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1.
J Vet Sci ; 25(3): e48, 2024 May.
Artigo em Inglês | MEDLINE | ID: mdl-38834516

RESUMO

IMPORTANCE: Early diagnosis of canine pancreatitis is challenging due to non-specific clinical signs. Currently, abdominal ultrasonography and measurement of canine pancreatic lipase (cPL) have been employed for the diagnosis of pancreatitis. OBJECTIVE: Many qualitative and quantitative commercial cPL tests have been developed and used in veterinary clinics. This study aimed to compare three different methodologies SNAP cPL, Spec cPL, and Vcheck cPL tests to assess the concordance of these assays. METHODS: Fifty serum samples were collected from 36 dogs with or without pancreatitis and subjected to SNAP cPL, Spec cPL, and Vcheck cPL tests. Agreement and correlation coefficients were calculated between the test results, and correlations were determined during the management of the patients. RESULTS: The results of the three cPL assays were strongly correlated in 47/50 serum samples (94%). Cohen's kappa analysis between the Spec cPL and Vcheck cPL showed near perfect agreement (κ = 0.960, p < 0.001), SNAP cPL and Vcheck cPL (κ = 0.920, p < 0.001), and Spec cPL and SNAP cPL (κ = 0.880, p < 0.001). The correlation coefficients (r) between data from Spec cPL and Vcheck cPL tests was calculated by Spearman's correlation test (r = 0.958, p < 0.001). Furthermore, the patterns of change in serum cPL concentrations determined using Spec cPL and Vcheck cPL were significantly consistent during the monitoring period in 11 patients. CONCLUSIONS AND RELEVANCE: Our data illustrated that Spec cPL and Vcheck cPL tests are compatible for clinical use in the diagnosis and monitoring of canine pancreatitis.


Assuntos
Doenças do Cão , Lipase , Pancreatite , Animais , Cães , Lipase/sangue , Pancreatite/veterinária , Pancreatite/diagnóstico , Pancreatite/sangue , Doenças do Cão/diagnóstico , Doenças do Cão/sangue , Masculino , Feminino , Pâncreas/enzimologia
2.
Biomed Pharmacother ; 176: 116853, 2024 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-38850663

RESUMO

Various adjuvants have been tested clinically for patients with problems with embryo implantation during in vitro fertilization (IVF)-embryo transfer (ET). Vitamin D3, an essential modulator of various physiological processes, has received attention as an important adjuvant for successful pregnancy, as many studies have shown a strong association between vitamin D deficiency and implantation failure and fetal growth restriction. However, vitamin D has been widely utilized in different protocols, resulting in non-reproducible and debatable outcomes. In the present study, we demonstrated that cyclic intrauterine administration of vitamin D3 increased endometrial receptivity and angiogenesis, which could be attributed to increased recruitment of uterus-resident natural killer cells. In particular, cyclic treatment of vitamin D3 promoted stable attachment of the embryo onto endometrial cells in vitro, suggesting its merit during the early stage of embryo implantation to support the initial maternal-fetal interactions. Our findings suggest that women with repeated implantation failure may benefit from the use of vitamin D3 as a risk-free adjuvant prior to IVF-ET procedures to improve the uterine environment, and make it favorable for embryo implantation.


Assuntos
Colecalciferol , Implantação do Embrião , Implantação do Embrião/efeitos dos fármacos , Feminino , Colecalciferol/farmacologia , Colecalciferol/administração & dosagem , Gravidez , Humanos , Animais , Endométrio/efeitos dos fármacos , Fertilização in vitro/métodos , Transferência Embrionária , Células Matadoras Naturais/efeitos dos fármacos , Neovascularização Fisiológica/efeitos dos fármacos , Útero/efeitos dos fármacos
3.
Theranostics ; 14(3): 954-972, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-38250040

