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Entamoeba histolytica is the causative agent of amebiasis, an infectious disease targeting the intestine and the liver in humans. Two types of intestinal infection are caused by this parasite: silent infection, which occurs in the majority of cases, and invasive disease, which affects 10% of infected persons. To understand the intestinal pathogenic process, several in vitro models, such as cell cultures, human tissue explants or human intestine xenografts in mice, have been employed. Nevertheless, our knowledge on the early steps of amebic intestinal infection and the molecules involved during human-parasite interaction is scarce, in part due to limitations in the experimental settings. In the present work, we took advantage of tissue engineering approaches to build a three-dimensional (3D)-intestinal model that is able to replicate the general characteristics of the human colon. This system consists of an epithelial layer that develops tight and adherens junctions, a mucus layer and a lamina propria-like compartment made up of collagen containing macrophages and fibroblast. By means of microscopy imaging, omics assays and the evaluation of immune responses, we show a very dynamic interaction between E. histolytica and the 3D-intestinal model. Our data highlight the importance of several virulence markers occurring in patients or in experimental models, but they also demonstrate the involvement of under described molecules and regulatory factors in the amoebic invasive process.
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Amebíase/parasitologia , Entamoeba histolytica/patogenicidade , Intestinos/microbiologia , Intestinos/patologia , Modelos Anatômicos , Amebíase/imunologia , Disenteria Amebiana/patologia , Entamoeba histolytica/imunologia , Interações Hospedeiro-Parasita , Humanos , Inflamação , Microscopia Confocal , VirulênciaRESUMO
Increased TNF-α levels have been associated with adverse pregnancy outcomes. Lipopolysaccharide (LPS), 1,1,1-trichloro-2,2-bis-(chlorophenyl)ethane (DDT), 1,1-bis-(chlorophenyl)-2,2-dichloroethene (DDE), and 1,1-dichloro-2,2-bis(chlorophenyl)ethane (DDD) induce TNF-α release in peripheral blood mononuclear cells (PBMC). Conversely, progesterone (P4) inhibits TNF-α secretion. Pregnant women in malaria endemic areas may be co-exposure to these compounds. Thus, this study was to investigate the synergistic effect of LPS and these pesticides in PBMC and to assess P4 influence on this synergy. Cultured PBMC were exposed to each pesticide in the presence of LPS, P4, or their combination. TNF-α was measured by ELISA. All pesticides enhanced TNF-α synthesis in PBMC. Co-exposure with LPS synergizes TNF-α production, which is blocked by progesterone. These results indicate that these organochlorines act synergistically with LPS to induce TNF-α secretion in PBMC. This effect is blocked by P4.
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DDT , Leucócitos Mononucleares/metabolismo , Lipopolissacarídeos , Progesterona/farmacologia , Fator de Necrose Tumoral alfa/metabolismo , Adulto , DDT/agonistas , DDT/farmacocinética , DDT/toxicidade , Feminino , Humanos , Leucócitos Mononucleares/patologia , Lipopolissacarídeos/agonistas , Lipopolissacarídeos/toxicidade , Malária/epidemiologia , Malária/metabolismo , Malária/patologia , Gravidez , Complicações Parasitárias na Gravidez/epidemiologia , Complicações Parasitárias na Gravidez/metabolismo , Complicações Parasitárias na Gravidez/patologiaRESUMO
Through research carried out in the last 25 years about the breast cancer etiology, it has been possible to estimate that less than 10 % of patients who are diagnosed with the condition are carriers of some germline or somatic mutation. The clinical reports of breast cancer patients with healthy twins and the development of disease in women without high penetrance mutations detected, warn the participation more factors in the transformation process. The high incidence of mammary adenocarcinoma in the modern woman and the urgent need for new methods of prevention and early detection have demanded more information about the role that environment and lifestyle have on the transformation of mammary gland epithelial cells. Obesity, alcoholism and smoking are factors that have shown a close correlation with the risk of developing breast cancer. And although these conditions affect different cell regulation levels, the study of its effects in the mechanisms of transcriptional and epigenetic regulation is considered critical for a better understanding of the loss of identity of epithelial cells during carcinogenesis of this tissue. The main objective of this review was to establish the importance of changes occurring to transcriptional level in the mammary gland as a consequence of acute or chronic exposure to harmful products such as obesity-causing foods, ethanol and cigarette smoke components. At analyze the main studies related to topic, it has concluded that the understanding of effects caused by the lifestyle factors in performance of the transcriptional mechanisms that determine gene expression of the mammary gland epithelial cells, may help explain the development of this disease in women without genetic propensity and different phenotypic manifestations of this cancer type.
