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1.
Int J Obes (Lond) ; 2024 Sep 04.
Artículo en Inglés | MEDLINE | ID: mdl-39232101

RESUMEN

BACKGROUND: Obesity paradox addressing all-cause mortality has been described in several chronic total occlusion (CTO) studies. However, the impact of aging on long-term cardiac events in patients with overweight and obesity with CTO recanalization were less studied. METHODS: A total of 458 patients (64.4 ± 11.3 years, 403 male) with CTO interventions were enrolled. The overweight/obesity group included 311 patients with body mass index (BMI) ≧24 kg/m2 and the non-obesity group included 147. With a median follow-up of 40.0 (17.9-61.4) months, 422 patients with successful true-lumen recanalization were further assessed for target lesion failure [TLF: cardiac death, target vessel myocardial infarction (TVMI), target lesion revascularization (TLR)]. RESULTS: At follow-up, the rates of cardiac death, TVMI, TLR, TLF, and stent thrombosis were 1.9%, 1.9%, 9.2%, 10.7%, and 0.5%, respectively. The TVMI-free survival was borderline better (p = 0.067 by log-rank test) in overweight/obesity than non-obesity group. Among patients <65 years of age, the TVMI-free survival was significantly better in the overweight/obesity group (p = 0.013 compared to non-obesity group by log-rank test). In multivariate Cox regression model, the non-obesity patients younger than 65 years were at a higher risk of TVMI, not only among those <65 years of age (hazard ratio = 11.0, 95% CI = 1.1-106.0) but also among the whole patients (hazard ratio=6.9, 95% CI = 1.4-35.1) with successful CTO recanalization. CONCLUSIONS: For those with true-lumen recanalized CTO, the higher risk of TVMI after successful recanalization was rather evident in patients <65 years of age and without overweight/obesity, suggesting that aging might attenuate prognostic significance of "obesity paradox" for CTO interventions.

2.
BMC Med Ethics ; 24(1): 59, 2023 08 04.
Artículo en Inglés | MEDLINE | ID: mdl-37542298

RESUMEN

BACKGROUND: Adherence to ethical guidelines and regulations and protecting and respecting the dignity and autonomy of participants by obtaining a valid informed consent form (ICF) prior to participation in research are crucial; The subjects did not add signatures next to the corrections made to signatures or dates on the ICF, Multiple signatures in other fields, ICF missing/missing signature, Incorrect ICF version Signed after modification, Correction tape used to correct signature, Impersonated signature, Non-research-member signature, however, ICFs are often not properly completed, which must be addressed. This study analyzed ICF signing errors and implemented measures to reduce or prevent these errors. METHODS: We used the plan-do-check-act (PDCA) cycle to help improve the correctness and validity of ICF signing. RESULTS: Interim and final reports from January 2016 to February 2020 including 363 ICFs were studied. The total proportion of correct ICF signatures (200, 83.3%) following the PDCA intervention was significantly higher than that before the intervention (P < 0.05). Analysis of the types of signing error demonstrated that signature errors were significantly reduced after the intervention, particularly for subjects did not add signatures next to the corrections made to signatures or dates on the ICF (16, 6.7%) and impersonated signature (0; P < 0.05). CONCLUSIONS: The proportions of other error types-multiple signatures in other fields, missing or unsigned ICF, incorrect signature order, incorrect ICF version, use of correction tape to correct signature, and non-medical profession members signing the ICF-did not differ significantly.


Asunto(s)
Formularios de Consentimiento , Mejoramiento de la Calidad , Humanos , Comprensión , Respeto , Consentimiento Informado
3.
Mar Drugs ; 20(8)2022 Jul 27.
Artículo en Inglés | MEDLINE | ID: mdl-36005485

RESUMEN

Overexpressed EGFR and mutant K-Ras play vital roles in therapeutic resistance in colorectal cancer patients. To search for an effective therapeutic protocol is an urgent task. A secondary metabolite in the sponge Hippospongia sp., Heteronemin, has been shown to induce anti-proliferation in several types of cancers. A thyroxine-deaminated analogue, tetrac, binds to integrin αvß3 to induce anti-proliferation in different cancers. Heteronemin- and in combination with tetrac-induced antiproliferative effects were evaluated. Tetrac enhanced heteronemin-induced anti-proliferation in HT-29 cells (KRAS WT CRC) and HCT-116 cells (KRAS MT CRC). Heteronemin and tetrac arrested cell cycle in different phases. Combined treatment increased the cell accumulation in sub-G1 and S phases. The combined treatment also induced the inactivation of EGFR signaling and downregulated the phosphorylated ERK1/2 protein in both cell lines. Heteronemin and the combination showed the downregulation of the phosphorylated and total PI3K protein in HT-29 cells (KRAS WT CRC). Results by NanoString technology and RT-qPCR revealed that heteronemin and combined treatment suppressed the expression of EGFR and downstream genes in HCT-116 cells (KRAS MT CRC). Heteronemin or combined treatment downregulated genes associated with cancer progression and decreased cell motility. Heteronemin or the combined treatment suppressed PD-L1 expression in both cancer cell lines. However, only tetrac and the combined treatment inhibited PD-L1 protein accumulation in HT-29 cells (KRAS WT CRC) and HCT-116 cells (KRAS MT CRC), respectively. In summary, heteronemin induced anti-proliferation in colorectal cancer cells by blocking the EGFR-dependent signal transduction pathway. The combined treatment further enhanced the anti-proliferative effect via PD-L1 suppression. It can be an alternative strategy to suppress mutant KRAS resistance for anti-EGFR therapy.