RESUMO

Background: Asherman's syndrome (AS) is a dreadful gynecological disorder of the uterus characterized by intrauterine adhesion with severe fibrotic lesions, resulting in a damaged basalis layer with infertility. Despite extensive research on overcoming AS, evidence-based effective and reproducible treatments to improve the structural and functional morphology of the AS endometrium have not been established. Methods: Endometrial organoids generated from human or mouse endometrial tissues were transplanted into the uterine cavity of a murine model of AS to evaluate their transplantable feasibility to improve the AS uterine environment. The successful engraftment of organoid was confirmed by detection of human mitochondria and cytosol (for human endometrial organoid) or enhanced green fluorescent protein signals (for mouse endometrial organoid) in the recipient endometrium. The therapeutic effects mediated by organoid transplantation were examined by the measurements of fibrotic lesions, endometrial receptivity and angiogenesis, and fertility assessment by recording the number of implantation sites and weighing the fetuses and placenta. To explore the cellular and molecular mechanisms underlying the recovery of AS endometrium, we evaluated the status of mitochondrial movement and biogenetics in organoid transplanted endometrium. Results: Successfully engrafted endometrial organoids with similar morphological and molecular features to the parental tissues dramatically repaired the AS-induced damaged endometrium, significantly reducing fibrotic lesions and increasing fertility outcomes in mice. Moreover, dysfunctional mitochondria in damaged tissues, which we propose might be a key cellular feature of the AS endometrium, was fully recovered by functional mitochondria transferred from engrafted endometrial organoids. Endometrial organoid-originating mitochondria restored excessive collagen accumulation in fibrotic lesions and shifted uterine metabolic environment to levels observed in the normal endometrium. Conclusions: Our findings suggest that endometrial organoid-originating mitochondria might be key players to mediate uterine repair resulting in fertility enhancement by recovering abrogated metabolic circumstance of the endometrium with AS. Further studies addressing the clinical applicability of endometrial organoids may aid in identifying new therapeutic strategies for infertility in patients with AS.


Assuntos
Infertilidade , Útero , Feminino , Gravidez , Humanos , Animais , Camundongos , Endométrio , Mitocôndrias , Organoides
4.
Biomol Ther (Seoul) ; 31(6): 682-691, 2023 Nov 01.
Artigo em Inglês | MEDLINE | ID: mdl-37899745

RESUMO

Cell transformation induced by epidermal growth factor (EGF) and 12-O-tetradecanoylphorbol-13-acetate (TPA) is a critical event in cancer initiation and progression, and understanding the underlying mechanisms is essential for the development of new therapeutic strategies. Licorice extract contains various bioactive compounds, which have been reported to have anticancer and anti-inflammatory effects. This study investigated the cancer preventive efficacy of licochalcone D (LicoD), a chalcone derivative in licorice extract, in EGF and TPA-induced transformed skin keratinocyte cells. LicoD effectively suppressed EGF-induced cell proliferation and anchorage-independent colony growth. EGF and TPA promoted the S phase of cell cycle, while LicoD treatment caused G1 phase arrest and down-regulated cyclin D1 and up-regulated p21 expression associated with the G1 phase. LicoD also induced apoptosis and increased apoptosis-related proteins such as cleaved-caspase-3, cleaved-caspase-7, and Bax (Bcl-2-associated X protein). We further investigated the effect of LicoD on the AKT signaling pathway involved in various cellular processes and found decreased p-AKT, p-GSK3ß, and p-NFκB expression. Treatment with MK-2206, an AKT pharmacological inhibitor, suppressed EGF-induced cell proliferation and transformed colony growth. In conclusion, this study demonstrated the potential of LicoD as a preventive agent for skin carcinogenesis.

5.
Plants (Basel) ; 12(8)2023 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-37111949

RESUMO

In total, four new eudesmane-type sesquiterpene glycosides, askoseosides A-D (1-4), and 18 known compounds (5-22) were isolated from the flowers of Aster koraiensis via chromatographic techniques. Chemical structures of the isolated compounds were identified by spectroscopic/spectrometric methods, including NMR and HRESIMS, and the absolute configuration of the new compounds (1 and 2) was performed by electronic circular dichroism (ECD) studies. Further, the anticancer activities of the isolated compounds (1-22) were evaluated using the epidermal growth factor (EGF)-induced as well as the 12-O-tetradecanoylphorbol 13-acetate (TPA)-induced cell transformation assay. Among the 22 compounds, compounds 4, 9, 11, 13-15, 17, 18, and 22 significantly inhibited both EGF- and TPA-induced colony growth. In particular, askoseoside D (4, EGF: 57.8%; TPA: 67.1%), apigenin (9, EGF: 88.6%; TPA: 80.2%), apigenin-7-O-ß-d-glucuronopyranoside (14, EGF: 79.2%; TPA: 70.7%), and 1-(3',4'-dihydroxycinnamoyl) cyclopentane-2,3-diol (22, EGF: 60.0%; TPA: 72.1%) showed higher potent activities.