A través de la investigación realizada en los últimos 25 años en torno a la etiología del cáncer de mama, ha sido posible estimar que menos del 10 % de las pacientes que son diagnosticadas con la enfermedad son portadoras de alguna mutación de línea germinal o somática. Los informes clínicos de pacientes de cáncer de mama con gemelas saludables y el desarrollo de la enfermedad en las mujeres sin mutaciones de alta penetrancia detectadas, advierten la participación de otros factores en el proceso de transformación. La alta incidencia de adenocarcinoma de mama en la mujer moderna y la necesidad urgente de nuevos métodos para la prevención y detección temprana han exigido una mayor información en relación al papel que el medio ambiente y el estilo de vida tienen en la transformación de las células epiteliales de la glándula mamaria. La obesidad, el alcoholismo y el tabaquismo son factores que han demostrado una estrecha correlación con el riesgo de desarrollar cáncer de mama. Y aunque estas condiciones pueden afectar distintos niveles de regulación celular, el estudio de sus efectos en los mecanismos de regulación transcripcional y epigenética, es considerado fundamental para un mayor entendimiento de la pérdida de identidad de las células epiteliales durante la carcinogénesis de este tejido. El objetivo principal de esta revisión fue establecer la importancia de los cambios que ocurren a nivel transcripcional en la glándula mamaria, como consecuencia de una exposición aguda o crónica a productos nocivos, tales como los alimentos que favorecen la obesidad, el etanol y los componentes del humo del cigarro. Al analizar los principales estudios relacionados con el tema, se ha llegado a la conclusión de que la comprensión de los efectos causados por los factores de estilo de vida sobre el desempeño de los mecanismos de regulación transcripcional responsables de la expresión génica de las células epiteliales de la glándula mamaria, pueden ayudar a explicar el desarrollo de esta enfermedad en las mujeres que no son genéticamente propensas así como las diferentes manifestaciones fenotípicas de este tipo de cáncer.
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Introduction: Breast cancer (BC) is the leading cause of death among women, primarily due to its potential for metastasis. As BC progresses, the extracellular matrix (ECM) produces more type-I collagen, resulting in increased stiffness. This alteration influences cellular behaviors such as migration, invasion, and metastasis. Specifically, cancer cells undergo changes in gene expression that initially promote an epithelial-to-mesenchymal transition (EMT) and subsequently, a transition from a mesenchymal to an amoeboid (MAT) migration mode. In this way, cancer cells can migrate more easily through the stiffer microenvironment. Despite their importance, understanding MATs remains challenging due to the difficulty of replicating in vitro the conditions for cell migration that are observed in vivo. Methods: To address this challenge, we developed a three-dimensional (3D) growth system that replicates the different matrix properties observed during the progression of a breast tumor. We used this model to study the migration and invasion of the Triple-Negative BC (TNBC) cell line MDA-MB-231, which is particularly subject to metastasis. Results: Our results indicate that denser collagen matrices present a reduction in porosity, collagen fiber size, and collagen fiber orientation, which are associated with the transition of cells to a rounder morphology with bleb-like protrusions. We quantified how this transition is associated with a more persistent migration, an enhanced invasion capacity, and a reduced secretion of matrix metalloproteinases. Discussion: Our findings suggest that the proposed 3D growth conditions (especially those with high collagen concentrations) mimic key features of MATs, providing a new platform to study the physiology of migratory transitions and their role in BC progression.
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Introduction: Breast cancer (BC) is the leading cause of cancer-related deaths among women, with triple-negative breast cancer (TNBC) representing one of the most aggressive and treatment-resistant subtypes. In this study, we aimed to evaluate the antitumor potential of C14 and P8 molecules in both TNBC and radioresistant TNBC cells. These compounds were chosen for their ability to stabilize the complex formed by the overactivated form of K-Ras4BG13D and its membrane transporter (PDE6δ). Methods: The antitumor potential of C14 and P8 was assessed using TNBC cell lines, MDA-MB-231, and the radioresistant derivative MDA-MB-231RR, both carrying the K-Ras4B> G13D mutation. We investigated the compounds' effects on K-Ras signaling pathways, cell viability, and tumor growth in vivo. Results: Western blotting analysis determined the negative impact of C14 and P8 on the activation of mutant K-Ras signaling pathways in MDA-MB-231 and MDA-MB-231RR cells. Proliferation assays demonstrated their efficacy as cytotoxic agents against K-RasG13D mutant cancer cells and in inducing apoptosis. Clonogenic assays proven their ability to inhibit TNBC and radioresistant TNBC cell clonogenicity. In In vivo studies, C14 and P8 inhibited tumor growth and reduced proliferation, angiogenesis, and cell cycle progression markers. Discussion: These findings suggest that C14 and P8 could serve as promising adjuvant treatments for TNBC, particularly for non-responders to standard therapies. By targeting overactivated K-Ras and its membrane transporter, these compounds offer potential therapeutic benefits against TNBC, including its radioresistant form. Further research and clinical trials are warranted to validate their efficacy and safety as novel TNBC treatments.