Asunto(s)
Neoplasias Colorrectales , Tiroxina , Antígeno B7-H1/metabolismo , Línea Celular Tumoral , Proliferación Celular , Neoplasias Colorrectales/metabolismo , Receptores ErbB/metabolismo , Humanos , Mutación , Proteínas Proto-Oncogénicas p21(ras)/genética , Proteínas Proto-Oncogénicas p21(ras)/metabolismo , Proteínas Proto-Oncogénicas p21(ras)/farmacología , Transducción de Señal , Terpenos , Tiroxina/análogos & derivados
4.
Int J Mol Sci ; 22(3)2021 Jan 30.
Artículo en Inglés | MEDLINE | ID: mdl-33573352

RESUMEN

The rice cell suspension culture system is a good way to produce recombinant human proteins, owing to its high biosafety and low production cost. Human Octamer-binding Transcription Factor 4 (Oct4) is a fundamental transcription factor responsible for maintaining human pluripotent embryonic stem cells. Recombinant Oct4 protein has been used to induce pluripotent stem cells. In this study, recombinant Oct4 proteins are produced via a sugar starvation-inducible αAmy3/RAmy3D promoter-signal peptide-based rice recombinant protein expression system. Oct4 mRNAs accumulate in the transgenic rice suspension cells under sugar starvation. The Oct4 recombinant protein is detected in the transgenic rice suspension cells, and its highest yield is approximately 0.41% of total cellular soluble proteins after one day of sugar starvation. The rice cell-synthesized recombinant human Oct4 protein show DNA-binding activity in vitro, which implies that the protein structure is correct for enabling specific binding to the target DNA motif.


Asunto(s)
Técnicas de Cultivo de Célula/métodos , Factor 3 de Transcripción de Unión a Octámeros/aislamiento & purificación , Oryza/citología , Células Cultivadas , Contención de Riesgos Biológicos , Regulación de la Expresión Génica de las Plantas , Factor 3 de Transcripción de Unión a Octámeros/genética , Factor 3 de Transcripción de Unión a Octámeros/metabolismo , Oryza/genética , Proteínas de Plantas/genética , Plantas Modificadas Genéticamente/citología , Plantas Modificadas Genéticamente/genética , Regiones Promotoras Genéticas/genética , Proteínas Recombinantes/genética , Proteínas Recombinantes/aislamiento & purificación , Proteínas Recombinantes/metabolismo , alfa-Amilasas/genética
5.
Mar Drugs ; 18(7)2020 Jul 02.
Artículo en Inglés | MEDLINE | ID: mdl-32630719

RESUMEN

BACKGROUND: Heteronemin, a marine sesterterpenoid-type natural product, possesses an antiproliferative effect in cancer cells. In addition, heteronemin has been shown to inhibit p53 expression. Our laboratory has demonstrated that the thyroid hormone deaminated analogue, tetrac, activates p53 and induces antiproliferation in colorectal cancer. However, such drug mechanisms are still to be studied in oral cancer cells. METHODS: We investigated the antiproliferative effects by Cell Counting Kit-8 and flow cytometry. The signal transduction pathway was measured by Western blotting analyses. Quantitative PCR was used to evaluate gene expression regulated by heteronemin, 3,3',5,5'-tetraiodothyroacetic acid (tetrac), or their combined treatment in oral cancer cells. RESULTS: Heteronemin inhibited not only expression of proliferative genes and Homo Sapiens Thrombospondin 1 (THBS-1) but also cell proliferation in both OEC-M1 and SCC-25 cells. Remarkably, heteronemin increased TGF-ß1 expression in SCC-25 cells. Tetrac suppressed expression of THBS-1 but not p53 expression in both cancer cell lines. Furthermore, the synergistic effect of tetrac and heteronemin inhibited ERK1/2 activation and heteronemin also blocked STAT3 signaling. Combined treatment increased p53 protein and p53 activation accumulation although heteronemin inhibited p53 expression in both cancer cell lines. The combined treatment induced antiproliferation synergistically more than a single agent. CONCLUSIONS: Both heteronemin and tetrac inhibited ERK1/2 activation and increased p53 phosphorylation. They also inhibited THBS-1 expression. Moreover, tetrac suppressed TGF-ß expression combined with heteronemin to further enhance antiproliferation and anti-metastasis in oral cancer cells.


