also have a link with regenerative neurogenesis; however, their role has not been fully elucidated. In this study, we investigated leukocyte infiltration and its influence on brain tissue regeneration in a trimethyltin (TMT)-injected mouse model of hippocampal regeneration. Immunohistochemically, CD3-positive T lymphocytes were found in the hippocampal lesion of TMT-injected mice. Prednisolone (PSL) treatment inhibited T lymphocyte infiltration and increased neuronal nuclei (NeuN)-positive mature neurons and doublecortin (DCX)-positive immature neurons in the hippocampus. Investigation of bromodeoxyuridine (BrdU)-labeled newborn cells revealed the percentage of BrdU/NeuN- and BrdU/DCX-positive cells increased by PSL treatment. These results indicate that infiltrated T lymphocytes prevent brain tissue regeneration by inhibiting hippocampal neurogenesis.
Assuntos
Células-Tronco Neurais , Linfócitos T , Compostos de Trimetilestanho , Camundongos , Animais , Bromodesoxiuridina , Hipocampo/patologia , Neurogênese/fisiologiaRESUMO
A 14-years-old squirrel monkey was euthanized due to weakness. Histopathological examination revealed multifocal growth of oval cells with severe atypia in the liver, spleen, and bone marrow. The neoplastic cells were positive for histiocytic markers (Iba1, HLA-DR, CD204). This is the fourth case of histiocytic sarcoma in non-human primates.
Assuntos
Sarcoma Histiocítico , Animais , Sarcoma Histiocítico/diagnóstico , Sarcoma Histiocítico/veterinária , Fígado , SaimiriRESUMO
Severe fever with thrombocytopenia syndrome (SFTS) is an emerging tickborne infectious disease caused by SFTS virus (SFTSV). We report 7 cases of spontaneous fatal SFTS in felines. Necropsies revealed characteristic lesions, including necrotizing lymphadenitis in 5 cases and necrotizing splenitis and SFTSV-positive blastic lymphocytes in all cases. We detected hemorrhagic lesions in the gastrointestinal tract in 6 cases and lungs in 3 cases, suggesting a more severe clinical course of SFTS in felids than in humans. We noted necrotic or ulcerative foci in the gastrointestinal tract in 3 cases, the lung in 2 cases, and the liver in 4 cases. We clarified that blastic lymphocytes are predominant targets of SFTSV and involved in induction of necrotic foci. We also found that thymic epithelial cells were additional targets of SFTSV. These results provide insights for diagnosing feline SFTS during pathological examination and demonstrate the similarity of feline and human SFTS cases.
Assuntos
Infecções por Bunyaviridae , Phlebovirus , Febre Grave com Síndrome de Trombocitopenia , Doenças Transmitidas por Carrapatos , Animais , Autopsia , Gatos , Humanos , JapãoRESUMO
Diabetes mellitus is a group of metabolic diseases characterized by hyperglycemia. Diabetic patients are known to have a higher prevalence and a higher risk of depression compared with the general population. The pathogenesis of diabetes-related depression is unclear, and the treatment is not well-established. Therefore, the prevention of diabetes-related depression is important for improving the quality of life of diabetic patients. Minocycline, a second-generation tetracycline antibiotic, has recently gained attention as a new agent for depression. In this study, we investigated the effect of minocycline on diabetes-related depression in a streptozotocin-induced mouse model of diabetes. Eight-week-old male C57BL/6 mice were injected with streptozotocin (200 mg/kg, i.p.). Seven days after injection, the mice received minocycline treatment through drinking water. We compared these mice with vehicle-treated control mice and diabetic mice not receiving minocycline treatment. On day 34, depression-like behavior was investigated using the forced swim test. On the following day, brain samples were collected, and formalin-fixed, paraffin-embedded specimens were prepared for immunohistochemistry. Compared with the control group, the diabetic mice not receiving minocycline treatment showed a prolonged duration of immobility in the forced swim test, the observation being interpreted as a depression-like behavior. Immunohistochemistry revealed an increase in microglia with an activated morphology in the diabetic mice without minocycline treatment. The expression of tumor necrosis factor alpha in microglia was increased. In addition, a decrease in the number of doublecortin-positive immature neurons was found in the hippocampus of diabetic mice. Minocycline treatment of diabetic animals prevented the depression-like behavior and microglial activation; however, minocycline did not reverse impaired hippocampal neurogenesis. These results indicate that minocycline has a preventive effect on diabetes-related depression. Inhibition of microglial activation would be a critical target for the antidepressant mechanism of minocycline. Impaired hippocampal neurogenesis was observed in diabetic mice; however, this may not be involved in the pathogenesis of depression.