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Heteropterys cotinifolia (Malpighiaceae) has been used in traditional Mexican medicine mainly for the treatment of nervous disorders. However, the specific neuropharmacological activities responsible for this use remain to be defined. The present study evaluates the antidepressant and anxiolytic effects produced by the methanolic extract of Heteropterys cotinifolia and the influence of such effects on motor activity in ICR mice. Our results show that the methanolic extract of Heteropterys cotinifolia produces a dose-dependent antidepressant effect in the forced swimming test in mice at doses from 31 to 310 mg/kg, with no reduction of mice locomotion. However, no anxiolytic properties were observed. Our findings suggest that the main extract compounds identified as chlorogenic acid and rutin may be involved in the antidepressant effects. To our knowledge, the present study constitutes the first report of pharmacological and phytochemical data of Heteropterys cotinifolia. The presence of flavonoids in the methanolic extract of Heteropterys cotinifolia may also provide further data to characterize taxonomically this species in order to be distinguished from others species closely related and belonging to the same genus.
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Antidepressivos , Comportamento Animal/efeitos dos fármacos , Malpighiaceae , Atividade Motora/efeitos dos fármacos , Extratos Vegetais , Animais , Antidepressivos/química , Antidepressivos/farmacologia , Malpighiaceae/química , Malpighiaceae/classificação , Camundongos , Camundongos Endogâmicos ICR , Neurofarmacologia/métodos , Extratos Vegetais/química , Extratos Vegetais/farmacologiaRESUMO
BACKGROUND: Gonadotropin-releasing hormone (GnRH) and its receptor (GnRHR) are both expressed by a number of malignant tumors, including those of the breast. In the latter, both behave as potent inhibitors of invasion. Nevertheless, the signaling pathways whereby the activated GnRH/GnRHR system exerts this effect have not been clearly established. In this study, we provide experimental evidence that describes components of the mechanism(s) whereby GnRH inhibits breast cancer cell invasion. METHODS: Actin polymerization and substrate adhesion was measured in the highly invasive cell line, MDA-MB-231 transiently expressing the wild-type or mutant DesK191 GnRHR by fluorometry, flow cytometric analysis, and confocal microscopy, in the absence or presence of GnRH agonist. The effect of RhoA-GTP on stress fiber formation and focal adhesion assembly was measured in MDA-MB-231 cells co-expressing the GnRHRs and the GAP domain of human p190Rho GAP-A or the dominant negative mutant GAP-Y1284D. Cell invasion was determined by the transwell migration assay. RESULTS: Agonist-stimulated activation of the wild-type GnRHR and the highly plasma membrane expressed mutant GnRHR-DesK191 transiently transfected to MDA-MB-231 cells, favored F-actin polymerization and substrate adhesion. Confocal imaging allowed detection of an association between F-actin levels and the increase in stress fibers promoted by exposure to GnRH. Pull-down assays showed that the effects observed on actin cytoskeleton resulted from GnRH-stimulated activation of RhoA GTPase. Activation of this small G protein favored the marked increase in both cell adhesion to Collagen-I and number of focal adhesion complexes leading to inhibition of the invasion capacity of MDA-MB-231 cells as disclosed by assays in Transwell Chambers. CONCLUSIONS: We here show that GnRH inhibits invasion of highly invasive breast cancer-derived MDA-MB-231 cells. This effect is mediated through an increase in substrate adhesion promoted by activation of RhoA GTPase and formation of stress fibers and focal adhesions. These observations offer new insights into the molecular mechanisms whereby activation of overexpressed GnRHRs affects cell invasion potential of this malignant cell line, and provide opportunities for designing mechanism-based adjuvant therapies for breast cancer.
