RESUMEN

BACKGROUND: Stress significantly affects both the physical and emotional health of individuals, particularly students in health-related fields. Medical students in Brazil face unique challenges due to the demanding nature of their studies, especially during assessment periods, which heighten academic pressure. These pressures often lead to poor coping strategies and mental health concerns. It is crucial to understand the complex dynamics of stress within medical education to develop strategies that improve student well-being and promote a healthier academic environment. This study aims to investigate the intricate relationship between assessment periods and stress levels among medical students. It seeks to understand how academic demands and sociodemographic factors contribute to stress dynamics during these periods. METHODS: An online observational, longitudinal, and prospective study was conducted from February to October 2022. Medical students were recruited through snowball sampling and participated in surveys administered via Google Forms at two timepoints: before (T1) and during (T2) assessment periods. The surveys collected sociodemographic data and stress symptoms using Lipp's Inventory of Stress Symptoms for Adults (LSSI). RESULTS: The transition from T1 to T2 was defined by a rise in the prevalence of stress from 59.6 to 84.2% (p = 0.001) and a decline in symptom-free students from 40.4 to 15.8% (p = 0.001). There was a significant increase in exhaustion, from 12.3 to 31.6% (p = 0.0001). Higher stress levels were notably more prevalent among younger students (≤ 24 years), females, those from wealthier families, students without scholarships, those without prior degrees, and those in the clinical phase of their studies. However, non significant correlations were found between these sociodemographic and academic factors and the increase in stress. CONCLUSION: The findings highlight significant concerns regarding the mental health of medical students during assessment periods, marked by increased stress and exhaustion levels. These results emphasize the need for proactive interventions to manage stress effectively in medical education, considering its profound impact on students' well-being.

Asunto(s)

Estrés Psicológico , Estudiantes de Medicina , Humanos , Estudiantes de Medicina/psicología , Estudios Prospectivos , Estudios Longitudinales , Femenino , Masculino , Estrés Psicológico/epidemiología , Brasil , Adulto Joven , Adulto , Educación de Pregrado en Medicina , Adaptación Psicológica , Encuestas y Cuestionarios

RESUMEN

Lactoferrin (Lf) is an iron-binding glycoprotein and a component of many external secretions with a wide diversity of functions. Structural studies are important to understand the mechanisms employed by Lf to exert so varied functions. Here, we used guanidine hydrochloride and high hydrostatic pressure to cause perturbations in the structure of bovine Lf (bLf) in apo and holo (unsaturated and iron-saturated, respectively) forms, and analyzed conformational changes by intrinsic and extrinsic fluorescence spectroscopy. Our results showed that the iron binding promotes changes on tertiary structure of bLf and increases its structural stability. In addition, we evaluated the effects of bLf structural change on the kinetics of bLf internalization in Vero cells by confocal fluorescence microscopy, and observed that the holo form was faster than the apo form. This finding may indicate that structural changes promoted by iron binding may play a key role in the intracellular traffic of bLf. Altogether, our data improve the comprehension of bLf stability and uptake, adding knowledge to its potential use as a biopharmaceutical.

RESUMEN

Malignant brain tumors are among the most aggressive cancers with poor prognosis and no effective treatment. Recently, we reported the oncolytic potential of Zika virus infecting and destroying the human central nervous system (CNS) tumors in vitro and in immunodeficient mice model. However, translating this approach to humans requires pre-clinical trials in another immunocompetent animal model. Here, we analyzed the safety of Brazilian Zika virus (ZIKVBR) intrathecal injections in three dogs bearing spontaneous CNS tumors aiming an anti-tumoral therapy. We further assessed some aspects of the innate immune and inflammatory response that triggers the anti-tumoral response observed during the ZIKVBR administration in vivo and in vitro. For the first time, we showed that there were no negative clinical side effects following ZIKVBR CNS injections in dogs, confirming the safety of the procedure. Furthermore, the intrathecal ZIKVBR injections reduced tumor size in immunocompetent dogs bearing spontaneous intracranial tumors, improved their neurological clinical symptoms significantly, and extended their survival by inducing the destruction specifically of tumor cells, sparing normal neurons, and activating an immune response. These results open new perspectives for upcoming virotherapy using ZIKV to destroy and induce an anti-tumoral immune response in CNS tumors for which there are currently no effective treatments.