Assuntos
Comportamento Animal/efeitos dos fármacos , Encéfalo/efeitos dos fármacos , Depressão/prevenção & controle , Minociclina/farmacologia , Animais , Antidepressivos/farmacologia , Encéfalo/metabolismo , Diabetes Mellitus Experimental/induzido quimicamente , Diabetes Mellitus Experimental/metabolismo , Diabetes Mellitus Experimental/patologia , Modelos Animais de Doenças , Hipocampo/efeitos dos fármacos , Hipocampo/patologia , Masculino , Camundongos Endogâmicos C57BL , Minociclina/metabolismo , Neurogênese/efeitos dos fármacos , Neurogênese/fisiologia , Qualidade de Vida , Estreptozocina/metabolismo , Estreptozocina/farmacologiaRESUMO
The brain has long been considered a site of "immune privilege"; however, recent evidence indicates the presence of brain-immune interactions in physiological and pathological conditions. Neurogenesis, a process of generating functionally integrated neurons, occurs in the adult brain of mammals. The adult neurogenesis predominantly takes place in the subgranular zone (SGZ) of the hippocampal dentate gyrus and the subventricular zone (SVZ). Several studies have shown that an immune reaction or alteration could affect adult neurogenesis activity, suggesting a link between the immune system and adult neurogenesis. Helminth infection is one of the activators of Th2 immune response. However, the influence of this type of immune reaction on adult neurogenesis is not well studied. In this study, we evaluated adult neurogenesis in mice infected with the helminth Nippostrongylus brasiliensis (Nb). Immunohistochemically, the number of both doublecortin-positive cells and doublecortin/5-bromodeoxyuridine (BrdU)-double-positive cells was decreased in the SGZ of Nb-infected mice by day 9 after infection. However, the total number of BrdU-positive newborn cells in the SGZ did not change. In no significant alterations were detected in the SVZ of infected mice. In addition, using reverse transcription-quantitative polymerase chain reaction, we observed no significant changes in the expression levels of neurotropic factors important for neurogenesis in the hippocampus. In conclusion, our results indicate that adult neurogenesis in SGZ, but not in SVZ, is inhibited by Nb infection. Th2 immune response might have a suppressive effect on hippocampal neurogenesis.
Assuntos
Hipocampo/citologia , Hipocampo/fisiologia , Imunidade Celular/fisiologia , Neurogênese/fisiologia , Nippostrongylus , Infecções por Strongylida/patologia , Animais , Proteína Duplacortina , Feminino , Camundongos , Distribuição Aleatória , Ratos , Ratos Sprague-Dawley , Infecções por Strongylida/imunologiaRESUMO
Epileptic seizure has been reported to enhance adult neurogenesis and induce aberrant synaptic reorganization in the human dentate gyrus in the hippocampal formation. However, adult neurogenesis in the extrahippocampal regions has not been well studied. To investigate seizure-enhanced neurogenesis in the extrahippocampal regions, we performed histological and immunohistochemical as well as western blot analyses on the cerebrum of Sprague-Dawley rats (n = 51, male, 7 weeks old, body weight 250-300 g) treated with intraperitoneal injection of kainic acid (KA, 10 mg/kg) to induce status epilepticus (SE) (n = 36) or normal saline solution (n = 15) followed by 5'-bromo-2-deoxyuridine (BrdU) injection to label newborn cells. Even though severe neuronal damage was found in the piriform cortex of rats having SE, immunohistochemistry for double cortin (DCX) revealed an increase in the number of immature neurons in the piriform cortex. Double immunofluorescence staining demonstrated that DCX-positive cells in the piriform cortex were positive for both BrdU and neuronal nuclear antigen. Immunohistochemistry and western blotting revealed increased expressions of synaptophysin and postsynaptic density protein 95 in the piriform cortex of rat having SE. These results suggested the enhanced neurogenesis and possible synaptic reorganization in the piriform cortex of the KA-treated rat.