6.
Expert Rev Mol Med ; 24: e43, 2022 10 25.
Artigo em Inglês | MEDLINE | ID: mdl-36281483

RESUMO

Globally, an aging population is increasing, and aging is a natural physiological process and a major risk factor for all age-related diseases. It seriously threatens personal health and imposes a great economic burden. Therefore, there is a growing scientific interest in strategies for well-aging with prevention and treatment of age-related diseases. The seed, root, stem or leaves of Cassia tora Linn. are useful for anti-bacteria, anti-hyperlipidemia and anti-obesity due to its pharmacological activities such as anti-inflammation and anti-oxidant both in vitro and in vivo. Nevertheless, no clinical trials have been attempted so far, therefore here we would like to understand the current preclinical activities for aging-related disease models including cataract, metabolic dysfunction and neurodegeneration, then discuss their preparation for clinical trials and perspectives.


Assuntos
Cassia , Catarata , Humanos , Idoso , Cassia/metabolismo , Extratos Vegetais/farmacologia , Extratos Vegetais/uso terapêutico , Antioxidantes/metabolismo , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Catarata/tratamento farmacológico , Catarata/metabolismo , Envelhecimento
7.
Int J Mol Sci ; 23(3)2022 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-35163115

RESUMO

Honeybee venom has recently been considered an anti-neurodegenerative agent, primarily due to its anti-inflammatory effects. The natural accumulation of amyloid-beta (Aß) in the brain is reported to be the natural cause of aging neural ability downfall, and oxidative stress is the main route by which Aß ignites its neural toxicity. Anti-neural oxidative stress is considered an effective approach for neurodegenerative therapy. To date, it is unclear how bee venom ameliorates neuronal cells in oxidative stress induced by Aß. Here, we evaluated the neuroprotective effect of bee venom on Aß-induced neural oxidative stress in both HT22 cells and an animal model. Our results indicate that bee venom protected HT22 cells against apoptosis induced by Aß1-42. This protective effect was explained by the increased nuclear translocation of nuclear factor erythroid 2-like 2 (Nrf2), consequently upregulating the production of heme oxygenase-1 (HO-1), a critical cellular instinct antioxidant enzyme that neutralizes excessive oxidative stress. Furthermore, bee venom treatment activated the tropomyosin-related kinase receptor B (TrkB)/cAMP response element-binding (CREB)/brain-derived neurotrophic factor (BDNF), which is closely related to the promotion of cellular antioxidant defense and neuronal functions. A mouse model with cognitive deficits induced by Aß1-42 intracerebroventricular (ICV) injections was also used. Bee venom enhanced animal cognitive ability and enhanced neural cell genesis in the hippocampal dentate gyrus region in a dose-dependent manner. Further analysis of animal brain tissue and serum confirmed that bee venom reduced oxidative stress, cholinergic system activity, and intercellular neurotrophic factor regulation, which were all adversely affected by Aß1-42. Our study demonstrates that bee venom exerts antioxidant and neuroprotective actions against neural oxidative stress caused by Aß1-42, thereby promoting its use as a therapeutic agent for neurodegenerative disorders.