Asunto(s)

Neoplasias Encefálicas/complicaciones , Neoplasias Encefálicas/terapia , Viroterapia Oncolítica/métodos , Seguridad del Paciente , Carga Tumoral , Infección por el Virus Zika/complicaciones , Virus Zika/inmunología , Animales , Neoplasias Encefálicas/inmunología , Neoplasias Encefálicas/patología , Línea Celular Tumoral , Técnicas de Cocultivo , Citocinas/metabolismo , Modelos Animales de Enfermedad , Perros , Inmunidad , Inyecciones Espinales , Masculino , Células Madre Mesenquimatosas/metabolismo , Células Madre Mesenquimatosas/virología , Monocitos/inmunología , Monocitos/virología , Neuronas/metabolismo , Neuronas/virología , Resultado del Tratamiento

RESUMEN

Malignant brain tumors are among the most aggressive cancers with poor prognosis and no effective treatment. Recently, we reported the oncolytic potential of Zika virus infecting and destroying the human central nervous system (CNS) tumors in vitro and in immunodeficient mice model. However, translating this approach to humans requires pre-clinical trials in another immunocompetent animal model. Here, we analyzed the safety of Brazilian Zika virus (ZIKVBR) intrathecal injections in three dogs bearing spontaneous CNS tumors aiming an anti-tumoral therapy. We further assessed some aspects of the innate immune and inflammatory response that triggers the anti-tumoral response observed during the ZIKVBR administration in vivo and in vitro. For the first time, we showed that there were no negative clinical side effects following ZIKVBR CNS injections in dogs, confirming the safety of the procedure. Furthermore, the intrathecal ZIKVBR injections reduced tumor size in immunocompetent dogs bearing spontaneous intracranial tumors, improved their neurological clinical symptoms significantly, and extended their survival by inducing the destruction specifically of tumor cells, sparing normal neurons, and activating an immune response. These results open new perspectives for upcoming virotherapy using ZIKV to destroy and induce an anti-tumoral immune response in CNS tumors for which there are currently no effective treatments.

RESUMEN

It is estimated that more than 600,000 new cases of cancer will be reported in Brazil during the 2018-2019 biennium, especially with regard to prostate, breast, lung and colorectal cancers. Due to the high prevalence, incidence and mortality rates of these diseases, cancer campaigns such as 'Pink October' and 'Blue November' were strongly promoted in the past decade throughout the country to raise awareness of breast and prostate cancer, respectively. Nevertheless, whether the implementation of these campaigns has been proven efficient is still unknown. In the present study, we analysed the effectiveness of these campaigns on eliciting population online interest for cancer information. The Google Trends database was evaluated for the relative Internet search popularity for the terms 'breast cancer' and 'prostate cancer' from 2014 to 2019. Aside from some regional differences, we found that there was a high demand for 'breast cancer' and, to a lesser extent, 'prostate cancer' searches in a seasonal fashion (during October and November, respectively). Despite the worldwide high incidence of lung and colorectal cancers, searches including these keywords did not show increases in any specific period of the year, demonstrating the efficiency of the 'Pink October' and 'Blue November' campaigns in engaging the interest of the Brazilian population on the subject. These results allow us to infer that campaigns are effective in mobilising the attention of the Brazilian population with regard to breast and prostate cancers, but the practical aspects in reducing incidence and mortality should still be discussed.

RESUMEN

There are many factors which make canine cancer like cancer in humans. The occurrence of spontaneous mammary tumors in pet dogs, tumor genetics, molecular targets and exposure to the same environmental risk factors are among these factors. Therefore, the study of canine cancer can provide useful information to the oncology field. This study aimed to establish and characterize a panel of primary mixed cell cultures obtained from spontaneous canine mammary tumors. Eight established cell cultures obtained from one normal mammary gland, one complex adenoma, one mixed adenoma, two complex carcinomas and two mixed carcinomas were analyzed. The gene expression levels of classic molecular cancer players such as fibroblast growth factor receptor (FGFR) 2, breast cancer (BRCA) 1, BRCA2 and estrogen receptor (ESR) 1 were evaluated. For the first time, three orphan nuclear receptors, estrogen-related receptors (ERRs) α, ß and γ were studied in canine mammary cancer. The highest expression level of ERRα was observed in complex carcinoma-derived cell culture, while the highest levels of ERRß and γ were observed in cells derived from a mixed carcinoma. Meanwhile, complex carcinomas presented the highest levels of expression of ESR1, BRCA1 and FGFR2 among all samples. BRCA2 was found exclusively in complex adenoma. The transcription factor GATA3 had its highest levels in mixed carcinoma samples and its lowest levels in complex adenoma. Proliferation assays were also performed to evaluate the mixed cell cultures response to ER ligands, genistein and DES, both in normoxia and hypoxic conditions. Our results demonstrate that morphological and functional studies of primary mixed cell cultures derived from spontaneous canine mammary tumors are possible and provide valuable tool for the study of various stages of mammary cancer development.