Assuntos
Neurogênese , Plasticidade Neuronal , Córtex Piriforme/patologia , Estado Epiléptico/patologia , Animais , Proteína 4 Homóloga a Disks-Large/metabolismo , Proteína Duplacortina , Quinases Semelhantes a Duplacortina , Filamentos Intermediários/efeitos dos fármacos , Ácido Caínico , Masculino , Neurônios/citologia , Neurônios/patologia , Córtex Piriforme/citologia , Córtex Piriforme/fisiopatologia , Proteínas Serina-Treonina Quinases/metabolismo , Ratos , Ratos Sprague-Dawley , Estado Epiléptico/induzido quimicamente , Estado Epiléptico/metabolismo , Estado Epiléptico/fisiopatologia , Sinaptofisina/metabolismoRESUMO
Colorectal cancer is one of the most common causes of cancer death worldwide. In patients with metastatic colorectal cancer, combination treatment with several anti-cancer drugs is employed and improves overall survival in some patients. Nevertheless, most patients with metastatic disease are not cured owing to the drug resistance. Cancer stem cells are known to regulate resistance to chemotherapy. In the previous study, we established a novel three-dimensional organoid culture model from tumor colorectal tissues of human patients using an air-liquid interface (ALI) method, which contained numerous cancer stem cells and showed resistance to 5-fluorouracil (5-FU) and Irinotecan. Here, we investigate which inhibitor for stem cell-related signal improves the sensitivity for anti-cancer drug treatment in tumor ALI organoids. Treatment with Hedgehog signal inhibitors (AY9944, GANT61) decreases the cell viability of organoids compared with Notch (YO-01027, DAPT) and Wnt (WAV939, Wnt-C59) signal inhibitors. Combination treatment of AY9944 or GANT61 with 5-FU, Irinotecan or Oxaliplatin decreases the cell viability of tumor organoids compared with each anti-cancer drug alone treatment. Treatment with AY9944 or GANT61 inhibits expression of stem cell markers c-Myc, CD44 and Nanog, likely through the decrease of their transcription factor, GLI-1 expression. Combination treatment of AY9944 or GANT61 with 5-FU or Irinotecan also prevents colony formation of colorectal cancer cell lines HCT116 and SW480. These findings suggest that Hedgehog signals mediate anti-cancer drug resistance in colorectal tumor patient-derived ALI organoids and that the inhibitors are useful as a combinational therapeutic strategy against colorectal cancer.
