RESUMEN
BACKGROUND: The transitional period between the oncosphere and the cysticercus of Taenia solium is the postoncospheral (PO) form, which has not yet been completely characterized. The aim of this work was to standardize a method to obtain T. solium PO forms by in vitro cultivation. We studied the morphology of the PO form and compared the expression of antigenic proteins among the PO form, oncosphere, and cysticerci stages. METHODOLOGY/PRINCIPAL FINDINGS: T. solium activated oncospheres were co-cultured with ten cell lines to obtain PO forms, which we studied at three stages of development--days 15, 30, and 60. A high percentage (32%) of PO forms was obtained using HCT-8 cells in comparison to the other cell lines. The morphology was observed by bright field, scanning, and transmission electron microscopy. Morphology of the PO form changed over time, with the six hooks commonly seen in the oncosphere stage disappearing in the PO forms, and vesicles and microtriches observed in the tegument. The PO forms grew as they aged, reaching a diameter of 2.5 mm at 60 days of culture. 15-30 day PO forms developed into mature cysticerci when inoculated into rats. Antigenic proteins expressed in the PO forms are also expressed by the oncosphere and cysticerci stages, with more cysticerci antigenic proteins expressed as the PO forms ages. CONCLUSIONS/SIGNIFICANCE: This is the first report of an in vitro production method of T. solium PO forms. The changes observed in protein expression may be useful in identifying new targets for vaccine development. In vitro culture of PO form will aid in understanding the host-parasite relationship, since the structural changes of the developing PO forms may reflect the parasite's immunoprotective mechanisms. A wider application of this method could significantly reduce the use of animals, and thus the costs and time required for further experimental investigations.
Asunto(s)
Antígenos Helmínticos/análisis , Taenia solium/anatomía & histología , Taenia solium/crecimiento & desarrollo , Animales , Western Blotting , Línea Celular , Técnicas de Cocultivo , Perfilación de la Expresión Génica , Humanos , Microscopía , Taenia solium/genéticaRESUMEN
BACKGROUND: Diminutive (≤ 5 mm) colorectal polyps are common, and overwhelmingly benign. Routinely, after polypectomy, they are examined pathologically to determine the surveillance intervals. Advances in equipment and techniques, such as narrow-band imaging (NBI) colonoscopy, now permit reliable real-time optical diagnosis. METHODS: We conducted a randomised single-masked study involving three institutions to determine whether optical diagnosis of diminutive colorectal polyps meets clinical practice standards and reduces the need for histopathology. We randomly assigned eligible patients undergoing routine high-definition colonoscopy to optical diagnosis using near focus versus standard view, using computer-generated block sequence. By validated criteria, we rendered an optical diagnosis and a confidence level (high vs low) for all polyps, using NBI. Our primary endpoint was the number of accurate high-confidence optical diagnoses compared with central blinded pathology in the two groups. We analysed data using intention to treat. FINDINGS: We enrolled 558 subjects, and randomly assigned 281 to near focus and 277 to standard view optical diagnosis. We detected 1309 predominantly diminutive (74.5%) and neoplastic (60.0%) polyps. Endoscopists were significantly more likely, OR 2.2 (95% CI 1.6 to 3.0, p<0.0001), to make a high-confidence optical diagnosis with near focus (85.1%) than standard (72.6%) view. High-confidence diagnoses had 96.4% and 92.0% negative predictive value, respectively. Of all polyps, 75.3% (95% CI71.3% to 78.9%) had a high-confidence accurate prediction using near focus, compared with 63.1% (95% CI 58.5% to 67.6%) using standard view. Optical versus histopathological diagnosis showed excellent agreement between the surveillance intervals, 93.5% in near focus and 92.2% in standard view. The median diagnosis time was 14 s. CONCLUSIONS: Real-time optical diagnosis using NBI colonoscopy may replace the pathology diagnosis for the majority of diminutive colorectal polyps. Using colonoscopy with near focus view increases the confidence level of the optical diagnosis. Optical diagnosis would be a paradigm shift in clinical practice of colonoscopy for colorectal cancer screening. TRIAL REGISTRATION NUMBER: ClinicalTrials.gov Identifier: NCT01288833.
Asunto(s)
Pólipos del Colon/diagnóstico , Colonoscopía/métodos , Neoplasias Colorrectales/diagnóstico , Imagen de Banda Estrecha/métodos , Diagnóstico Diferencial , Estudios de Seguimiento , Humanos , Valor Predictivo de las Pruebas , Estudios Retrospectivos , Método Simple CiegoRESUMEN
The specific mechanisms underlying Taenia solium oncosphere adherence and penetration in the host have not been studied previously. We developed an in vitro adhesion model assay to evaluate the mechanisms of T. solium oncosphere adherence to the host cells. The following substrates were used: porcine intestinal mucosal scrapings (PIMS), porcine small intestinal mucosal explants (PSIME), Chinese hamster ovary cells (CHO cells), epithelial cells from ileocecal colorectal adenocarcinoma (HCT-8 cells), and epithelial cells from colorectal adenocarcinoma (Caco-2 cells). CHO cells were used to compare oncosphere adherence to fixed and viable cells, to determine the optimum time of oncosphere incubation, to determine the role of sera and monolayer cell maturation, and to determine the effect of temperature on oncosphere adherence. Light microscopy, scanning microscopy, and transmission microscopy were used to observe morphological characteristics of adhered oncospheres. This study showed in vitro adherence of activated T. solium oncospheres to PIMS, PSIME, monolayer CHO cells, Caco-2 cells, and HCT-8 cells. The reproducibility of T. solium oncosphere adherence was most easily measured with CHO cells. Adherence was enhanced by serum-binding medium with >5% fetal bovine serum, which resulted in a significantly greater number of oncospheres adhering than the number adhering when serum at a concentration less than 2.5% was used (P < 0.05). Oncosphere adherence decreased with incubation of cells at 4 degrees C compared with the adherence at 37 degrees C. Our studies also demonstrated that T. solium oncospheres attach to cells with elongated microvillus processes and that the oncospheres expel external secretory vesicles that have the same oncosphere processes.