Asunto(s)

Enfermedades de los Perros/metabolismo , Neoplasias Mamarias Animales/metabolismo , Animales , Proteína BRCA1/metabolismo , Proteína BRCA2/metabolismo , Ciclo Celular , Línea Celular Tumoral , Proliferación Celular , Células Cultivadas , Perros , Receptor alfa de Estrógeno/metabolismo , Femenino , Citometría de Flujo , Ploidias , Cultivo Primario de Células , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptor Tipo 2 de Factor de Crecimiento de Fibroblastos/metabolismo

RESUMEN

OBJECTIVE: Liver fibrosis results from the perpetuation of the normal wound healing response to several types of injury. Despite the wealth of knowledge regarding the involvement of intracellular and extracellular signaling pathways in liver fibrogenesis, information about the role of intercellular communication mediated by gap junctions is scarce. METHODS: In this study, liver fibrosis was chemically induced by carbon tetrachloride in mice lacking connexin32, the major liver gap junction constituent. The manifestation of liver fibrosis was evaluated based on a series of read-outs, including collagen morphometric and mRNA analysis, oxidative stress, apoptotic, proliferative and inflammatory markers. RESULTS: More pronounced liver damage and enhanced collagen deposition were observed in connexin32 knockout mice compared to wild-type animals in experimentally triggered induced liver fibrosis. No differences between both groups were noticed in apoptotic signaling nor in inflammation markers. However, connexin32 deficient mice displayed decreased catalase activity and increased malondialdehyde levels. CONCLUSION: These findings could suggest that connexin32-based signaling mediates tissue resistance against liver damage by the modulation of the antioxidant capacity. In turn, this could point to a role for connexin32 signaling as a therapeutic target in the treatment of liver fibrosis.

Asunto(s)

Tetracloruro de Carbono/toxicidad , Enfermedad Hepática Inducida por Sustancias y Drogas/metabolismo , Conexinas/deficiencia , Cirrosis Hepática Experimental/metabolismo , Hígado/efectos de los fármacos , Animales , Antioxidantes/metabolismo , Enfermedad Hepática Inducida por Sustancias y Drogas/etiología , Enfermedad Hepática Inducida por Sustancias y Drogas/patología , Colágeno/metabolismo , Conexinas/genética , Citocinas/inmunología , Ensayo de Inmunoadsorción Enzimática , Femenino , Hígado/inmunología , Hígado/metabolismo , Hígado/patología , Cirrosis Hepática Experimental/etiología , Cirrosis Hepática Experimental/patología , Pruebas de Función Hepática , Ratones Noqueados , Reacción en Cadena en Tiempo Real de la Polimerasa , Proteína beta1 de Unión Comunicante

RESUMEN

Pyrrolizidine alkaloids (PAs) are toxins that are exclusively biosynthesized by plants and are commonly present in foods and herbs. PAs are usually associated with poisoning events in livestock and human beings. The aim of the present study was to evaluate the behavioral and neurochemical effects of prenatal exposure to PA integerrimine N-oxide of rats in adulthood. Pregnant Wistar rats received integerrimine N-oxide from the butanolic residue of Senecio brasiliensis by gavage on gestational days 6-20 at doses of 3, 6 and 9 mg/kg. During adulthood of the offspring, the following behavioral tests were performed: open-field, plus-maze, forced swimming, catalepsy and stereotypy. Histological analyses and monoamine levels were measured. Male offspring from dams that were exposed to 9 mg/kg showed an increase in locomotion in the open-field test, an increased frequency of entries and time spent in open arms in elevated plus-maze test, as well as decreased swimming time. In the female offspring from dams that were exposed to 9 mg/kg, there was an increased time of climbing in forced swimming and intensity of stereotyped behavior. The histological study indicates an increase in the number of multinucleated cells in the liver (6 and 9 mg/kg). In neurotransmitter analysis, specifically in the striatum, we observed change in dopamine and serotonin levels in the middle dose. Thus, our results indicate that prenatal exposure to integerrimine N-oxide changed behavior in adulthood and neurotransmitter levels in the striatum. Our results agree with previous studies, which showed that integerrimine N-oxide impaired physical and neurobehavioral development in childhood that can persist until adulthood.

Asunto(s)

Antineoplásicos Fitogénicos/farmacología , Conducta Animal/efectos de los fármacos , Cuerpo Estriado/efectos de los fármacos , Neurotransmisores/metabolismo , Efectos Tardíos de la Exposición Prenatal , Alcaloides de Pirrolicidina/farmacología , Factores de Edad , Alanina Transaminasa/sangre , Animales , Antineoplásicos Fitogénicos/química , Aspartato Aminotransferasas/sangre , Proteínas Sanguíneas/metabolismo , Catalepsia/inducido químicamente , Cuerpo Estriado/metabolismo , Relación Dosis-Respuesta a Droga , Conducta Exploratoria/efectos de los fármacos , Femenino , Masculino , Embarazo , Efectos Tardíos de la Exposición Prenatal/inducido químicamente , Efectos Tardíos de la Exposición Prenatal/patología , Efectos Tardíos de la Exposición Prenatal/fisiopatología , Alcaloides de Pirrolicidina/química , Ratas , Ratas Wistar , Factores Sexuales , Conducta Estereotipada/efectos de los fármacos , Natación/psicología , gamma-Glutamiltransferasa/sangre