Assuntos
Neoplasias Colorretais/metabolismo , Resistencia a Medicamentos Antineoplásicos , Proteínas Hedgehog/antagonistas & inibidores , Organoides/efeitos dos fármacos , Camptotecina/análogos & derivados , Camptotecina/farmacologia , Fluoruracila/farmacologia , Células HCT116 , Humanos , Receptores de Hialuronatos/genética , Receptores de Hialuronatos/metabolismo , Irinotecano , Células-Tronco Neoplásicas/efeitos dos fármacos , Compostos Organoplatínicos/farmacologia , Oxaliplatina , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Piridinas/farmacologia , Pirimidinas/farmacologia , Proteína GLI1 em Dedos de Zinco/genética , Proteína GLI1 em Dedos de Zinco/metabolismo , Dicloridrato de trans-1,4-Bis(2-clorobenzaminometil)ciclo-hexano/farmacologiaRESUMO
Dog spontaneously develop prostate cancer (PC) like humans. Because most dogs with PC have a poor prognosis, they could be used as a translational model for advanced PC in humans. Stem cell-derived 3-D organoid culture could recapitulate organ structures and physiology. Using patient tissues, a human PC organoid culture system was established. Recent study has shown that urine cells also possess the characteristic of stem cells. However, urine cell-derived PC organoids have never been produced. Therefore, we generated PC organoids using the dog urine samples. Urine organoids were successfully generated from each dog with PC. Each organoid showed cystic structures and resembled the epithelial structures of original tissues. Expression of an epithelial cell marker, E-cadherin, and a myofibloblast marker, α-SMA, was observed in the urine organoids. The organoids also expressed a basal cell marker, CK5, and a luminal cell marker, CK8. CD49f-sorted basal cell organoids rapidly grew compared with CD24-sorted luminal cell organoids. The population of CD44-positive cells was the highest in both organoids and the original urine cells. Tumors were successfully formed with the injection of the organoids into immunodeficient mice. Treatment with a microtubule inhibitor, docetaxel, but not a cyclooxygenase inhibitor, piroxicam, and an mTOR inhibitor, rapamycin, decreased the cell viability of organoids. Treatment with a Hedgehog signal inhibitor, GANT61, increased the radiosensitivity in the organoids. These findings revealed that PC organoids using urine might become a useful tool for investigating the mechanisms of the pathogenesis and treatment of PC in dogs.
Assuntos
Técnicas de Cultura de Células/métodos , Modelos Animais de Doenças , Células-Tronco Neoplásicas/patologia , Organoides , Neoplasias da Próstata , Urina/citologia , Animais , Cães , Xenoenxertos , Masculino , Camundongos , Camundongos Endogâmicos NOD , Camundongos SCIDRESUMO
Recurrent seizures without interictal resumption (status epilepticus) have been reported to induce neuronal death in the midline thalamic region that has functional roles in memory and decision-making; however, the pathogenesis underlying status epilepticus-induced thalamic neuronal death is yet to be determined. We performed histological and immunohistochemical studies as well as cerebral blood flow measurement using 4.7 tesla magnetic resonance imaging spectrometer on midline thalamic region in Sprague-Dawley rats (n = 75, male, 7 weeks after birth, body weight 250-300 g) treated with intraperitoneal injection of kainic acid (10 mg/kg) to induce status epilepticus (n = 55) or normal saline solution (n = 20). Histological study using paraffin-embedded specimens revealed neuronal death showing ischemic-like changes and Fluoro-Jade C positivity with calcium deposition in the midline thalamic region of epileptic rats. The distribution of neuronal death was associated with focal loss of immunoreactivity for excitatory amino acid transporter 2 (EAAT2), stronger immunoreaction for glutamate and increase in number of Iba-1-positive microglial cells showing swollen cytoplasm and long processes. Double immunofluorescence study demonstrated co-expression of interleukin-1 beta (IL-1ß) and inducible nitric oxide synthase (iNOS) within microglial cells, and loss of EAAT2 immunoreactivity in reactive astrocytes. These microglial alterations and astrocytic EAAT2 downregulation were also observed in tissue without obvious neuronal death in kainic acid-treated rats. These results suggest the possible role of glutamate excitotoxicity in neuronal death in the midline thalamic region following kainic acid-induced status epilepticus due to astrocytic EAAT2 downregulation following microglial activation showing upregulation of IL-1ß and iNOS.