Asunto(s)
Adhesión Celular/fisiología , Células Epiteliales/parasitología , Membrana Mucosa/parasitología , Taenia solium/fisiología , Animales , Células CHO , Células CACO-2 , Cricetinae , Cricetulus , Humanos , Microscopía Electrónica de Rastreo , Microscopía Electrónica de Transmisión , Reproducibilidad de los Resultados , Suero , Porcinos , TemperaturaRESUMEN
Mallory bodies (MBs) are characteristic of several liver disorders, and consist primarily of keratins with transglutaminase-generated keratin crosslinks. We tested the effect of the transglutaminase-2 (TG2) inhibitor KCC009 on MB formation in a mouse model fed 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC). KCC009 decreased DDC-induced liver enlargement without affecting MB formation or extent of liver injury. TG2 protein and activity increased after DDC feeding and localized within and outside hepatocytes. KCC009 inhibited DDC-induced hepatomegaly by affecting hepatocyte cell size rather than proliferation. Hence, TG2 is a potential mediator of injury-induced hepatomegaly via modulation of hepatocyte hypertrophy, and KCC009-mediated TG2 inhibition does not affect mouse MB formation.
Asunto(s)
Inhibidores Enzimáticos/administración & dosificación , Proteínas de Unión al GTP/antagonistas & inhibidores , Cuerpos de Inclusión/efectos de los fármacos , Isoxazoles/administración & dosificación , Hepatopatías/tratamiento farmacológico , Proteínas/metabolismo , Transglutaminasas/antagonistas & inhibidores , Animales , Tamaño de la Célula/efectos de los fármacos , Enfermedad Hepática Inducida por Sustancias y Drogas , Dicarbetoxidihidrocolidina/administración & dosificación , Dicarbetoxidihidrocolidina/toxicidad , Proteínas de Unión al GTP/metabolismo , Hepatomegalia/inducido químicamente , Hepatomegalia/tratamiento farmacológico , Hepatomegalia/enzimología , Hepatomegalia/patología , Humanos , Cuerpos de Inclusión/metabolismo , Cuerpos de Inclusión/patología , Queratinas/metabolismo , Hepatopatías/enzimología , Hepatopatías/patología , Ratones , Ratones Endogámicos C3H , Proteína Glutamina Gamma Glutamiltransferasa 2 , Transglutaminasas/metabolismoRESUMEN
A 43-year-old woman underwent mitral valve replacement for severe mitral regurgitation nine years after orthotopic heart transplant. Histopathology showed chronic rejection of the mitral valve with lymphocytic infiltrates. The patient is well at one year follow-up. This report describes an identified case of chronic mitral valve rejection requiring valve replacement.
Asunto(s)
Implantación de Prótesis de Válvulas Cardíacas , Insuficiencia de la Válvula Mitral/cirugía , Adulto , Femenino , Rechazo de Injerto , Trasplante de Corazón , Humanos , Insuficiencia de la Válvula Mitral/etiología , Insuficiencia de la Válvula Mitral/patología , Complicaciones PosoperatoriasRESUMEN
Whipple disease (WD) is a systemic disorder caused by the bacterium Tropheryma whipplei. Since the recognition of a bacterial etiology in 1961, many attempts have been made to cultivate this bacterium in vitro. It was eventually isolated, in 2000, from an infected heart valve, in coculture with human fibroblasts. Here we report the isolation of 2 new strains of T. whipplei from cerebrospinal fluid (CSF) of 2 patients with intestinal WD but no neurological signs or symptoms. One culture-positive specimen was obtained before treatment; the other was obtained 12 months after discontinuation of therapy, at a time of intestinal remission. In both cases, 15 passages of the cultures were completed over 17 months. Bacterial growth was measured by quantitative polymerase chain reaction, which suggested a generation time of 4 days. Staining with YO-PRO nucleic-acid dye showed characteristic rod-shaped bacteria arranged in chains. Fluorescent in situ hybridization with a T. whipplei-specific oligonucleotide probe, a broad-range bacterial probe, and a nonspecific nucleic-acid stain indicated that all visible bacteria were T. whipplei. Scanning electron microscopy and transmission electron microscopy showed both intracellular and extracellular bacteria. This first isolation of T. whipplei from CSF provides clear evidence of viable bacteria in the central nervous system in individuals with WD, even after prolonged antibiotic therapy.