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Actinas/metabolismo , Movimento Celular , Receptores LHRH/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo , Neoplasias da Mama/genética , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Busserrelina/metabolismo , Busserrelina/farmacologia , Linhagem Celular Tumoral , Ativação Enzimática/efeitos dos fármacos , Feminino , Citometria de Fluxo , Fluorometria , Adesões Focais/efeitos dos fármacos , Hormônio Liberador de Gonadotropina/farmacologia , Humanos , Immunoblotting , Células MCF-7 , Microscopia Confocal , Mutação , Invasividade Neoplásica , Polimerização/efeitos dos fármacos , Receptores LHRH/agonistas , Receptores LHRH/genética , Fibras de Estresse/metabolismo , Transfecção , Proteína rhoA de Ligação ao GTP/genéticaRESUMO
Recent studies of Hibiscus sabdariffa Linn. have demonstrated that it presents diuretic, natriuretic, and potassium sparing effects. However, the mechanism that induces these effects has not yet been elucidated. The aim of this study was to explore the possible mechanism of action for the diuretic effect of Hibiscus sabdariffa extract and its fractions.The aqueous extract from this plant and the fractions obtained with solvents of different polarities were administered to adrenalectomized rats, and the diuretic effect was measured in the presence of deoxycorticosterone acetate (aldosterone analog).The effect on renal filtration was also evaluated in an in situ kidney model, and finally, the effect of diuretic active extracts on gene expression of the alpha subunit from the transporter (αENaC) of renal epithelial cell was quantified. The subsequent results were obtained: The aqueous extract of Hibiscus sabdariffa presented the following chemical composition, 32.4 mg/g delphinidin-3-O-sambubioside, 11.5 mg/g cyanidin-3-O-sambubioside, 11.5 mg/g quercetin, and chlorogenic acid 2.7 mg/g. The concentration of anthocyanins was diminished until disappearance due to decrease of the polarity of the solvents used in the extraction process, in contrast to the flavonoids and chlorogenic acid, which had their concentration increased. The diuretic effect caused by adrenalectomy in rats was reversed by deoxycorticosterone acetate activity. However, the effect of deoxycorticosterone acetate was antagonized by spironolactone, the aqueous extract of Hibiscus sabdariffa, and the acetonitrileâ:âmethanol 5â:â5 mixture extract, administered orally. A similar effect was observed on renal filtration obtained from the isolated kidney model.When the gene expression levels of αENaC was measured in adrenalectomized rats, it was observed that spironolactone, the aqueous extract of Hibiscus sabdariffa, the acetonitrileâ:âmethanol 5â:â5 mixture, as well as the acetonitrile extract significantly decreased the expression of this protein.The conclusion of this work is that the diuretic, natriuretic, and potassium sparing effects of Hibiscus sabdariffa are due in part to the modulation of aldosterone activity by the presence in the extract of this plant of compounds potentially responsible for this modulation, as anthocyanins, flavonoids, and chlorogenic acid.
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Diuréticos/farmacologia , Canais Epiteliais de Sódio/efeitos dos fármacos , Hibiscus/química , Rim/efeitos dos fármacos , Antagonistas de Receptores de Mineralocorticoides/farmacologia , Extratos Vegetais/farmacologia , Adrenalectomia , Animais , Antocianinas/análise , Antocianinas/farmacologia , Ácido Clorogênico/análise , Dissacarídeos , Relação Dose-Resposta a Droga , Flores/química , Medicina Tradicional , México , Quercetina/análise , Ratos , Ratos Sprague-DawleyRESUMO
Although there are currently several factors that allow measuring the risk of having breast cancer or predicting its progression, the underlying causes of this malignancy have remained unknown. Several molecular studies have described some mechanisms involved in the progress of breast cancer. These have helped in identifying new targets with therapeutic potential. However, despite the therapeutic strategies implemented from the advances achieved in breast cancer research, a large percentage of patients with breast cancer die due to the spread of malignant cells to other tissues or organs, such as bones and lungs. Therefore, determining the processes that promote the migration of malignant cells remains one of the greatest challenges for oncological research. Several research groups have reported evidence on how the dedifferentiation of tumor cells leads to the acquisition of stemness characteristics, such as invasion, metastasis, the capability to evade the immunological response, and resistance to several cytotoxic drugs. These phenotypic changes have been associated with a complex reprogramming of gene expression in tumor cells during the Epithelial- Mesenchymal Transition (EMT). Considering the determining role that the transcriptional regulation plays in the expression of the specific characteristics and attributes of breast cancer during ETM, in the present work, we reviewed and analyzed several transcriptional mechanisms that support the mesenchymal phenotype. In the same way, we established the importance of transcription factors with a therapeutic perspective in the progress of breast cancer.