Asunto(s)
Carcinoma/tratamiento farmacológico , Proliferación Celular/efectos de los fármacos , Neoplasias Gingivales/tratamiento farmacológico , Terpenos/farmacología , Tiroxina/análogos & derivados , Antineoplásicos/administración & dosificación , Antineoplásicos/farmacología , Línea Celular Tumoral , Regulación Neoplásica de la Expresión Génica/efectos de los fármacos , Humanos , Terpenos/administración & dosificación , Tiroxina/administración & dosificación , Tiroxina/farmacología
6.
Mol Cell ; 43(5): 811-22, 2011 Sep 02.
Artículo en Inglés | MEDLINE | ID: mdl-21884981

RESUMEN

Epithelial-mesenchymal transition (EMT) is important for organ development, metastasis, cancer stemness, and organ fibrosis. Molecular mechanisms to coordinately regulate hypoxia-induced EMT remain elusive. Here, we show that HIF-1α-induced histone deacetylase 3 (hdac3) is essential for hypoxia-induced EMT and metastatic phenotypes. Change of specific chromatin states is associated with hypoxia-induced EMT. Under hypoxia, HDAC3 interacts with hypoxia-induced WDR5, recruits the histone methyltransferase (HMT) complex to increase histone H3 lysine 4 (H3K4)-specific HMT activity, and activates mesenchymal gene expression. HDAC3 also serves as an essential corepressor to repress epithelial gene expression. Knockdown of WDR5 abolishes mesenchymal gene activation but not epithelial gene repression during hypoxia. These results indicate that hypoxia induces different chromatin modifiers to coordinately regulate EMT through distinct mechanisms.


Asunto(s)
Transición Epitelial-Mesenquimal/fisiología , Histona Desacetilasas/metabolismo , N-Metiltransferasa de Histona-Lisina/metabolismo , Animales , Hipoxia de la Célula , Células Cultivadas , Transición Epitelial-Mesenquimal/genética , Expresión Génica , Histona Desacetilasas/genética , N-Metiltransferasa de Histona-Lisina/genética , Humanos , Ratones , Ratones Endogámicos NOD , Ratones SCID , Transfección
7.
Am J Emerg Med ; 34(1): 79-82, 2016 Jan.
Artículo en Inglés | MEDLINE | ID: mdl-26472508

RESUMEN

OBJECTIVE: The study aims to assess if electrocardiographic (ECG) abnormalities could predict the development of neurogenic pulmonary edema (NPE) within 24 hours in cases of spontaneous subarachnoid hemorrhage (SAH). METHODS: We studied prospectively a cohort of 269 adult patients with nontraumatic SAH in an emergency department of a university-affiliated medical center. A 12-lead ECG was taken for these patients. The patients were stratified into NPE and non-NPE based on serially clinical and radiologic findings. The ECG abnormalities were compared between these 2 groups of patients. RESULTS: Compared with the non-NPE (n = 229), the NPE (n = 40) had significantly higher World Federation of Neurological Surgeons class (P < .001), higher Hunt-Hess scale (P < .001), and higher prevalence of diabetes mellitus (P = .033). In addition, the percentage of ECG morphological abnormality was significantly higher in NPE, in which nonspecific ST- or T-wave changes (NSSTTCs) are significantly higher. Multiple logistic regression model identified World Federation of Neurological Surgeons class (95% confidence interval [CI], 2.6-13.3; P < .001), abnormal Q or QS wave (95% CI, 1.1-9.1; P = .038), and NSSTTCs (95% CI, 1.2-7.5; P = .016) as the significant variables associated with NPE. CONCLUSIONS: Electrocardiographic abnormalities, especially abnormal Q or QS wave and NSSTTCs, may predict the development of NPE within 24 hours in adult patients with spontaneous SAH.


Asunto(s)
Electrocardiografía , Servicio de Urgencia en Hospital , Edema Pulmonar/etiología , Hemorragia Subaracnoidea/fisiopatología , Adulto , Anciano , Femenino , Frecuencia Cardíaca , Humanos , Masculino , Persona de Mediana Edad , Estudios Prospectivos , Factores de Riesgo , Hemorragia Subaracnoidea/complicaciones , Factores de Tiempo
8.
J Interprof Care ; 30(4): 536-8, 2016 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-27332501

RESUMEN

This article reports the results from a study that employed an interprofessional crew resource management (CRM) education programme in the emergency and critical care departments. The study aimed to investigate the effectiveness of this intervention of participants' satisfaction and safety attitude changes using a satisfaction questionnaire and the Human Factors Attitude Survey (HFAS). Overall, participants responded positively to the CRM training-93.4% were satisfied, 93.1% agreed that it enhanced patient safety and care quality, 85.7% agreed that it increased their confidence, 86.4% agreed that it reduced practice errors, and 90.8% agreed that it would change their behaviours. Overall, the participants reported positive changes in their attitudes regarding 22 of the 23 HFAS questions.