Assuntos
Peptídeos beta-Amiloides/toxicidade , Venenos de Abelha/farmacologia , Disfunção Cognitiva/tratamento farmacológico , Regulação da Expressão Gênica/efeitos dos fármacos , Doenças Neurodegenerativas/tratamento farmacológico , Neurônios/efeitos dos fármacos , Estresse Oxidativo , Fragmentos de Peptídeos/toxicidade , Animais , Apoptose , Fator Neurotrófico Derivado do Encéfalo/genética , Fator Neurotrófico Derivado do Encéfalo/metabolismo , Disfunção Cognitiva/induzido quimicamente , Disfunção Cognitiva/metabolismo , Disfunção Cognitiva/patologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Heme Oxigenase-1/genética , Heme Oxigenase-1/metabolismo , Masculino , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos ICR , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Doenças Neurodegenerativas/induzido quimicamente , Doenças Neurodegenerativas/metabolismo , Doenças Neurodegenerativas/patologia , Neurônios/metabolismo , Neurônios/patologia , Fármacos Neuroprotetores/farmacologia , Receptor trkB/genética , Receptor trkB/metabolismo
8.
World J Mens Health ; 40(2): 316-329, 2022 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-35021315

RESUMO

PURPOSE: To build an age prediction model, we measured CD4+ and CD8+ cells, and humoral components in canine peripheral blood. MATERIALS AND METHODS: Large Belgian Malinois (BGM) and German Shepherd Dog (GSD) breeds (n=27), aged from 1 to 12 years, were used for this study. Peripheral bloods were obtained by venepuncture, then plasma and peripheral blood mononuclear cells (PBMCs) were separated immediately. Six myokines, including interleukin (IL)-6, IL-8, IL-15, leukemia inhibitory factor (LIF), growth differentiation factor 8 (GDF8), and GDF11 were measured from plasma and CD4+/CD8+ T-lymphocytes ratio were measured from PBMC. These parameters were then tested with age prediction models to find the best fit model. RESULTS: We found that the T-lymphocyte ratio (CD4+/CD8+) was significantly correlated with age (r=0.46, p=0.016). Among the six myokines, only GDF8 showed a significant correlation with age (r=0.52, p=0.005). Interestingly, these two markers showed better correlations in male dogs than females, and BGM breed than GSD. Using these two age biomarkers, we could obtain the best fit in a quadratic linear mixed model (r=0.77, p=3×10-6). CONCLUSIONS: Age prediction is a challenging task because of complication with biological age. Our quadratic linear mixed model using CD4+/CD8+ ratio and GDF8 level showed a meaningful age prediction.

9.
Int J Mol Sci ; 21(18)2020 Sep 18.
Artigo em Inglês | MEDLINE | ID: mdl-32962182

RESUMO

Global environmental pollution has led to human exposure to ultraviolet (UV) radiation due to the damaged ozone layer, thereby increasing the incidence and death rate of skin cancer including both melanoma and non-melanoma. Overexpression and activation of V-akt murine thymoma viral oncogene homolog (AKT, also known as protein kinase B) and related signaling pathways are major factors contributing to many cancers including lung cancer, esophageal squamous cell carcinoma and skin cancer. Although BRAF inhibitors are used to treat melanoma, further options are needed due to treatment resistance and poor efficacy. Depletion of AKT expression and activation, and related signaling cascades by its inhibitors, decreases the growth of skin cancer and metastasis. Here we have focused the effects of AKT and related signaling (PI3K/AKT/mTOR) pathways by regulators derived from plants and suggest the need for efficient treatment in skin cancer therapy.


Assuntos
Antineoplásicos/farmacologia , Melanoma/tratamento farmacológico , Extratos Vegetais/farmacologia , Plantas Medicinais/química , Inibidores de Proteínas Quinases/farmacologia , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Neoplasias Cutâneas/tratamento farmacológico , Animais , Antineoplásicos/química , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Humanos , Melanoma/metabolismo , Camundongos , Extratos Vegetais/química , Extratos Vegetais/uso terapêutico , Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas c-akt/química , Proteínas Proto-Oncogênicas c-akt/genética , Transdução de Sinais/efeitos dos fármacos , Transdução de Sinais/genética , Neoplasias Cutâneas/metabolismo , Neoplasias Cutâneas/patologia
10.
PLoS Genet ; 16(3): e1008617, 2020 03.
Artigo em Inglês | MEDLINE | ID: mdl-32130226