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Antineoplásicos , Neoplasias da Mama , Antineoplásicos/farmacologia , Neoplasias da Mama/patologia , Linhagem Celular Tumoral , Transição Epitelial-Mesenquimal/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Fatores de Transcrição/genéticaRESUMO
Breast cancer (BC) is the most commonly occurring cancer and primary cause of cancerrelated mortality in women worldwide. Investigations into BC have been conducted in in vitro and in vivo models. Of these models, the cultivation of tumor cell lines in twodimensional models is the most widely employed in vitro model to study tumor physiology. However, this approach does not accurately model all aspects observed in tumors. To address these limitations, threedimensional (3D) in vitro models have been developed. In these, it is possible to reproduce the interaction between tumor cells and the extracellular matrix, as well as the interrelationship between tumor cells and stromal cells, in order to replicate the interactions observed within the 3D environment of in vivo tumors. The present review summarizes the most common 3D in vitro models used to study BC, including spheroid models, organonachip models, hydrogel models and bioprinted models, with a discussion of their particular advantages and limitations.
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Bioimpressão/métodos , Neoplasias da Mama/patologia , Dispositivos Lab-On-A-Chip , Esferoides Celulares/patologia , Técnicas de Cultura de Células/instrumentação , Técnicas de Cultura de Células/métodos , Linhagem Celular Tumoral , Feminino , Humanos , Hidrogéis , Microambiente TumoralRESUMO
Implementations of suitable in vitro cell culture systems of the human intestine have been essential tools in the study of the interaction among organs, commensal microbiota, pathogens and parasites. Due to the great complexity exhibited by the intestinal tissue, researchers have been developing in vitro/ex vivo systems to diminish the gap between conventional cell culture models and the human intestine. These models are able to reproduce different structures and functional aspects of the tissue. In the present review, information is recapitulated on the most used models, such as cell culture, intestinal organoids, scaffold-based three-dimensional models, and organ-on-a-chip and their use in studying the interaction between human intestine and microbes, and their advantages and limitations are also discussed.
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Microbioma Gastrointestinal , Mucosa Intestinal/metabolismo , Mucosa Intestinal/microbiologia , Intestinos/fisiologia , Modelos Biológicos , Interações Hospedeiro-Patógeno , Humanos , OrganoidesRESUMO
Human gonadotropin-releasing hormone receptor (GnRHR) and its natural ligand human gonadotropin-releasing hormone (GnRH) were initially described as signaling complexes that play a key role in reproductive functions. By binding to specific receptors present on pituitary gonadotropes, GnRH regulates the sperm and ovum maturation, as well as steroidogenesis within the context of the hypothalamus-hypophysis axis. The expression of GnRH and its receptor has clearly been established in many extra-pituitary organs. Some of them are tumors from non-reproductive tissues such as liver, larynx, pancreas, colon, lymphoma, kidney, skin, blood and brain as well as tissues from reproductive track, for example ovary, endometrium, prostate and breast or tumors derived from these organs. Expression of GnRH and its receptor in these organs has gained much attention and several research groups have established their role during cell proliferation and cell motility. Although the signaling pathways and their effector proteins in these samples remain unclear, the molecular mechanism employed for GnRH and its receptor in extra-pituitary tissues could be related with non-classical GnRHR-signaling pathways. In the present review, we explore the vast literature reported on GnRH and GnRHR principally in tumors, describing how cross-talk between GnRHR and growth factor receptor, the coupling between GnRHR and many G proteins depending on cell context, and the regulation of several proteins associated with cell proliferation and cell motility are employed by GnRHR/GnRH to regulate their extra-pituitary activities.