Asunto(s)
Conocimientos, Actitudes y Práctica en Salud , Personal de Salud/educación , Capacitación en Servicio , Relaciones Interprofesionales , Grupo de Atención al Paciente , Seguridad del Paciente , Femenino , Recursos en Salud/organización & administración , Humanos , Masculino , Evaluación de Programas y Proyectos de Salud , Encuestas y Cuestionarios
9.
Proteins ; 82(7): 1519-26, 2014 Jul.
Artículo en Inglés | MEDLINE | ID: mdl-24470206

RESUMEN

Uridine diphosphate N-acetylglucosamine (UDP-GlcNAc) 2-epimerase catalyzes the interconversion of UDP-GlcNAc to UDP-N-acetylmannosamine (UDP-ManNAc), which is used in the biosynthesis of cell surface polysaccharides in bacteria. Biochemical experiments have demonstrated that mutation of this enzyme causes changes in cell morphology and the thermoresistance of the cell wall. Here, we present the crystal structures of Methanocaldococcus jannaschii UDP-GlcNAc 2-epimerase in open and closed conformations. A comparison of these crystal structures shows that upon UDP and UDP-GlcNAc binding, the enzyme undergoes conformational changes involving a rigid-body movement of the C-terminal domain. We also present the crystal structure of Bacillus subtilis UDP-GlcNAc 2-epimerase in the closed conformation in the presence of UDP and UDP-GlcNAc. Although a structural overlay of these two closed-form structures reveals that the substrate-binding site is evolutionarily conserved, some areas of the allosteric site are distinct between the archaeal and bacterial UDP-GlcNAc 2-epimerases. This is the first report on the crystal structure of archaeal UDP-GlcNAc 2-epimerase, and our results clearly demonstrate the changes between the open and closed conformations of this enzyme.


Asunto(s)
Proteínas Arqueales , Methanocaldococcus/enzimología , Uridina Difosfato N-Acetilglucosamina , Secuencia de Aminoácidos , Proteínas Arqueales/química , Proteínas Arqueales/metabolismo , Carbohidrato Epimerasas/química , Carbohidrato Epimerasas/metabolismo , Dominio Catalítico , Cristalografía por Rayos X , Isomerismo , Modelos Moleculares , Datos de Secuencia Molecular , Conformación Proteica , Alineación de Secuencia , Uridina Difosfato N-Acetilglucosamina/química , Uridina Difosfato N-Acetilglucosamina/metabolismo
10.
Patient Educ Couns ; 123: 108206, 2024 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-38422949

RESUMEN

OBJECTIVE: To examine the effectiveness of health education and counseling on the stages of change, decisional balance, and smoking cessations elf-efficacy in smokers with no intention of quitting. METHODS: A prospective self-controlled design was conducted between December 2020 and December 2022. The research period was divided into a control stage (first to fourth weeks) and an experimental stage (fifth to eighth weeks). Patients with coronary artery disease (CAD) and habitually smoked were recruited. Pearson correlation and a one-factor repeated-measurement analysis were performed to assess the effectiveness of the intervention. RESULTS: In total, 108 male CAD patients with a mean age of 58.1 years were recruited. After 4 weeks of the intervention, 55 (51%) exhibited behavior change (X 2 = 18.03, p = .001). The decisional balance and smoking cessation self-efficacy scores significantly improved in the experimental stage. No significant differences were observed in the control stage. CONCLUSIONS: Four weeks of health education and counseling could effectively improve participants' stage of change, decisional balance, and smoking cessation self-efficacy. PRACTICE IMPLICATION: Healthcare professionals can play key roles in helping CAD patients successfully quit smoking through individual education and counseling.