RESUMEN

Hebanthe paniculata roots (formerly Pfaffia paniculata and popularly known as Brazilian ginseng) show antineoplastic, chemopreventive, and antiproliferative properties. Functional properties of these roots and their extracts are usually attributed to the pfaffosidic fraction, which is composed mainly by pfaffosides A-F. However, the therapeutic potential of this fraction in cancer cells is not yet entirely understood. This study aimed to analyze the antitumoral effects of the purified pfaffosidic fraction or saponinic fraction on the human hepatocellular carcinoma HepG2 cell line. Cellular viability, proliferation, and apoptosis were evaluated, respectively, by MTT assay, BrdU incorporation, activated caspase-3 immunocytochemistry, and DNA fragmentation assay. Cell cycle was analyzed by flow cytometry and the cell cycle-related proteins were analyzed by quantitative PCR and Western blot. The cells exposed to pfaffosidic fraction had reduced viability and cellular growth, induced G2/M at 48 h or S at 72 h arrest, and increased sub-G1 cell population via cyclin E downregulation, p27(KIP1) overexpression, and caspase-3-induced apoptosis, without affecting the DNA integrity. Antitumoral effects of pfaffosidic fraction from H. paniculata in HepG2 cells originated by multimechanisms of action might be associated with cell cycle arrest in the S phase, by CDK2 and cyclin E downregulation and p27(KIP1) overexpression, besides induction of apoptosis through caspase-3 activation.

RESUMEN

Lung cancer is the leading cause of cancer-related mortality in both men and women throughout the world. This disease is strongly associated with tobacco smoking. The aim of this manuscript was to establish an in vitro model that mimics the chronic exposures of alveolar epithelial type II cells to the tobacco-specific nitrosamine carcinogen, NNK. Immortalized non-neoplastic alveolar epithelial cells type II, (E10 cells), from BALB/c mice were exposed to low concentration of NNK (100 pM) during 5, 10, 15, and 20 cycles of 48 h. NNK-transformed cells showed an increase of proliferation rate and motility. Moreover, these cells underwent epithelial-to-mesenchymal transition (EMT). Increased migratory capacity and EMT were correlated to the time of exposure to NNK. NNK-transformed cells were tested for their growth and metastatic capacity in vivo. Subcutaneous injection of cells exposed to NNK for 20 cycles (E10-NNK20 clone) into BALB/c mice led to the formation of subcutaneous tumors that arose after 40 ± 17 d in all animals, which died 95 ± 18 d after cell inoculation, with lymph nodes and lung metastasis. The morphological characteristics of tumors were compatible with metastatic undifferentiated carcinoma. Cells exposed to NNK for 5-10 cycles did not display metastatic capacity, while those exposed for 15 cycles displayed low capacity. Our results show that prolonged exposures to NNK led the cells to increasingly acquire malignant properties. The cellular model presented in this study is suitable for studying the molecular events involved in the different stages of malignant transformation.

Asunto(s)

Carcinógenos/toxicidad , Transformación Celular Neoplásica/efectos de los fármacos , Neoplasias Pulmonares/patología , Nicotiana , Nitrosaminas/toxicidad , Alveolos Pulmonares/patología , Animales , Western Blotting , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Modelos Animales de Enfermedad , Embrión de Mamíferos/citología , Embrión de Mamíferos/efectos de los fármacos , Embrión de Mamíferos/metabolismo , Transición Epitelial-Mesenquimal , Femenino , Técnica del Anticuerpo Fluorescente Indirecta , Técnicas para Inmunoenzimas , Técnicas In Vitro , Neoplasias Pulmonares/tratamiento farmacológico , Neoplasias Pulmonares/metabolismo , Ratones , Ratones Endogámicos BALB C , Metástasis de la Neoplasia , Alveolos Pulmonares/efectos de los fármacos , Alveolos Pulmonares/metabolismo , Cicatrización de Heridas