RESUMO

The oligosaccharyl transferase (OST) protein complex mediates the N-linked glycosylation of substrate proteins in the endoplasmic reticulum (ER), which regulates stability, activity, and localization of its substrates. Although many OST substrate proteins have been identified, the physiological role of the OST complex remains incompletely understood. Here we show that the OST complex in C. elegans is crucial for ER protein homeostasis and defense against infection with pathogenic bacteria Pseudomonas aeruginosa (PA14), via immune-regulatory PMK-1/p38 MAP kinase. We found that genetic inhibition of the OST complex impaired protein processing in the ER, which in turn up-regulated ER unfolded protein response (UPRER). We identified vitellogenin VIT-6 as an OST-dependent glycosylated protein, critical for maintaining survival on PA14. We also showed that the OST complex was required for up-regulation of PMK-1 signaling upon infection with PA14. Our study demonstrates that an evolutionarily conserved OST complex, crucial for ER homeostasis, regulates host defense mechanisms against pathogenic bacteria.


Assuntos
Proteínas de Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/metabolismo , Retículo Endoplasmático/metabolismo , Proteostase/fisiologia , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Hexosiltransferases/metabolismo , Imunidade Inata/fisiologia , Sistema de Sinalização das MAP Quinases/fisiologia , Proteínas de Membrana/metabolismo , Pseudomonas aeruginosa/metabolismo , Transdução de Sinais/fisiologia , Fatores de Transcrição/metabolismo , Resposta a Proteínas não Dobradas/fisiologia , Regulação para Cima/fisiologia , Vitelogeninas/metabolismo
11.
World J Surg ; 43(11): 2699-2709, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31399794

RESUMO

BACKGROUND: To compare the effectiveness of a custom-made skills simulator (CMSS) with the commercially available da Vinci® skills simulator (DVSS) that help improving surgical skills for effective and safe robotic surgical interventions. METHODS: A randomized control study was conducted to determine the performance of participants after undergoing robotic surgical training. Total 64 students who had no previous experience with robotic surgery enrolled this study. After 5 min-introduction of robotic surgical system, the participants got random-assignment into two groups to perform either CMSS-or DVSS-exercises. After 15 min-practicing the corresponding simulator, task-execution performance and individual questionnaires were compared between participants trained with the CMSS and those trained with the DVSS. RESULTS: Regardless of simulator the participants used, the system understanding and manipulation ability of the participants was found to be higher than after completing the simulation-based robotic surgical training (p < 0.05). However, there were no significant differences in terms of the required time to complete the tasks, and improvement of understanding the concept of robotic surgery, or surgical skill capacity between two groups (p > 0.05). CONCLUSIONS: The training effectiveness of CMSS was not significantly different to DVSS. It can be synergetic tool to DVSS for novice trainees of robotic surgery to get accustomed to the robotic surgical system and to improve their basic robotic surgical skills.


Assuntos
Procedimentos Cirúrgicos Robóticos/educação , Treinamento por Simulação , Adulto , Competência Clínica , Feminino , Humanos , Masculino , Análise e Desempenho de Tarefas , Adulto Jovem
12.
J Vet Sci ; 20(2): e6, 2019 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-30944529

RESUMO

The recent emergence of Staphylococcus schleiferi in dogs with otitis externa or skin and soft tissue infections has become a significant zoonotic issues. In the current study, we investigated 1) the carriage rates of S. schleiferi among major staphylococci in healthy dogs and dogs with otitis externa, 2) antibiotic susceptibility profiles of S. schleiferi, particularly methicillin resistance (MR), and 3) virulence factors associated with skin and soft tissue infections such as ability to form biofilm, resistance to cationic antimicrobial peptides (CAMPs), and carriage of staphylococcal enterotoxin genes. Among the 21 S. schleiferi isolates, 5 isolates (24%) were determined to be methicillin-resistant (MRSS). Staphylococcal cassette chromosome mec (SCCmec) typing revealed the presence of SCCmec type V in 4 MRSS isolates and type VII in one MRSS. Higher levels of antibiotic resistance, especially multidrug resistance, were observed in MRSS isolates compared to the methicillin-susceptible S. schleiferi (MSSS) isolates. In addition, MRSS isolates exhibited enhanced ability to form biofilm under static condition and all the 5 MRSS isolates carried three or more enterotoxin genes. However, there were no significant differences in resistance to CAMPs between MRSS and MSSS isolates. These findings suggest that coagulase-negative S. schleiferi is becoming more prevalent in canine otitis externa cases. Our results also highlight the presence of multidrug-resistant MRSS isolates with enhanced biofilm production and carriage of multiple enterotoxins.