Asunto(s)
Cese del Hábito de Fumar , Humanos , Masculino , Persona de Mediana Edad , Cese del Hábito de Fumar/psicología , Estudios Prospectivos , Modelo Transteórico , Consejo , Educación en Salud , Atención a la Salud
11.
Ann Med ; 56(1): 2396076, 2024 Dec.
Artículo en Inglés | MEDLINE | ID: mdl-39193705

RESUMEN

BACKGROUND: As the burden and distribution of calcification within chronic total occlusion (CTO) lesions can be diverse, its effect on CTO recanalization using multiple devices and techniques is debatable. This study investigated the role of calcification in wiring-based intraplaque tracking techniques for CTO recanalization. METHODS: A modified J-CTO score without counting calcification was used to analyze the procedures of 458 consecutive patients who underwent CTO interventions. Failed guidewire crossing and intraplaque tracking were considered procedural failures. Recanalization time details were analyzed for successful procedures. RESULTS: In patients with calcified CTO, the rate of procedural success only significantly declined to be lower than that of noncalcified CTO when the modified J-CTO score was ≥3 (77% vs. 94%, p = 0.008). In 422 patients with successful procedures, the presence of calcification was irrelevant to guidewire crossing time, but was accompanied with longer time from guidewire cross to final angiogram when the modified J-CTO score was 1-2 (53 ± 35 vs. 35 ± 17 [noncalcified] min, p < 0.001). Multivariate analyses showed that calcification was independently associated with procedural failure (odds ratio [OR] = 5.1, 95% confidence interval [CI] = 1.4-18.3) in lesions with modified J-CTO ≥3, and prolonged angioplasty/stenting procedures >60 min (OR = 4.8, 95% CI = 2.2-10.2) in successfully recanalized lesions with modified J-CTO score 1-2. CONCLUSIONS: Using intraplaque guidewire tracking, calcification was unfavorable for very difficult CTO lesions, and caused prolongation of angioplasty time for lesions with moderate complexity. This suggested that the role of calcification in the J-CTO score could be altered when different recanalization techniques were applied for CTO interventions.


Since several commonly used scoring systems for grading the difficulty of CTO-PCI are derived from multiple recanalization techniques and devices, their application should be fundamental. However, most CTO interventionists usually have their own favored recanalization techniques in the real-world. As one of the parameters of J-CTO score, the findings of the study suggest that the interpretation of calcification during CTO-PCI could be altered and should be cautious if different recanalization technique was used.


Asunto(s)
Oclusión Coronaria , Humanos , Masculino , Femenino , Anciano , Persona de Mediana Edad , Oclusión Coronaria/diagnóstico por imagen , Oclusión Coronaria/cirugía , Intervención Coronaria Percutánea/efectos adversos , Intervención Coronaria Percutánea/métodos , Intervención Coronaria Percutánea/instrumentación , Calcificación Vascular/diagnóstico por imagen , Angiografía Coronaria/métodos , Estudios Retrospectivos , Índice de Severidad de la Enfermedad , Calcinosis/diagnóstico por imagen , Enfermedad Crónica , Resultado del Tratamiento
12.
Bioorg Med Chem Lett ; 23(3): 699-701, 2013 Feb 01.
Artículo en Inglés | MEDLINE | ID: mdl-23260351

RESUMEN

2-(3-Alkylaminophenyl)-6-(pyrrolidin-1-yl)quinolin-4-ones 1-3 were synthesized and screened for anti-proliferative activity against three human cancer cell lines, as well as the normal cell line Detroit 551. All of the synthesized target compounds 1-3 demonstrated potent cytotoxic activity against the cancer cell lines, but weak inhibitory activity toward the normal cell line. 2-(3-Methyl aminophenyl)-6-(pyrrolidin-1-yl)quinolin-4-one (1), one of the potent compounds in vitro, was also tested in an in vivo Hep3B xenograft nude mice model, and its significant anticancer activity was reconfirmed. Therefore, compound 1 merits further investigation as an antitumor clinical trial candidate and potential anticancer agent.


Asunto(s)
Compuestos de Anilina/síntesis química , Compuestos de Anilina/farmacología , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Quinolonas/síntesis química , Quinolonas/farmacología , Compuestos de Anilina/química , Animales , Línea Celular , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Humanos , Concentración 50 Inhibidora , Ratones , Ratones Desnudos , Estructura Molecular , Pirrolidinas , Quinolonas/química , Ensayos Antitumor por Modelo de Xenoinjerto
13.
Artículo en Inglés | MEDLINE | ID: mdl-23722855

RESUMEN

SaeR is the response regulator of the SaeRS two-component signal transduction system, which is involved in regulating bacterial autolysis and biofilm formation. SaeR comprises an N-terminal receiver domain and a C-terminal effector domain. The effector domain possesses DNA-binding and transactivation functions. Here, the effector domain of SaeR from Staphylococcus epidermidis was purified and crystallized using the sitting-drop vapour-diffusion method. The crystals diffracted to a resolution of 2.15 Å and belonged to space group P2(1)2(1)2(1), with unit-cell parameters a = 34.20, b = 53.78, c = 111.66 Å. Determining the structure will provide insights into the mechanisms underlying DNA binding.