RESUMEN

The results of our previous study demonstrated that ptaquiloside, the main toxic agent found in Pteridium aquilinum, suppresses natural killer (NK) cell-mediated cytotoxicity. However, the ability of ptaquiloside to suppress the cytotoxicity of NK cells was prevented by selenium supplementation. NK cells play an important role in the innate immune response and have the ability to kill tumor cells. Therefore, we hypothesized that selenium may prevent the higher susceptibility to urethane-induced lung carcinogenesis that has been observed in mice treated with P. aquilinum. The immunosuppressive effects of ptaquiloside have been associated with a higher number of urethane-induced lung nodules in mice. Hence, we assessed the effects of P. aquilinum-induced immunosuppression on urethane-induced lung carcinogenesis in C57BL/6 mice that had been supplemented with selenium. For these experiments, mice were treated with both an aqueous extract of P. aquilinum (20 g/kg/day) and selenium (1.3 mg/kg) by gavage once daily for 14 days followed by a once-weekly intraperitoneal injection of urethane (1 g/kg) for 10 weeks that was accompanied by gavage 5 days a week. Lung adenomas in mice that had been treated with P. aquilinum plus urethane occurred with a frequency that was 44% higher than that in mice that had been treated with only urethane. In mice that had been supplemented with selenium and treated with P. aquilinum plus urethane, the occurrence of lung adenomas was reduced to 26%. These results suggest that selenium prevents the immunosuppressive effects of P. aquilinum on urethane-induced lung carcinogenesis.

Asunto(s)

Adenoma/prevención & control , Carcinógenos/farmacología , Suplementos Dietéticos , Indanos , Neoplasias Pulmonares/prevención & control , Pteridium/química , Selenio/farmacología , Sesquiterpenos , Uretano , Adenoma/inducido químicamente , Adenoma/patología , Animales , Femenino , Indanos/efectos adversos , Indanos/farmacología , Neoplasias Pulmonares/inducido químicamente , Neoplasias Pulmonares/patología , Ratones , Sesquiterpenos/efectos adversos , Sesquiterpenos/farmacología , Uretano/efectos adversos , Uretano/farmacología

RESUMEN

Connexins (Cxs) are proteins that form the communicating gap junctions, and reportedly have a role in carcinogenesis. Here, we evaluated the importance of Connexin43 (Cx43) in spontaneous and 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK)-induced lung carcinogenesis. Male wild-type (Cx43(+/+) ) and hemizygote (Cx43(+/-) ) CD1 × AJ F1 mice were injected with NNK or saline. After 60 weeks mice were euthanized; lung nodules were counted, measured, and fixed in formalin or snap frozen. Immunohistochemistry for Cx43 and Beta-catenin (ß-catenin) was performed and Cx43 mRNA expression was evaluated by real-time PCR. Cx43 deletion significantly increased the incidence and number of spontaneous nodules in the CD1 × AJ F1 mice and the number of gross lesions and the aggressiveness of lesions in NNK-treated mice. Cx43 mRNA increased significantly and was correlated with the aggressiveness of tumors, although lesions from Cx43(+/-) mice expressed less Cx43 RNAm than their counterparts. Lung parenchyma presented a Cx43 immunostaining pattern with points or plaques between cells. In hyperplasias and adenomas, Cx43 was found in the membrane and in cytoplasm. Malignant lesions presented increased Cx43 in cytoplasm and a few membrane spots of immunostaining. ß-catenin was weakly expressed in lung parenchyma. Though hyperplasias presented some cells with nuclear ß-catenin, NNK-induced tumors contained a higher number of this staining pattern. Also, no difference in ß-catenin occurred between both genotypes independently of the histological grade. In summary, our results indicate that Cx43 acts as a tumor suppressor gene in early lung tumorigenesis and loses this property in advanced carcinogenesis. Therefore, Cxs are better classified as conditional tumor suppressors.

Asunto(s)

Carcinógenos/toxicidad , Transformación Celular Neoplásica/patología , Conexina 43/fisiología , Neoplasias Pulmonares/etiología , Nitrosaminas/toxicidad , beta Catenina/metabolismo , Animales , Western Blotting , Susceptibilidad a Enfermedades , Femenino , Técnicas para Inmunoenzimas , Neoplasias Pulmonares/metabolismo , Neoplasias Pulmonares/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , ARN Mensajero/genética , Reacción en Cadena en Tiempo Real de la Polimerasa , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , beta Catenina/genética