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Hormônio Liberador de Gonadotropina/fisiologia , Sistema Hipotálamo-Hipofisário/metabolismo , Neoplasias/metabolismo , Receptores LHRH/fisiologia , Transdução de Sinais , Proteínas de Ligação ao GTP/metabolismo , HumanosRESUMO
Although significant progress has been made in the implementation of new breast cancer treatments over the last three decades, this neoplasm annually continues to show high worldwide rates of morbidity and mortality. In consequence, the search for novel therapies with greater effectiveness and specificity has not come to a stop. Among the alternative therapeutic targets, the human gonadotropin-releasing hormone type I and type II (hGnRH-I and hGnRH-II, respectively) and its receptor, the human gonadotropin-releasing hormone receptor type I (hGnRHR-I), have shown to be powerful therapeutic targets to decrease the adverse effects of this disease. In the present review, we describe how the administration of GnRH analogs is able to reduce circulating concentrations of estrogen in premenopausal women through their action on the hypothalamus-pituitary-ovarian axis, consequently reducing the growth of breast tumors and disease recurrence. Also, it has been mentioned that, regardless of the suppression of synthesis and secretion of ovarian steroids, GnRH agonists exert direct anticancer action, such as the reduction of tumor growth and cell invasion. In addition, we discuss the effects on breast cancer of the hGnRH-I and hGnRH-II agonist and antagonist, non-peptide GnRH antagonists, and cytotoxic analogs of GnRH and their implication as novel adjuvant therapies as antitumor agents for reducing the adverse effects of breast cancer. In conclusion, we suggest that the hGnRH/hGnRHR system is a promising target for pharmaceutical development in the treatment of breast cancer, especially for the treatment of advanced states of this disease.
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The Gonadotropin-Releasing Hormone Receptor (GnRHR) is expressed mainly in the gonadotrope membrane of the adenohypophysis and its natural ligand, the Gonadotropin-Releasing Hormone (GnRH), is produced in anterior hypothalamus. Furthermore, both molecules are also present in the membrane of cells derived from other reproductive tissues such as the breast, endometrium, ovary, and prostate, as well as in tumors derived from these tissues. The functions of GnRH receptor and its hormone in malignant cells have been related with the decrease of proliferation and the invasiveness of those tumors however, little is known about the molecules associated with the signaling pathways regulated by both molecules in malignant cells. To further analyze the potential mechanisms employed by the GnRHR/GnRH system to reduce the tumorigenesis of the highly invasive breast cancer cell line MDA-MB-231, we performed microarrays experiments to evaluated changes in genes expression and validate these modifications by functional assays. We show that activation of human GnRHR is able to diminish the expression and therefore functions of the Rho GTPase-Activating Protein 18 (ARHGAP18). Decrease of this GAP following GnRHR activation, correlates to the higher of cell adhesion and also with reduction of tumor cell invasion, supporting the notion that GnRHR triggers intracellular signaling pathways that acts through ARHGAP18. On the contrary, although a decline of cellular proliferation was observed during GnRHR activation in MDA-MB-231, this was independent of ARHGAP18 showing the complex system in which is involved the signaling pathways regulated by the GnRHR/GnRH system.
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Regulação para Baixo/genética , Proteínas Ativadoras de GTPase/genética , Receptores LHRH/metabolismo , Adesão Celular , Linhagem Celular Tumoral , Proliferação de Células , Colágeno Tipo I/metabolismo , Proteínas Ativadoras de GTPase/metabolismo , Regulação Neoplásica da Expressão Gênica , Humanos , Invasividade Neoplásica , Reprodutibilidade dos Testes , CicatrizaçãoRESUMO
The pathogenic amoeba Entamoeba histolytica is able to migrate within various compartments of the human body. The present article reviews progress in understanding the mechanisms of cell motility in E. histolytica during human intestinal invasion and, in particular, how the three-dimensional characteristics of the environment regulate the parasite's behaviour. The amoeboid mode of migration that applies to E. histolytica's displacements on two-dimensional surfaces is also expected to apply to the three-dimensional environment in the human intestine although several unknown, distinct modalities may be involved. Recent advances in the field of tissue engineering have provided clues on how the construction of a human colon model could help us to understand the host's intestinal physiology and its changes following amoebic infection.
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Disenteria Amebiana/parasitologia , Entamoeba histolytica/fisiologia , Intestinos/fisiopatologia , Intestinos/parasitologia , Animais , Colo/parasitologia , Colo/fisiopatologia , Entamoeba histolytica/patogenicidade , Humanos , Movimento , Técnicas de Cultura de Órgãos , Engenharia TecidualRESUMO
Recently, an increasing amount of evidence indicates that human gonadotropin-releasing hormone (hGnRH) and its receptor (hGnRHR) are important regulatory components not only to the reproduction process but also in the regulation of some cancer cell functions such as cell proliferation, in both hormone-dependent and -independent types of tumors. The hGnRHR is a naturally misfolded protein that is retained mostly in the endoplasmic reticulum; however, this mechanism can be overcome by treatment with several pharmacoperones, therefore, increasing the amount of receptors in the cell membrane. In addition, several reports indicate that the expression level of hGnRHR in tumor cells is even lower than in pituitary or gonadotrope cells. The signal transduction pathways activated by hGnRH in both gonadotrope and different cancer cell types are described in the present review. We also discuss how the rescue of misfolded receptors in tumor cells could be a promising strategy for cancer therapy.