Asunto(s)
Proteínas Bacterianas/química , ADN Bacteriano/química , Staphylococcus epidermidis , Proteínas Bacterianas/metabolismo , Cristalización , ADN Bacteriano/metabolismo , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Unión Proteica/fisiología , Estructura Terciaria de Proteína , Factores de Transcripción , Difracción de Rayos X
14.
Artículo en Inglés | MEDLINE | ID: mdl-23295494

RESUMEN

MJ0927 is a member of the Nif3 family and is widely distributed across living organisms. Although several crystal structures of Nif3 proteins have been reported, structural information on archaeal Nif3 is still limited. To understand the structural differences between bacterial and archaeal Nif3 proteins, MJ0927 from Methanocaldococcus jannaschii was purified and crystallized using the sitting-drop vapour-diffusion method. The crystals diffracted to a resolution of 2.47 Šand belonged to the orthorhombic space group C222, with unit-cell parameters a = 81.21, b = 172.94, c = 147.42 Å. Determination of this structure may provide insights into the function of MJ0927.


Asunto(s)
Proteínas Arqueales/química , Proteínas Arqueales/aislamiento & purificación , Methanococcaceae/química , Proteínas Arqueales/genética , Clonación Molecular , Cristalización , Cristalografía por Rayos X , Conformación Proteica
15.
BMC Vet Res ; 9: 161, 2013 Aug 13.
Artículo en Inglés | MEDLINE | ID: mdl-23937712

RESUMEN

BACKGROUND: Chicken anemia virus (CAV) is an important viral pathogen that causes anemia and severe immunodeficiency syndrome in chickens worldwide. Generally, CAV infection occurs via vertical transmission in young chicks that are less than two weeks old, which are very susceptible to the disease. Therefore, epidemiological investigations of CAV infection and/or the evaluation of the immunization status of chickens is necessary for disease control. Up to the present, systematically assessing viral protein antigenicity and/or determining the immunorelevant domain(s) of viral proteins during serological testing for CAV infection has never been performed. The expression, production and antigenic characterization of CAV viral proteins such as VP1, VP2 and VP3, and their use in the development of diagnostic kit would be useful for CAV infection prevention. RESULTS: Three CAV viral proteins VP1, VP2 and VP3 was separately cloned and expressed in recombinant E. coli. The purified recombinant CAV VP1, VP2 and VP3 proteins were then used as antigens in order to evaluate their reactivity against chicken sera using indirect ELISA. The results indicated that VP2 and VP3 show good immunoreactivity with CAV-positive chicken sera, whereas VP1 was found to show less immunoreactivity than VP2 and VP3. To carry out the further antigenic characterization of the immunorelevant domains of the VP2 and VP3 proteins, five recombinant VP2 subunit proteins (VP2-435N, VP2-396N, VP2-345N, VP2-171C and VP2-318C) and three recombinant VP3 subunit proteins (VP3-123N, VP3-246M, VP3-366C), spanning the defined regions of VP2 and VP3 were separately produced by an E. coli expression system. These peptides were then used as antigens in indirect ELISAs against chicken sera. The results of these ELISAs using truncated recombinant VP2 and VP3 subunit proteins as coating antigen showed that VP2-345N, VP2-396N and VP3-246M gave good immunoreactivity with CAV-positive chicken sera compared to the other subunit proteins. Moreover, the VP2-396N and VP2-345 based ELISAs had better sensitivity (97.5%) and excellent specificity (100%) during serodiagnosis testing using a mean plus three standard deviations cut-off. The VP3-246M based ELISA showed a sensitivity of 85% and a specificity of 100% at the same cut-off value. CONCLUSIONS: This is the first report to systematically assess the antigenic characteristics of CAV viral proteins for sero-diagnosis purposes. Purified recombinant VP2-396N and VP2-345N subunit proteins, which span defined regions of VP2, were demonstrated to have good antigenicity and higher sensitivities than VP3-246M and were able to recognize CAV-positive chicken serum using an ELISA assay. The defined antigenicity potential of these chimeric subunit proteins produced by expression in E. coli seem to have potential and could be useful in the future for the development of the CAV diagnostic tests based on a subunit protein ELISA system.