Assuntos
Transportador 2 de Aminoácido Excitatório/metabolismo , Núcleos da Linha Média do Tálamo/metabolismo , Núcleos da Linha Média do Tálamo/patologia , Neurônios/fisiologia , Estado Epiléptico/metabolismo , Estado Epiléptico/patologia , Animais , Astrócitos/metabolismo , Morte Celular , Regulação para Baixo , Ácido Caínico , Espectroscopia de Ressonância Magnética , Masculino , Microglia/metabolismo , Núcleos da Linha Média do Tálamo/irrigação sanguínea , Neurônios/citologia , Ratos , Ratos Sprague-Dawley , Estado Epiléptico/induzido quimicamenteRESUMO
This study aimed to investigate the effects of rolipram, a phosphodiesterase inhibitor, on brain tissue regeneration. Trimethyltin-injected mice, an animal model of hippocampal tissue regeneration, was created by a single injection of trimethyltin chloride (2.2â mg/kg, intraperitoneally). Daily rolipram administration (10â mg/kg, intraperitoneally) was performed from the day after trimethyltin injection until the day before sampling. In Experiment 1, brain samples were collected on day 7 postinjection of trimethyltin following the forced swim test. In Experiment 2, bromodeoxyuridine (150â mg/kg, intraperitoneally/day) was administered on days 3-5 and sampling was on day 21 postinjection of trimethyltin. Samples were routinely embedded in paraffin and sections were obtained for histopathological investigation. In Experiment 1, rolipram-treated mice showed shortened immobility times in the forced swim test. Histopathology revealed that rolipram treatment had improved the replenishment of neuronal nuclei-positive neurons in the dentate gyrus, which was accompanied by an increase in the percentage of phosphorylated cyclic AMP response element-binding protein-positive cells. In addition, rolipram had decreased the percentage of ionized calcium-binding adapter protein 1-positive microglia with activated morphology and the number of tumor necrosis factor-alpha-expressing cells. In Experiment 2, double immunofluorescence for bromodeoxyuridine/neuronal nuclei revealed an increase of double-positive cells in rolipram-treated mice. These results demonstrate that rolipram effectively promotes brain tissue regeneration by enhancing the survival of newborn neurons and inhibiting neuroinflammation.
Assuntos
Hipocampo , Rolipram , Compostos de Trimetilestanho , Animais , Compostos de Trimetilestanho/toxicidade , Rolipram/farmacologia , Hipocampo/efeitos dos fármacos , Hipocampo/patologia , Hipocampo/metabolismo , Camundongos , Masculino , Doenças Neurodegenerativas/induzido quimicamente , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/patologia , Regeneração Nervosa/efeitos dos fármacos , Regeneração Nervosa/fisiologia , Neurônios/efeitos dos fármacos , Neurônios/patologia , Microglia/efeitos dos fármacos , Microglia/patologiaRESUMO
Skin is the largest organ in a mammal body, and it exhibits most significant range of adaptations to different habitats. It is a complex, biological composite structure, consisting of epidermis, dermis and subcutaneous tissues and is used for the therapeutic application of medical devices to improve healthcare. Extensive studies have been performed on the roles of the skin; however, little is known on its physiological characteristics in relation to body size among different species. The purpose of this study was therefore to evaluate the allometric scaling of skin weight (SW) and thickness (ST) to body weight (BW) in relation to genetics and habitats. Also analysed the relationship of BW to thicknesses of epidermis, dermis and subcutaneous tissues. This study used 249 adult animals of both sexes, belonging to 144 species, clustered in 18 taxonomic orders and five types of habitats. The animals were obtained from various sources in Japan. SW and BW were weighed, and ST was measured using a calliper followed by data analysis. Results showed that SW and ST were related to BW [log SW = 0.969 × logBW - 0636, adjust. R2 : 0.975]. The BW increased with increasing skin dermal thickness (y = 0.3916x + 1.5253, adjust. R2 : 0.6921), slightly with epidermal thickness (y = 0.2495x + 0.3984, adjust. R2 : 0.3402), but not all with the thickness of subcutaneous tissues (y = 0.1454x + 2.2437, adjust. R2 : 0.0752). The ratio of SW to BW (SW/BW) distributed over a large range from 0.06 to 0.64 values and varied among animal taxonomic orders and their dwelling habitats. Close relationship of BW to SW/BW was observed in species weighing ≥200 g but not in species weighing <200 g. In conclusion, SW and ST in mammals are determined by BW. The SW/BW varies based on BW, taxonomic orders and habitat and is large in small mammals weighing ≥200 g to provide a mechanism used for survival strategy.