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Hormônio Liberador de Gonadotropina/metabolismo , Neoplasias/metabolismo , Reprodução , Sequência de Aminoácidos , Neoplasias da Mama/metabolismo , Membrana Celular/metabolismo , Neoplasias do Endométrio/metabolismo , Feminino , Regulação da Expressão Gênica , Hormônios/metabolismo , Humanos , Masculino , Dados de Sequência Molecular , Neoplasias Ovarianas/metabolismo , Hipófise/metabolismo , Neoplasias da Próstata/metabolismo , Domínios Proteicos , Dobramento de Proteína , Receptores LHRH/metabolismo , Transdução de SinaisRESUMO
Posttranslational modifications occurring during the biosynthesis of G protein-coupled receptors include glycosylation and palmitoylation at conserved cysteine residues located in the carboxyl-terminus of the receptor. In a number of these receptors, these modifications play an important role in receptor function and particularly, in intracellular trafficking. In the present study, the three cysteine residues present in the carboxyl-terminus of the human FSHR were replaced with glycine by site-directed mutagenesis. Wild-type and mutant (Cys627/629/655Gly) FSHRs were then transiently expressed in HEK-293 cells and analyzed for cell-surface plasma membrane expression, agonist-stimulated signaling and internalization, and postendocytic processing in the absence and presence of lysosome and/or proteasome inhibitors. Compared with the wild-type FSHR, the triple mutant FSHR exhibited ~70% reduction in plasma membrane expression as well as a profound attenuation in agonist-stimulated cAMP production and ERK1/2 phosphorylation. Incubation of HEK-293 cells expressing the wild-type FSHR with 2-bromopalmitate (palmitoylation inhibitor) for 6 h, decreased plasma membrane expression of the receptor by ~30%. The internalization kinetics and ß-arrestin 1 and 2 recruitment were similar between the wild-type and triple mutant FSHR as disclosed by assays performed in non-equilibrium binding conditions and by confocal microscopy. Cells expressing the mutant FSHR recycled the internalized FSHR back to the plasma membrane less efficiently than those expressing the wild-type FSHR, an effect that was counteracted by proteasome but not by lysosome inhibition. These results indicate that replacement of the cysteine residues present in the carboxyl-terminus of the FSHR, impairs receptor trafficking from the endoplasmic reticulum/Golgi apparatus to the plasma membrane and its recycling from endosomes back to the cell surface following agonist-induced internalization. Since in the FSHR these cysteine residues are S-palmitoylated, the data presented emphasize on this posttranslational modification as an important factor for both upward and downward trafficking of this receptor.
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The 240-bp alpha exon of the tight junction (TJ) protein ZO-1 pre-mRNA is alternatively spliced. Expression of both ZO-1alpha+/ZO-1alpha- isoforms results in hermetic TJs, and these become leaky when ZO-1alpha- expression prevails. The alpha exon inclusion/skipping mechanism was studied by in vivo RT-PCR splicing assays in neural and epithelial cells, utilizing a canine minigene construct containing the alpha exon, and the flanking introns and exons. Inclusion of the alpha exon always occurs in wild-type MDCK cells and it is detectable in transfected HeLa cells. However, the alpha exon is skipped in transfected neural cells. Accordingly, both 5' and 3' splice sites surrounding the alpha exon appear to be suboptimal and no cis-acting splicing control elements were found in this exon. Deletion analysis revealed an 83-bp splicing enhancer in the downstream exon and a 35-bp splicing silencer at the beginning of the upstream exon. In epithelial cells all constructs rendered alpha exon inclusion. We conclude that, in neural cells, skipping of the alpha exon depends on two antagonistic exonic elements located in the flanking constitutive exons.