Asunto(s)
Proteínas de la Cápside/inmunología , Virus de la Anemia del Pollo/inmunología , Enfermedades de las Aves de Corral/inmunología , Animales , Antígenos Virales/biosíntesis , Antígenos Virales/inmunología , Proteínas de la Cápside/biosíntesis , Virus de la Anemia del Pollo/metabolismo , Pollos/virología , Infecciones por Circoviridae/diagnóstico , Infecciones por Circoviridae/inmunología , Infecciones por Circoviridae/veterinaria , Infecciones por Circoviridae/virología , Clonación Molecular , Ensayo de Inmunoadsorción Enzimática/veterinaria , Escherichia coli/metabolismo , Enfermedades de las Aves de Corral/diagnóstico , Enfermedades de las Aves de Corral/metabolismo , Enfermedades de las Aves de Corral/virología , Proteínas Recombinantes/biosíntesis , Proteínas Recombinantes/inmunología , Pruebas Serológicas/métodos , Pruebas Serológicas/veterinaria
16.
Molecules ; 18(7): 8028-45, 2013 Jul 08.
Artículo en Inglés | MEDLINE | ID: mdl-23884134

RESUMEN

Phosphorylation of 2-(3-hydroxy-5-methoxyphenyl)-6,7-methylenedioxy-1H-quinolin-4-one (1) afforded diphosphate 2. We found that, upon treatment with methanol under mild conditions, 2 can undergo facile and highly regioselective dephosphorylation to give the monophosphate 3, with a phosphate group remaining on the phenyl ring. The details of the dephosphorylation process were postulated and then probed by LC-MS and HPLC analyses. Furthermore, as a preliminary study, the water soluble monophosphate prodrug 4 was tested for antitumor activity against a MCF-7 xenograft nude mice model.


Asunto(s)
Neoplasias/tratamiento farmacológico , Profármacos/síntesis química , Profármacos/farmacocinética , Compuestos de Quinolinio/farmacología , Animales , Antineoplásicos/síntesis química , Antineoplásicos/farmacología , Difosfatos/química , Difosfatos/metabolismo , Humanos , Células MCF-7/efectos de los fármacos , Ratones , Fosforilación , Profármacos/química , Compuestos de Quinolinio/síntesis química , Solubilidad , Relación Estructura-Actividad , Ensayos Antitumor por Modelo de Xenoinjerto
17.
BMC Biotechnol ; 12: 27, 2012 Jun 06.
Artículo en Inglés | MEDLINE | ID: mdl-22672291

RESUMEN

BACKGROUND: Apoptin, a nonstructural protein encoded by the VP3 gene of chicken anemia virus (CAV), has been shown to not only induce apoptosis when introduced into the precursors of chicken thymocytes, but has been found to specifically kill human cancer cells, tumor cell and transformed cells without affecting the proliferation of normal cells. This tumor-specific apoptotic characteristic of the protein potentially may allow the development of a protein drug that has applications in tumor therapy. However, several major problems, which include poor expression and poor protein solubility, have hampered the production of apoptin in bacteria. RESULTS: Significantly increased expression of recombinant full-length apoptin that originated from chicken anemia virus was demonstrated using an E. coli expression system. The CAV VP3 gene was fused with a synthetic sequence containing a trans-acting activator of transcription (TAT) protein transduction domain (PTD). The resulting construct was cloned into various different expression vectors and these were then expressed in various E. coli strains. The expression of the TAT-Apoptin in E. coli was significantly increased when TAT-Apoptin was fused with GST-tag rather than a His-tag. When the various rare amino acid codons of apoptin were optimized, the expression level of the GST-TAT-Apoptin(opt) in E. coli BL21(DE3) was significantly further increased. The highest protein expression level obtained was 8.33 g/L per liter of bacterial culture after induction with 0.1 mM IPTG for 4 h at 25 °C. Moreover, approximately 90% of the expressed GST-TAT-Apoptin(opt) under these conditions was soluble. After purification by GST affinity chromatography, the purified recombinant TAT-Apoptin(opt) protein was used to evaluate the recombinant protein's apoptotic activity on tumor cells. The results demonstrated that the E. coli-expressed GST-TAT-apoptin(opt) showed apoptotic activity and was able to induce human premyelocytic leukemia HL-60 cells to enter apoptosis. CONCLUSIONS: On expression in E. coli, purified recombinant TAT-Apoptin(opt) that has been fused to a GST tag and had its codons optimized, was found to have great potential. This protein may in the future allow the development of a therapeutic protein that is able to specifically kill tumor cells.


Asunto(s)
Proteínas de la Cápside/genética , Proteínas de la Cápside/metabolismo , Virus de la Anemia del Pollo/genética , Neoplasias/tratamiento farmacológico , Ingeniería de Proteínas , Apoptosis/efectos de los fármacos , Secuencia de Bases , Proteínas de la Cápside/aislamiento & purificación , Proteínas de la Cápside/uso terapéutico , Línea Celular Tumoral , Virus de la Anemia del Pollo/metabolismo , Codón , Escherichia coli/genética , Escherichia coli/metabolismo , Expresión Génica , Humanos , Datos de Secuencia Molecular , Neoplasias/fisiopatología , Proteínas Recombinantes de Fusión/genética , Proteínas Recombinantes de Fusión/aislamiento & purificación , Proteínas Recombinantes de Fusión/metabolismo , Proteínas Recombinantes de Fusión/uso terapéutico
18.
Acta Crystallogr Sect F Struct Biol Cryst Commun ; 68(Pt 10): 1234-6, 2012 Oct 01.
Artículo en Inglés | MEDLINE | ID: mdl-23027755