RESUMEN

Resveratrol is a promising chemopreventive agent that mediates many cellular targets involved in cancer signaling pathways. p53 has been suggested to play a role in the anticancer properties of resveratrol. We investigated resveratrol-induced cytotoxicity in H1299 cells, which are non-small lung cancer cells that have a partial deletion of the gene that encodes the p53 protein. The results for H1299 cells were compared with those for three cell lines that constitutively express wild-type p53: breast cancer MCF-7, adenocarcinomic alveolar basal epithelia A549 and non-small lung cancer H460. Cell viability assays revealed that resveratrol reduced the viability of all four of these cell lines in a dose- and time-dependent manner. MCF-7, A549 and H460 cells were more sensitive to resveratrol than were H1299 cells when exposed to the drug for 24 h at concentrations above 100 µM. Resveratrol also increased the p53 protein levels in MCF-7 cells without altering the p53 mRNA levels, suggesting a post-translational modulation of the protein. The resveratrol-induced cytotoxicity in these cells was partially mediated by p53 and involved the activation of caspases 9 and 7 and the cleavage of PARP. In H1299 cells, resveratrol-induced cytotoxicity was less pronounced and (in contrast to MCF-7 cells) cell death was not accompanied by caspase activation. These findings are consistent with the observation that MCF-7 cells were positively labeled by TUNEL following exposure to 100 µM resveratrol whereas H1299 cells under similar conditions were not labeled by TUNEL. The transient transfection of a wild-type p53-GFP gene caused H1299 cells to become more responsive to the pro-apoptotic properties of resveratrol, similarly to findings in the p53-positive MCF-7 cells. Our results suggest a possible therapeutic strategy based on the use of resveratrol for the treatment of tumors that are typically unresponsive to conventional therapies because of the loss of normal p53 function.

Asunto(s)

Antineoplásicos Fitogénicos/farmacología , Apoptosis/efectos de los fármacos , Apoptosis/genética , Estilbenos/farmacología , Proteína p53 Supresora de Tumor/genética , Antineoplásicos Fitogénicos/toxicidad , Neoplasias de la Mama/genética , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Supervivencia Celular/genética , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Neoplasias Pulmonares/genética , Células MCF-7 , Masculino , Resveratrol , Estilbenos/toxicidad , Activación Transcripcional/efectos de los fármacos , Transfección , Proteína p53 Supresora de Tumor/metabolismo

RESUMEN

Over 50% of all human cancers lose p53 function. To evaluate the role of aggregation in cancer, we asked whether wild-type (WT) p53 and the hot-spot mutant R248Q could aggregate as amyloids under physiological conditions and whether the mutant could seed aggregation of the wild-type form. The central domains (p53C) of both constructs aggregated into a mixture of oligomers and fibrils. R248Q had a greater tendency to aggregate than WT p53. Full-length p53 aggregated into amyloid-like species that bound thioflavin T. The amyloid nature of the aggregates was demonstrated using x-ray diffraction, electron microscopy, FTIR, dynamic light scattering, cell viabilility assay, and anti-amyloid immunoassay. The x-ray diffraction pattern of the fibrillar aggregates was consistent with the typical conformation of cross ß-sheet amyloid fibers with reflexions of 4.7 Å and 10 Å. A seed of R248Q p53C amyloid oligomers and fibrils accelerated the aggregation of WT p53C, a behavior typical of a prion. The R248Q mutant co-localized with amyloid-like species in a breast cancer sample, which further supported its prion-like effect. A tumor cell line containing mutant p53 also revealed massive aggregation of p53 in the nucleus. We conclude that aggregation of p53 into a mixture of oligomers and fibrils sequestrates the native protein into an inactive conformation that is typical of a prionoid. This prion-like behavior of oncogenic p53 mutants provides an explanation for the negative dominance effect and may serve as a potential target for cancer therapy.

Asunto(s)

Amiloide/química , Mutación Missense , Neoplasias/química , Priones , Multimerización de Proteína , Proteína p53 Supresora de Tumor/química , Sustitución de Aminoácidos , Amiloide/genética , Amiloide/metabolismo , Benzotiazoles , Humanos , Neoplasias/genética , Neoplasias/metabolismo , Neoplasias/terapia , Estructura Cuaternaria de Proteína , Estructura Secundaria de Proteína , Tiazoles/química , Proteína p53 Supresora de Tumor/genética , Proteína p53 Supresora de Tumor/metabolismo , Difracción de Rayos X

RESUMEN

Primary hepatic neoplasms (PHN) account for 0.6 to 1.5% of canine tumours and constitute a major portion of chronic liver disease. The canine hepatic neoplasms can be classified as hepatocellular adenoma/carcinoma (HA/HCC), cholangiocellular adenoma/carcinoma (CCA/CCC), neuroendocrine carcinoma of tumors of mesenchymal origin, vascular or hematopoietic. Thus, because of its importance in veterinary oncology and hepatology, we performed a retrospective study of primary canine liver cancer in the Department of Pathology, School of Veterinary Medicine and Animal Science, University of Sao Paulo, between 1999 and 2012. Was evaluated 6886 histopathological analysis, 106 (1.5%) were related to canine liver diseases, in which the tumors accounted for 27 cases (25.47%). Liver neoplasms, were classified as HA (2 cases -7.4%); CCA (5 cases - 18.5%); HCC (5 cases - 18.5%); CCC (6 cases - 22.2%) and 9 tumors of mesenchymal, hematopoietic or vascular (33.3%). Epidemiological data showed similarities to previously reported data, predominantly CCC, HCC and CCA in older animals and females.(AU)

Asunto(s)