Assuntos
Antibacterianos/uso terapêutico , Doenças do Cão/microbiologia , Otite Externa/veterinária , Infecções Cutâneas Estafilocócicas/veterinária , Staphylococcus , Animais , Peptídeos Catiônicos Antimicrobianos/farmacologia , Portador Sadio/microbiologia , Portador Sadio/veterinária , Doenças do Cão/tratamento farmacológico , Cães , Farmacorresistência Bacteriana , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Testes de Sensibilidade Microbiana/veterinária , Otite Externa/tratamento farmacológico , Otite Externa/microbiologia , Infecções Cutâneas Estafilocócicas/tratamento farmacológico , Infecções Cutâneas Estafilocócicas/microbiologia , Staphylococcus/efeitos dos fármacos , Staphylococcus/isolamento & purificação , Fatores de Virulência
13.
J Microbiol Biotechnol ; 28(11): 1937-1945, 2018 Nov 28.
Artigo em Inglês | MEDLINE | ID: mdl-30562885

RESUMO

Malassezia pachydermatis is a commensal yeast found on the skin of dogs. However, M. pachydermatis is also considered an opportunistic pathogen and is associated with various canine skin diseases including otitis externa and atopic dermatitis, which usually require treatment using an azole antifungal drug, such as ketoconazole. In this study, we isolated a ketoconazole-resistant strain of M. pachydermatis, designated "KCTC 27587," from the external ear canal of a dog with otitis externa and analyzed its resistance mechanism. To understand the mechanism underlying ketoconazole resistance of the clinical isolate M. pachydermatis KCTC 27587, the whole genome of the yeast was sequenced using the PacBio platform and was compared with M. pachydermatis type strain CBS 1879. We found that a ~84-kb region in chromosome 4 of M. pachydermatis KCTC 27587 was tandemly quadruplicated. The quadruplicated region contains 52 protein coding genes, including the homologs of ERG4 and ERG11, whose overexpression is known to be associated with azole resistance. Our data suggest that the quadruplication of the ~84-kb region may be the cause of the ketoconazole resistance in M. pachydermatis KCTC 27587.


Assuntos
Antifúngicos/farmacologia , Doenças do Cão/microbiologia , Farmacorresistência Fúngica/genética , Genômica , Cetoconazol/farmacologia , Malassezia/efeitos dos fármacos , Otite Externa/veterinária , Animais , Duplicação Cromossômica , Cães , Perfilação da Expressão Gênica , Malassezia/genética , Malassezia/isolamento & purificação , Testes de Sensibilidade Microbiana , Otite Externa/microbiologia
14.
Anal Sci ; 34(12): 1427-1432, 2018 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-30224566

RESUMO

A novel fluorescent staining protocol to detect phosphoproteins in sodium dodecyl sulfate-polyacrylamide gels using a fluorescence sensor, 1-(2-hydroxy-1-naphthylazo)-2-naphthol-4-sulfonic acid sodium salt (Calcon), was developed. This method yields results within 135 min, with the sensitivities of 15 ng of α-casein and ß-casein, and 62.5 ng of κ-casein, respectively. Since non-phosphoproteins have shown negative signals that are distinctly different from positive signals of phosphoproteins, this detection method allows one to monitor phosphoproteins with high specificity. Furthermore, a total protein profile can be achieved before a destaining step using a scanner with rapid and low-cost without further total protein staining.