Assuntos
Mamíferos , Pele , Masculino , Feminino , Animais , Mamíferos/fisiologia , Tamanho Corporal , Peso Corporal/fisiologia , EcossistemaRESUMO
BACKGROUND/AIM: The activation of phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway has been implicated in canine soft tissue sarcoma (STS) and may serve as a prognostic marker. This study investigated the correlation between PI3K/Akt activation in tumor cells and tumor-infiltrating lymphocytes (TILs). MATERIALS AND METHODS: A total of 59 STS samples were labeled via immunohistochemistry to calculate the density of TILs, including CD3+ T cells, CD8+ T cells, CD20+ B cells, and FOXP3+ regulatory T cells. RESULTS: Forty-eight samples (81.3%) had intra-tumoral TILs with a high density of CD3+ T cells (mean: 283.3 cells/mm2) and CD8+ T cells (mean: 134.8 cells/mm2). Conversely, CD20+ B cells (mean: 73.6 cells/mm2) and FOXP3+ regulatory T cells (mean: 9.2 cells/mm2) were scarce. The abundance of CD3+/CD8+, CD3+/CD20+, and CD8+/CD20+ TILs were highly correlated in multivariate analyses (r=0.895, 0.946, and 0.856, respectively). Nonetheless, TIL density was unrelated to clinicopathological parameters (sex, age, tumor location, breed) and tumor grade. The abundance of CD8+ T cells was positively correlated with the activation of PI3K/Akt, indicating that samples with high levels of phospho-Akt and phospho-S6 tend to have a higher CD8+ T cell density (p=0.0032 and 0.0218, respectively). Furthermore, TIL density was correlated with the Ki-67 index, a tumor proliferation and growth marker. Samples with a high Ki-67 index had a significantly higher abundance of CD3+ T cells, CD8+ T cells, and CD20+ B cells (p=0.0392, 0.0254, 0.0380, respectively). CONCLUSION: PI3K/Akt pathway activation may influence the infiltration of CD8+ T cells within the tumor microenvironment in canine STS. Prospective studies involving a higher number of cases are warranted to confirm these findings.
Assuntos
Linfócitos T CD8-Positivos , Antígeno Ki-67 , Linfócitos do Interstício Tumoral , Proteínas Proto-Oncogênicas c-akt , Sarcoma , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Animais , Proteínas Proto-Oncogênicas c-akt/metabolismo , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Sarcoma/veterinária , Sarcoma/patologia , Sarcoma/imunologia , Sarcoma/metabolismo , Antígeno Ki-67/metabolismo , Cães , Feminino , Masculino , Imuno-Histoquímica , Transdução de Sinais , Doenças do Cão/imunologia , Doenças do Cão/patologia , Doenças do Cão/metabolismoRESUMO
Canine tumours including urothelial carcinoma, lung adenocarcinoma, mammary gland tumour, squamous cell carcinoma, and melanoma have been identified as causes of death, but effective therapies are limited due to insufficient knowledge of the molecular mechanisms involved. Within the tumour microenvironment, hypoxia activates hypoxia-inducible factor 1α (HIF1α) in tumour cells. High HIF1α expression correlates with enhanced glycolysis and poorer outcomes in human cancers. However, the molecular mechanisms underlying hypoxic tumour cells remain elusive in dogs. In our study, we investigated upregulated genes in a canine malignant melanoma cell line during hypoxia using RNA-sequencing analysis. Glycolysis and HIF1 signalling pathways were upregulated in hypoxic melanoma cells. HIF1α knockout melanoma cells revealed that the glycolysis marker MCT4 is regulated by HIF1α activation. Hypoxia induces high lactate secretion due to enhanced glycolysis in canine melanoma cells. Furthermore, we examined monocarboxylate transporter 4 (MCT4) expression in malignant melanoma and eight other types of canine tumour tissues using immunohistochemistry (IHC). Membrane-localized MCT4 protein was mostly detected in urothelial carcinoma and lung adenocarcinoma rather than malignant melanoma. We conclude that canine MCT4 protein plays a role in lactic acid efflux from glycolytic cells and may serve as a marker for hypoxia and glycolysis in canine tumours. These findings could inform future therapeutic strategies targeting MCT4.