Animales , Femenino , Perros , Enfermedades de los Perros/epidemiología , Neoplasias Hepáticas/epidemiología , Estudios Retrospectivos , Hígado

RESUMEN

Primary hepatic neoplasms (PHN) account for 0.6 to 1.5% of canine tumours and constitute a major portion of chronic liver disease. The canine hepatic neoplasms can be classified as hepatocellular adenoma/carcinoma (HA/HCC), cholangiocellular adenoma/carcinoma (CCA/CCC), neuroendocrine carcinoma of tumors of mesenchymal origin, vascular or hematopoietic. Thus, because of its importance in veterinary oncology and hepatology, we performed a retrospective study of primary canine liver cancer in the Department of Pathology, School of Veterinary Medicine and Animal Science, University of Sao Paulo, between 1999 and 2012. Was evaluated 6886 histopathological analysis, 106 (1.5%) were related to canine liver diseases, in which the tumors accounted for 27 cases (25.47%). Liver neoplasms, were classified as HA (2 cases -7.4%); CCA (5 cases - 18.5%); HCC (5 cases - 18.5%); CCC (6 cases - 22.2%) and 9 tumors of mesenchymal, hematopoietic or vascular (33.3%). Epidemiological data showed similarities to previously reported data, predominantly CCC, HCC and CCA in older animals and females.

RESUMEN

GJA1 gene (Connexin43, also known as Cx43) is the most abundant gap junction protein isoform in animal cells and is associated with bone development in embryos. The objective of the present work was to evaluate in vivo osteal development in GJA1-deficient fetal mice through determination of the histological and molecular alterations induced by partial or total deletion of the GJA1 gene. Heterozygous C57BL/6 mice (HT) harboring a null mutation of the GJA1 gene were mated, and pregnant females were submitted to euthanasia and Caesarean section from 12.5 to 19.5 days post coitum (dpc). HT (GJA1(+/-)) and homozygous (GJA1(-/- )) knockout (KO) mutants and wild-type (WT) fetuses were identified by polymerase chain reaction (PCR), and development curves were constructed on the basis of fetus weight and crown-rump length. Histopathological, histochemical, and real-time PCR analyses were performed in order to assess the expression of markers associated with bone development, namely, osteocalcin, osteopontin, alkaline phosphatase, RUNX2, GJA1, GJC1 (Cx45), and GJA3 (Cx46). HT and KO fetuses exhibited delays in the differentiation of osteoblasts and, consequently, in bone development in comparison with the WT group. Additionally, less deposition of mineralized and osteoid matrix was observed in GJA1-deficient fetuses. Bone development in KO fetuses was delayed through the moment of birth, but in HT animals the delay only extended until 17.5 dpc, following which development was normalized. The expression of genes coding for osteocalcin, osteopontin, alkaline phosphatise, and RUNX2 were also delayed in GJA1-deficient fetuses. Animals that exhibited a lower expression of GJA1 presented delayed expression of the GJC1 and GJA3 genes and their corresponding protein products in the bone tissue. The results of the present study contribute to our understanding of the function of GJA1 during bone development and suggest that GJC1 could play a role in restoring intercellular communication in GJA1-deficient mice.

Asunto(s)

Conexina 43/deficiencia , Osteoblastos/citología , Osteogénesis/fisiología , Análisis de Varianza , Animales , Biomarcadores/metabolismo , Diferenciación Celular/genética , Conexina 43/genética , Conexina 43/metabolismo , Subunidad alfa 1 del Factor de Unión al Sitio Principal/biosíntesis , Subunidad alfa 1 del Factor de Unión al Sitio Principal/genética , Largo Cráneo-Cadera , Femenino , Peso Fetal/genética , Uniones Comunicantes/genética , Eliminación de Gen , Histocitoquímica , Ratones , Ratones Noqueados , Osteoblastos/metabolismo , Osteogénesis/genética , Embarazo , Costillas/química

RESUMEN

Swainsonine is a natural α-mannosidase inhibitor found in numerous poisonous plants, such as Astragalus lentiginosus. Its mechanism of action is through the inhibition of Golgi α-mannosidase II activity in the N-glycan biosynthesis pathway. As a result, swainsonine inhibits the production of complex ß1,6-branched N-linked glycans, which are related to the malignant phenotype of tumor cells. In this study, we investigated whether treatment with swainsonine affects the sensitivity of Ehrlich ascites carcinoma (EAC) cells to cisplatin. To this end, male C57BL/6 mice were treated with swainsonine (SW--0.5 mg/kg, i.p., twice-daily for ten days) and/or cisplatin (Cis--0.25 mg/kg, i.p., every other day for a total of five applications) two days after transplantation with EAC cells. The results showed a greater reduction in the ascites volume in mice from the CisSW group (63.5%) than in mice from the Cis group (45.7%), an elevated induction of apoptosis by CisSW treatment when compared to Cis alone, as demonstrated by higher percentage of cells in the subG1 phase in that group (p<0.0001 Kruskal-Wallis, p<0.0001 control vs. CisSW, p<0.001 Co vs. Cis post-test Dunn), and an increase in the median survival from 12.5 days observed in the control group to 27 days in the CisSW group, which corresponds to a 116% survival increase (p=0.0022 Co vs. CisSW Log-rank test). In addition, the mice from the Cis group had a median survival of only 15 days, an increase of just 20% compared to controls. Our results indicate that swainsonine increases the sensitivity of EAC cells to cisplatin.