RESUMEN

The RecQ proteins are a highly conserved group of DNA helicases which play crucial roles in the maintenance of genome stability. DrRecQ from the radioresistant bacterium Deinococcus radiodurans is a special member of the RecQ family because it contains three Helicase-and-RNase-D-C-terminal (HRDC) domains at the C-terminus. The helicase catalytic core is essential for ATPase and DNA-unwinding activities. In this work, the helicase catalytic core of DrRecQ was expressed in Escherichia coli, purified and crystallized. Crystals were obtained using the sitting-drop vapour diffusion method and X-ray diffraction data were collected to 2.9 Šresolution. The crystals belong to space group P2(1)2(1)2(1), with unit-cell parameters a = 84.75, b = 95.61, c = 183.83 Å.


Asunto(s)
Deinococcus/enzimología , RecQ Helicasas/química , Dominio Catalítico , Cristalización , Expresión Génica , RecQ Helicasas/genética , RecQ Helicasas/aislamiento & purificación , Difracción de Rayos X
19.
Artículo en Inglés | MEDLINE | ID: mdl-22750871

RESUMEN

The D-alanyl lipoteichoic acids (D-alanyl LTAs) present in the cell walls of Gram-positive bacteria play crucial roles in autolysis, cation homeostasis and biofilm formation. The alanylation of LTAs requires the D-alanyl carrier protein DltC to transfer D-Ala onto a membrane-associated LTA. Here, DltC from Staphylococcus epidermidis (SeDltC) was purified and crystallized using the sitting-drop vapour-diffusion method. The crystals diffracted to a resolution of 1.83 Šand belonged to space group P2, with unit-cell parameters a = 66.26, b = 53.28, c = 88.05 Å, ß = 98.22°. The results give a preliminary crystallographic analysis of SeDltC and shed light on the functional role of DltC in the alanylation of LTAs.


Asunto(s)
Proteínas Bacterianas/química , Proteínas Portadoras/química , Staphylococcus epidermidis/química , Proteínas Bacterianas/genética , Proteínas Bacterianas/aislamiento & purificación , Proteínas Portadoras/genética , Proteínas Portadoras/aislamiento & purificación , Cristalización , Cristalografía por Rayos X , Expresión Génica
20.
Am J Emerg Med ; 30(5): 732-6, 2012 Jun.
Artículo en Inglés | MEDLINE | ID: mdl-21641747

RESUMEN

OBJECTIVE: The aim of this study was to investigate if the electrocardiographic (ECG) abnormalities assessed early in the emergency department (ED) are associated with the in-hospital mortality of the patients with spontaneous subarachnoid hemorrhage (SAH). METHODS: We studied prospectively a cohort of 222 adult patients with spontaneous SAH in an ED. A 12-lead ECG was performed for these patients in the ED. The patients were stratified into nonsurvivors and survivors based on the in-hospital mortality. The clinical characteristics, heart rate, corrected QT interval (QTc) and 7 predefined morphologic abnormalities were compared between these 2 groups of patients. RESULTS: Compared with the survivors (n=178), the nonsurvivors (n=44) had significantly slower heart rate (75±23 vs 83±16, P=.018) and more prolonged QTc (492±58 vs 458±40, P=.001). There were significantly higher frequency of occurrence of ECG morphologic abnormalities (66% vs 37%, P=.001) and nonspecific ST- or T-wave changes (NSSTTCs; 32% vs 12%, P=.015) in the nonsurvivors compared with those in the survivors. Multiple logistic regression model identified QTc (odds ratio, 1.0; 95% confidence interval, 1.0-1.0; P=.005) and NSSTTC (odds ratio, 3.3; 95% confidence interval, 1.0-10.7; P=.047) as the significant ECG variables associated with in-hospital mortality. CONCLUSIONS: The occurrence of NSSTTC and prolonged QTc assessed early in the ED are independently associated with the in-hospital mortality in adult patients with spontaneous SAH.


Asunto(s)
Electrocardiografía/métodos , Frecuencia Cardíaca/fisiología , Hemorragia Subaracnoidea/diagnóstico , Distribución de Chi-Cuadrado , Servicio de Urgencia en Hospital , Femenino , Mortalidad Hospitalaria , Humanos , Modelos Logísticos , Masculino , Persona de Mediana Edad , Oportunidad Relativa , Valor Predictivo de las Pruebas , Pronóstico , Estudios Prospectivos , Hemorragia Subaracnoidea/mortalidad , Hemorragia Subaracnoidea/fisiopatología
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