Assuntos
Doenças do Cão , Animais , Cães , Doenças do Cão/genética , Doenças do Cão/metabolismo , Linhagem Celular Tumoral , Neoplasias/veterinária , Neoplasias/metabolismo , Neoplasias/genética , Regulação Neoplásica da Expressão Gênica , Melanoma/veterinária , Melanoma/genética , Melanoma/metabolismo , Glicólise/genética , Transportadores de Ácidos Monocarboxílicos/genética , Transportadores de Ácidos Monocarboxílicos/metabolismo , Hipóxia/veterinária , Hipóxia/metabolismo , Hipóxia/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genéticaRESUMO
Soft tissue sarcoma (STS) is a relatively common tumor in dogs. However, very few canine STS cell lines are available. This study aimed to establish a new cell line, STS-YU1, derived from a recurrence of myxosarcoma in an 11-year-old mixed-breed dog. We examined STS-YU1 for in vitro cell proliferation, migration, anticancer drug sensitivity, transcriptome analysis using next-generation sequencing (RNA-seq), and in vivo tumorigenicity in mice and compared it with previously established STS cell lines, MUMA-G and A72. The cell proliferation and migration of STS-YU1 were higher than MUMA-G although MUMA-G only exhibited tumorigenicity in mice. STS-YU1 showed dose-dependent cytotoxicity to anticancer drugs, but with weak effects. RNA-seq analysis revealed the molecular phenotype of STS-YU1 was different from that of a previously reported cell line, A72. Hence, the use of STS-YU1 would help in efficient drug screening against canine STS in vitro.
Assuntos
Antineoplásicos , Doenças do Cão , Doenças dos Roedores , Sarcoma , Animais , Cães , Camundongos , Sarcoma/veterinária , Linhagem Celular , Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Proliferação de Células , Doenças do Cão/diagnósticoRESUMO
BACKGROUND: The anti-programmed death 1 (PD-1) antibody has led to durable clinical responses in a wide variety of human tumors. We have previously developed the caninized anti-canine PD-1 antibody (ca-4F12-E6) and evaluated its therapeutic properties in dogs with advance-staged oral malignant melanoma (OMM), however, their therapeutic effects on other types of canine tumors remain unclear. OBJECTIVE: The present clinical study was carried out to evaluate the safety profile and clinical efficacy of ca-4F12-E6 in dogs with advanced solid tumors except for OMM. METHODS: Thirty-eight dogs with non-OMM solid tumors were enrolled prospectively and treated with ca-4F12-E6 at 3 mg/kg every 2 weeks of each 10-week treatment cycle. Adverse events (AEs) and treatment efficacy were graded based on the criteria established by the Veterinary Cooperative Oncology Group. RESULTS: One dog was withdrawn, and thirty-seven dogs were evaluated for the safety and efficacy of ca-4F12-E6. Treatment-related AEs of any grade occurred in 13 out of 37 cases (35.1%). Two dogs with sterile nodular panniculitis and one with myasthenia gravis and hypothyroidism were suspected of immune-related AEs. In 30 out of 37 dogs that had target tumor lesions, the overall response and clinical benefit rates were 6.9% and 27.6%, respectively. The median progression-free survival and overall survival time were 70 days and 215 days, respectively. CONCLUSIONS: The present study demonstrated that ca-4F12-E6 was well-tolerated in non-OMM dogs, with a small number of cases showing objective responses. This provides evidence supporting large-scale clinical trials of anti-PD-1 antibody therapy in dogs.