Asunto(s)

Antineoplásicos Fitogénicos/uso terapéutico , Carcinoma de Ehrlich/tratamiento farmacológico , Cisplatino/uso terapéutico , Swainsonina/uso terapéutico , Animales , Antineoplásicos Fitogénicos/farmacología , Recuento de Células Sanguíneas , Ciclo Celular/efectos de los fármacos , Cisplatino/farmacología , Sinergismo Farmacológico , Quimioterapia Combinada , Masculino , Ratones , Ratones Endogámicos C57BL , Análisis de Supervivencia , Swainsonina/farmacología , Aumento de Peso/efectos de los fármacos

RESUMEN

Mast cell tumor (MCT) is one of the most prevalent neoplasms that affect skin and soft tissue in dogs. Because mast cell tumors present a great variety of clinical appearance and behavior, their treatment becomes a challenge. Trichostatin A (TSA), an antifungal antibiotic, has shown inhibitory effects on the proliferation and induction of apoptosis in various types of cancer cells. In order to evaluate the potential of trichostatin A as a therapeutic drug, cells of grade 3 MCT were cultured and treated with concentrations of 1 nM to 400 nM of TSA. MTT assay and trypan blue exclusion assays were performed to estimate cell growth and cell viability, and cell cycle analysis was evaluated. TSA treatment showed a reduction in numbers of viable cells and an increase of cell death by apoptosis. The cell cycle analysis showed an increase of hypodiploid cells and a reduction of G0/G1 and G2/M -phases. According to these results, trichostatin A may be an interesting potential chemotherapeutic agent for the treatment of canine MCT.

Asunto(s)

Inhibidores de Histona Desacetilasas/farmacología , Ácidos Hidroxámicos/farmacología , Sarcoma de Mastocitos/veterinaria , Acetilación , Animales , Apoptosis/efectos de los fármacos , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Supervivencia Celular/efectos de los fármacos , Perros , Femenino , Inhibidores de Histona Desacetilasas/administración & dosificación , Histonas/metabolismo , Ácidos Hidroxámicos/administración & dosificación , Sarcoma de Mastocitos/tratamiento farmacológico , Sarcoma de Mastocitos/patología , Ratones , Ratones Desnudos , Sales de Tetrazolio/química , Tiazoles/química , Azul de Tripano/química

RESUMEN

Gap junction channels, formed by connexins (Cx), are involved in the maintenance of tissue homeostasis, cell growth, differentiation, and development. Several studies have shown that Cx43 is involved in the control of wound healing in dermal tissue. However, it remains unknown whether Cx43 plays a role in the control of liver fibrogenesis. Our study investigated the roles of Cx43 heterologous deletion on carbon tetrachloride (CCl(4))-induced hepatic fibrosis in mice. We administered CCl(4) to both Cx43-deficient (Cx43(+/-)) and wild-type mice and examined hepatocellular injury and collagen deposition by histological and ultrastructural analyses. Serum biochemical analysis was performed to quantify liver injury. Hepatocyte proliferation was analyzed immunohistochemically. Protein and messenger RNA (mRNA) expression of liver connexins were evaluated using immunohistochemistry as well as immunoblotting analysis and quantitative real-time PCR. We demonstrated that Cx43(+/-) mice developed excessive liver fibrosis compared with wild-type mice after CCl(4) -induced chronic hepatic injury, with thick and irregular collagen fibers. Histopathological evaluation showed that Cx43(+/-) mice present less necroinflammatory lesions in liver parenchyma and consequent reduction of serum aminotransferase activity. Hepatocyte cell proliferation was reduced in Cx43(+/-) mice. There was no difference in Cx32 and Cx26 protein or mRNA expression in fibrotic mice. Protein expression of Cx43 increased in CCl(4)-treated mice, although with aberrant protein location on cytoplasm of perisinusoidal cells. Our results demonstrate that Cx43 plays an important role in the control and regulation of hepatic fibrogenesis.

Asunto(s)

Quimiocina CCL4/toxicidad , Conexina 43/deficiencia , Cirrosis Hepática/inducido químicamente , Hígado/patología , Patología Molecular , Animales , Ratones