Assuntos
Compostos Azo/química , Eletroforese em Gel de Poliacrilamida/métodos , Corantes Fluorescentes/química , Fosfoproteínas/análise , Coloração e Rotulagem/métodos , Caseínas/análise , Fluorescência , Sensibilidade e Especificidade , Soroalbumina Bovina/análise
15.
Methods Mol Biol ; 1853: 53-64, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30097930

RESUMO

A fast and matrix-assisted laser desorption/ionization-mass spectrometry compatible protein staining method in one- and two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis is described. It is based on the counterion dye staining method that employs oppositely charged two dyes, zincon and ethyl violet to form an ion-pair complex. The protocol including fixing, staining, and quick washing steps can be completed in 1-1.5 h depending upon gel thickness. It has the sensitivity comparable to the colloidal Coomassie Brilliant Blue G stain using phosphoric acid as a component of staining solution (4-8 ng). The counterion dye stain does not induce protein modifications that complicate interpretation of peptide mapping data from mass spectrometry. Considering the speed, sensitivity, and compatibility with mass spectrometry, the counterion dye stain may be more practical than any other dye-based protein stains for routine proteomic researches.


Assuntos
Corantes , Eletroforese em Gel Bidimensional , Eletroforese em Gel de Poliacrilamida , Proteínas , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , Coloração e Rotulagem , Corantes/química , Eletroforese em Gel Bidimensional/métodos , Eletroforese em Gel de Poliacrilamida/métodos , Proteínas/análise , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz/métodos , Coloração e Rotulagem/métodos
16.
Methods Mol Biol ; 1853: 65-73, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30097931

RESUMO

In order to detect phosphoproteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), an easy and fast fluorescent detection method is described. 8-Quinolinol can form ternary complexes in the gel matrix contributed by the affinity of aluminum ion to the phosphate groups on the proteins and the metal chelating property of 8-Quinolinol, exhibiting strong fluorescence in ultraviolet light. It can visualize as little as 4-8 ng of α-casein and ß-casein, 15-31 ng of ovalbumin and κ-casein within 70 min. The approach utilizing 8-quinolinol could be an alternative staining method for phosphoproteomics.


Assuntos
Eletroforese em Gel de Poliacrilamida , Oxiquinolina , Fosfoproteínas , Coloração e Rotulagem , Animais , Eletroforese em Gel Bidimensional , Eletroforese em Gel de Poliacrilamida/métodos , Camundongos , Oxiquinolina/química , Fosfoproteínas/análise , Fosfoproteínas/química , Coloração e Rotulagem/métodos
17.
J Vet Med Sci ; 80(9): 1445-1449, 2018 Sep 26.
Artigo em Inglês | MEDLINE | ID: mdl-29998924

RESUMO

A 12-year-old neutered male domestic shorthair cat was presented with a 3-year history of multiple nonpruritic, ulcerated, plaque-like skin lesions but no other clinical signs. A systemic examination revealed mild lymphadenopathy. Histopathologic analysis of the skin lesions revealed multicentric basosquamous carcinoma (BSC). Immunohistochemical analysis, PCR, and sequencing detected Felis catus papillomavirus type 2 (FcaPV-2) within the tumors. As BSC is rare in cats, clinical behavior has not been established. To our knowledge, this is the first case report to demonstrate detection of FcaPV-2 within a BSC in a domestic cat.


Assuntos
Carcinoma Basoescamoso/veterinária , Doenças do Gato/virologia , Papillomaviridae/isolamento & purificação , Neoplasias Cutâneas/veterinária , Animais , Carcinoma Basoescamoso/virologia , Gatos , DNA Viral , Masculino , Neoplasias Cutâneas/virologia
18.
J Vet Sci ; 19(5): 585-591, 2018 Sep 30.
Artigo em Inglês | MEDLINE | ID: mdl-29929355

RESUMO

Dogs serve human society in various ways by working at tasks that are based on their superior olfactory sensitivity. However, it has been reported that only about half of all trained dogs may qualify as working dogs through conventional breeding management because proper temperament and health are needed in addition to their innate scent detection ability. To overcome this low efficiency of breeding qualified working dogs, and to reduce the enormous costs of maintaining unqualified dogs, somatic cell nuclear transfer has been applied in the propagation of working dogs. Herein, we review the history of cloning working dogs and evaluate the health development, temperaments, and behavioral similarities among the cloned dogs. We also discuss concerns about dog cloning including those related to birth defects, lifespan, and cloning efficiency.