Assuntos
Processamento Alternativo , Proteínas de Membrana/genética , Fosfoproteínas/genética , Animais , Cães , Epitélio/metabolismo , Éxons , Regulação da Expressão Gênica , Humanos , Proteínas de Membrana/metabolismo , Fosfoproteínas/metabolismo , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Precursores de RNA/metabolismo , Proteína da Zônula de Oclusão-1RESUMO
Bouvardia ternifolia has been used medicinally to treat inflammation. In the present study, we investigate the anti-Alzheimer's potential effect of the hydroalcoholic extract of B. ternifolia through evaluation of anti-inflammatory and antioxidant activities, quantification of the percentage inhibition of acetylcholinesterase activity, protection effect against ß-amyloid fibrillar-induce neurotoxicity, and the identification of the main constituents. Our results show that B. ternifolia extract and ethyl acetate fraction induced anti-inflammatory effects by reducing inflammation by >70 %, while antioxidant test revealed significant IC50 values for flavonoid content fraction (30.67 ± 2.09 µg/ml) and ethyl acetate fraction (42.66 ± 0.93 µg/ml). The maximum inhibition of acetylcholinesterase was exhibited by scopoletin content fraction (38.43 ± 3.94 %), while ethyl acetate fraction exerted neuroprotective effect against ß-amyloid peptide (83.97 ± 5.03 %). Phytochemical analysis, showed the presence of 3-O-quercetin glucopyranoside (415 mg/g), rutin (229.9 mg/g), ursolic and oleanolic acid (54 and 20.8 mg/g respectively), 3-O-quercetin rhamnopyranoside (12.8 mg/g), chlorogenic acid (9.5 mg/g), and scopoletin (1.38 mg/g). Our findings support the use of B. ternifolia since the extract induced significant neuroprotection against ß-amyloid peptide, anti-inflammatory, antioxidant and anti-acetylcholinesterase effects that could be attributed to its contents of polyphenols, coumarins, and triterpenes, and encourage further studies for development of this extract as therapeutic agent in treatment of Alzheimer's disease.
Assuntos
Doença de Alzheimer/tratamento farmacológico , Anti-Inflamatórios/farmacologia , Antioxidantes/farmacologia , Inibidores da Colinesterase/farmacologia , Fármacos Neuroprotetores/farmacologia , Extratos Vegetais/farmacologia , Rubiaceae/química , Doença de Alzheimer/metabolismo , Peptídeos beta-Amiloides/antagonistas & inibidores , Anti-Inflamatórios/isolamento & purificação , Antioxidantes/isolamento & purificação , Sobrevivência Celular/efeitos dos fármacos , Inibidores da Colinesterase/isolamento & purificação , Cumarínicos/isolamento & purificação , Cumarínicos/farmacologia , Flavonoides/farmacologia , Humanos , Fármacos Neuroprotetores/isolamento & purificação , Fragmentos de Peptídeos/antagonistas & inibidores , Fenóis/isolamento & purificação , Fenóis/farmacologia , Componentes Aéreos da Planta/química , Extratos Vegetais/química , Triterpenos/isolamento & purificação , Triterpenos/farmacologiaRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Heteropterys brachiata is a plant species that has been used in traditional Mexican medicine for the treatment of nervous disorders. AIM OF THE STUDY: To evaluate the anxiolytic, anticonvulsant, antidepressant and sedative effects produced by the methanolic extract of Heteropterys brachiata (HbMeOH) in ICR mice. Additionally, we determine the acute toxicity profiles of the extract and the presence of its main constituents. MATERIAL AND METHODS: The neuropharmacological effects of the extract were evaluated using a variety of models, such as the elevated plus maze (EPM), the forced swimming test (FST), the pentobarbital potentiation test (PTBt), pentylenetetrazole-induced seizures test (PTZt), and the open field test (OFT). HPLC was employed for obtention of phytochemical profile. RESULTS: HbMeOH produced a significant antidepressant effect in FST at 500 and 750 mg/kg doses, while doses from 500 to 1500 mg/kg exhibited a clear dose-dependent anxiolytic activity in EPM. A dose of 500 mg/kg showed a significant anticonvulsant activity in PTZt and an absence of sedation effects in PTBt. The main compounds of HbMeOH were chlorogenic acid and chlorogenic acid methyl ester, as well as less abundant terpene-type compounds. Furthermore, the extract was either safe with no deaths in mice treated orally with 2000 mg/kg. CONCLUSIONS: HbMeOH extract which contains mainly hydroxycinnamic acids and triterpene-type compounds, possesses antidepressant, anxiolytic and anticonvulsive properties and can be considered safe or of low toxicity when orally administrated. These findings lend pharmacological justification to the traditional use of Heteropterys brachiata in the treatment of nervous disorders.