Assuntos
Doenças do Cão , Melanoma , Neoplasias Cutâneas , Cães , Animais , Humanos , Melanoma/tratamento farmacológico , Melanoma/veterinária , Melanoma/patologia , Receptor de Morte Celular Programada 1 , Neoplasias Cutâneas/tratamento farmacológico , Neoplasias Cutâneas/veterinária , Neoplasias Cutâneas/patologia , Resultado do Tratamento , Doenças do Cão/tratamento farmacológicoRESUMO
Immunotherapy is a breakthrough in human cancer therapy and has become a major concern in veterinary oncology. However, in cats, many unclear points of the tumor microenvironment exist, including immune checkpoint molecules. A reason is that very few monoclonal antibodies have been proven to react with feline molecules. Therefore, this study investigated whether anti-human programmed cell death ligand 1 (PD-L1) monoclonal antibody, clone 28-8, which is currently commercially available, can also recognize feline PD-L1 by flow cytometry, immunoprecipitation, and immunohistochemical (IHC) staining. We confirmed that the antibody's specificity by flow cytometry and immunoprecipitation using NIH3T3 cells transfected with feline PD-L1. Additionally, we revealed that PD-L1 was expressed on the surface of some feline cell lines by flow cytometry and clone 28-8 antibody unbound to the cells where feline PD-L1 was knocked out. Furthermore, IHC analysis revealed that PD-L1 was expressed in macrophages in the spleen and lymph nodes from healthy cats and mast cell tumor cells. Therefore, we indicated that the clone 28-8 antibody is a valuable tool in detecting feline PD-L1, and further analysis of tumor tissues is expected in the future.
Assuntos
Anticorpos Monoclonais , Antígeno B7-H1 , Camundongos , Gatos , Animais , Imuno-Histoquímica , Ligantes , Células NIH 3T3 , Células Clonais/química , Células Clonais/metabolismo , ApoptoseRESUMO
The abnormal activity of PP2A, a dominant member of type 2A serine/threonine protein phosphatase, has been implicated in the development of Alzheimer's disease (AD) and dementia with Lewy bodies (DLB). PP2A is a holoenzyme, and protein methylation of the catalytic subunit, PP2Ac, alters the complex composition. A decrease in PP2Ac methylation levels has been reported in AD and DLB. Aging is the most common risk factor for AD and DLB, but the relationship between aging and PP2A has not been studied in detail. Cynomolgus monkey show increased phosphorylation levels of tau and α-synuclein with aging. In this study, we investigated the alterations in the PP2A activity regulation with aging in monkey brains from 2 to 43 years of age using fractionated proteins. We found that type 2A protein phosphatase activity decreased with aging in cytoplasmic and nuclear-soluble fractions. PP2Ac methylation level was decreased in cytoplasmic and sarkosyl-insoluble fractions. A principal component analysis using PP2Ac, demethylated PP2Ac and PP2A methylesterase PME-1 levels in cytoplasmic and nuclear-soluble fractions as attributes showed that aged monkeys were in the same cluster. Our results show that brain aging in cynomolgus monkeys is closely related to changes in PP2A methylation.
Assuntos
Doença de Alzheimer , Proteína Fosfatase 2 , Animais , Proteína Fosfatase 2/metabolismo , Macaca fascicularis/metabolismo , Projetos Piloto , Metilação , Doença de Alzheimer/metabolismo , Fosforilação , Encéfalo/metabolismoRESUMO
Canine soft tissue sarcoma (STS) is relatively common in dogs and is the generic term for tumours that originate from mesenchymal cells. While histopathological grade and immunolabelling with Ki-67 have been used for estimating prognosis, additional indicators are needed for predicting prognosis. Aberrant cell signalling pathways may contribute to disease activity and, therefore, prognostic markers. However, their role in canine STS remains poorly understood. The aim of this study was to investigate expression of phosphorylated Akt (phospho-Akt) and phosphorylated S6 (phospho-S6) as potential prognostic indicators. Immunohistochemical labelling was conducted on clinical samples of canine STS (n = 67). We found that phospho-Akt expression was positively correlated with histopathological grade (P = 0.001) and Ki-67 index (P <0.01). There was no apparent relationship between the type of STS and the expression of phospho-Akt. The number of cases that expressed phospho-S6, which is the downstream molecule of the Akt signalling pathway, was higher in immunopositive phospho-Akt cases than in immunonegative phospho-Akt cases (P <0.0001). Furthermore, phospho-Akt expression was significantly higher in recurrent and metastatic cases. We also confirmed that phosphorylation of Akt occurred in conjunction with S6 phosphorylation in three canine STS cell lines. These results suggest that immunolabelling for phospho-Akt, phospho-S6 and Ki-67 could potentially be used as a prognostic indicator and therapeutic target in canine STS.