Assuntos
Clonagem de Organismos/veterinária , Cães/psicologia , Técnicas de Transferência Nuclear/veterinária , Temperamento , Animais , Comportamento Animal
19.
Integr Biol (Camb) ; 9(11): 857-867, 2017 Nov 13.
Artigo em Inglês | MEDLINE | ID: mdl-29098213

RESUMO

RIG-I-like receptors (RLRs) are cytoplasmic sensors of viral RNA that trigger the signaling cascade that leads to type I interferon (IFN) production. Transcriptional induction of RLRs by IFN is believed to play the role of positive feedback to further amplify viral sensing. We found that RLRs and several other IFN-stimulated genes (ISGs) are induced early in viral infection independent of IFN. Expression of these early ISGs requires IRF3/IRF7 and is highly correlated amongst them. Simultaneous detection of mRNA of IFNB1, viral replicase, and ISGs revealed distinct populations of IFNB1 expressing and non-expressing cells which are highly correlated with the levels of early ISGs but are uncorrelated with IFN-dependent ISGs and viral gene expression. Individual expression of RLRs made IFNB1 expression more robust and earlier, suggesting a causal relation between levels of RLR and induction of IFN.


Assuntos
Regulação da Expressão Gênica , Interferon beta/metabolismo , Análise de Célula Única , Animais , Embrião de Galinha , Chlorocebus aethiops , Citoplasma/metabolismo , Proteína DEAD-box 58/metabolismo , Células HeLa , Células Hep G2 , Humanos , Imunidade Inata , Hibridização in Situ Fluorescente , Fator Regulador 3 de Interferon/metabolismo , Fator Regulador 7 de Interferon/metabolismo , Helicase IFIH1 Induzida por Interferon/metabolismo , Interferência de RNA , RNA Mensageiro/metabolismo , Receptores Imunológicos , Transdução de Sinais , Processos Estocásticos , Células Vero
20.
Electrophoresis ; 38(24): 3079-3085, 2017 12.
Artigo em Inglês | MEDLINE | ID: mdl-28833374

RESUMO

Protein phosphorylation, one of the most important post-translational modifications, plays critical roles in many biological processes. Thus, it is necessary to precisely detect, identify and understand the phosphoproteins from protein mixture for the study of cell biology. We introduce a sensitive and specific detection method for phosphoproteins in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Anthracene Chrome Red A (ACRA) combined with the trivalent metal ion (Al3+ ) is converted to fluorescent complex and the fluorescence is sharply increased by a change of pH environment. Phosphoproteins and non-phosphoproteins can be easily distinguished by the fluorescence quenching due to the structural change of ACRA-Al3+ -phosphoprotein complex, unlike non-phosphoprotein complex. The method using ACRA is a negative staining based on the fluorescence quenching and has a high sensitivity comparable to Pro-Q Diamond stain. ACRA stain can detect 1-2 ng of α-casein and ß-casein, 8-16 ng of ovalbumin (OVA) and κ-casein within 130 min. Moreover, the ACRA stain showed similar linear dynamic ranges and RSD to Pro-Q stain. The linear dynamic ranges of ACRA and the values of correlation coefficient were for OVA (8-500 ng, correlation coefficient r = 0.999), α-casein (4-500 ng, r = 0.992), ß-casein (4-500 ng, r = 0.996), and κ-casein (8-500 ng, 0.998), respectively. On the other hand, the values of the relative standard deviations (RSD) ranged from 2.33 to 3.56% for ACRA. The method is sensitive, specific, simple, rapid and compatible with total protein stain such as SYPRO Ruby stain. Therefore, ACRA stain can be an advanced method for phosphoprotein detection in gels.


Assuntos
Antracenos/química , Corantes/química , Eletroforese em Gel de Poliacrilamida/métodos , Fosfoproteínas/análise , Coloração e Rotulagem/métodos , Antracenos/análise , Corantes/análise , Modelos Lineares , Fosfoproteínas/química , Reprodutibilidade dos Testes , Sensibilidade e Especificidade
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