Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 74
Filtrar
Mais filtros

Base de dados
País/Região como assunto
Tipo de documento
Intervalo de ano de publicação
1.
Malar J ; 19(1): 285, 2020 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-32778117

RESUMO

BACKGROUND: Extracellular vesicles (EVs) have been broadly studied in malaria for nearly a decade. These vesicles carry various functional biomolecules including RNA families such as microRNAs (miRNA). These EVs-derived microRNAs have numerous roles in host-parasite interactions and are considered promising biomarkers for disease severity. However, this field lacks clinical studies of malaria-infected samples. In this study, EV specific miRNAs were isolated from the plasma of patients from Thailand infected with Plasmodium vivax and Plasmodium falciparum. In addition, it is postulated that these miRNAs were differentially expressed in these groups of patients and had a role in disease onset through the regulation of specific target genes. METHODS: EVs were purified from the plasma of Thai P. vivax-infected patients (n = 19), P. falciparum-infected patients (n = 18) and uninfected individuals (n = 20). EV-derived miRNAs were then prepared and abundance of hsa-miR-15b-5p, hsa-miR-16-5p, hsa-let-7a-5p and hsa-miR-150-5p was assessed in these samples. Quantitative polymerase chain reaction was performed, and relative expression of each miRNA was calculated using hsa-miR-451a as endogenous control. Then, the targets of up-regulated miRNAs and relevant pathways were predicted by using bioinformatics. Receiver Operating Characteristic with Area under the Curve (AUC) was then calculated to assess their diagnostic potential. RESULTS: The relative expression of hsa-miR-150-5p and hsa-miR-15b-5p was higher in P. vivax-infected patients compared to uninfected individuals, but hsa-let-7a-5p was up-regulated in both P. vivax-infected patients and P. falciparum-infected patients. Bioinformatic analysis revealed that these miRNAs might regulate genes involved in the malaria pathway including the adherens junction and the transforming growth factor-ß pathways. All up-regulated miRNAs could potentially be used as disease biomarkers as determined by AUC; however, the sensitivity and specificity require further investigation. CONCLUSION: An upregulation of hsa-miR-150-5p and hsa-miR-15b-5p was observed in P. vivax-infected patients while hsa-let-7a-5p was up-regulated in both P. vivax-infected and P. falciparum-infected patients. These findings will require further validation in larger cohort groups of malaria patients to fully understand the contribution of these EVs miRNAs to malaria detection and biology.


Assuntos
Vesículas Extracelulares/metabolismo , Malária Falciparum/fisiopatologia , Malária Vivax/fisiopatologia , MicroRNAs/sangue , Adulto , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Plasmodium falciparum/isolamento & purificação , Plasmodium vivax/isolamento & purificação , Tailândia , Adulto Jovem
2.
Blood ; 129(12): 1669-1679, 2017 03 23.
Artigo em Inglês | MEDLINE | ID: mdl-28096086

RESUMO

Clinical studies indicate that thrombocytopenia correlates with the development of severe falciparum malaria, suggesting that platelets either contribute to control of parasite replication, possibly as innate parasite killer cells or function in eliciting pathogenesis. Removal of platelets by anti-CD41 mAb treatment, platelet inhibition by aspirin, and adoptive transfer of wild-type (WT) platelets to CD40-KO mice, which do not control parasite replication, resulted in similar parasitemia compared with control mice. Human platelets at a physiologic ratio of 1 platelet to 9 red blood cells (RBCs) did not inhibit the in vitro development or replication of blood-stage Plasmodium falciparum The percentage of Plasmodium-infected (iRBCs) with bound platelets during the ascending parasitemia in Plasmodium chabaudi- and Plasmodium berghei-infected mice and the 48-hour in vitro cycle of P falciparum was <10%. P chabaudi and P berghei iRBCs with apoptotic parasites (TdT+) exhibited minimal platelet binding (<5%), which was similar to nonapoptotic iRBCs. These findings collectively indicate platelets do not kill bloodstage Plasmodium at physiologically relevant effector-to-target ratios. P chabaudi primary and secondary parasitemia was similar in mice depleted of platelets by mAb-injection just before infection, indicating that activation of the protective immune response does not require platelets. In contrast to the lack of an effect on parasite replication, adoptive transfer of WT platelets to CD40-KO mice, which are resistant to experimental cerebral malaria, partially restored experimental cerebral malaria mortality and symptoms in CD40-KO recipients, indicating platelets elicit pathogenesis and platelet CD40 is a key molecule.


Assuntos
Plaquetas/fisiologia , Malária/imunologia , Animais , Plaquetas/parasitologia , Antígenos CD40 , Células Cultivadas , Eritrócitos/parasitologia , Humanos , Imunidade Celular , Malária/sangue , Malária Cerebral/etiologia , Camundongos , Plasmodium chabaudi
3.
FASEB J ; 31(7): 2817-2827, 2017 07.
Artigo em Inglês | MEDLINE | ID: mdl-28314769

RESUMO

Microvesicles (MVs) are involved in cell-cell interactions, including disease pathogenesis. Nondestructive Fourier-transform infrared (FTIR) spectra from MVs were assessed as a technique to provide new biochemical insights into a LPS-induced monocyte model of septic shock. FTIR spectroscopy provided a quick method to investigate relative differences in biomolecular content of different MV populations that was complementary to traditional semiquantitative omics approaches, with which it is difficult to provide information on relative changes between classes (proteins, lipids, nucleic acids, carbohydrates) or protein conformations. Time-dependent changes were detected in biomolecular contents of MVs and in the monocytes from which they were released. Differences in phosphatidylcholine and phosphatidylserine contents were observed in MVs released under stimulation, and higher relative concentrations of RNA and α-helical structured proteins were present in stimulated MVs compared with MVs from resting cells. FTIR spectra of stimulated monocytes displayed changes that were consistent with those observed in the corresponding MVs they released. LPS-stimulated monocytes had reduced concentrations of nucleic acids, α-helical structured proteins, and phosphatidylcholine compared with resting monocytes but had an increase in total lipids. FTIR spectra of MV biomolecular content will be important in shedding new light on the mechanisms of MVs and the different roles they play in physiology and disease pathogenesis.-Lee, J., Wen, B., Carter, E. A., Combes, V., Grau, G. E. R., Lay, P. A. Infrared spectroscopic characterization of monocytic microvesicles (microparticles) released upon lipopolysaccharide stimulation.


Assuntos
Micropartículas Derivadas de Células/fisiologia , Lipopolissacarídeos/toxicidade , Monócitos/efeitos dos fármacos , Monócitos/fisiologia , Espectroscopia de Infravermelho com Transformada de Fourier , Linhagem Celular , Citometria de Fluxo , Humanos
4.
Malar J ; 17(1): 192, 2018 May 11.
Artigo em Inglês | MEDLINE | ID: mdl-29747626

RESUMO

BACKGROUND: Cerebral malaria (CM) is a fatal complication of Plasmodium infection, mostly affecting children under the age of five in the sub-Saharan African region. CM pathogenesis remains incompletely understood, although sequestered infected red blood cells, inflammatory cells aggregating in the cerebral blood vessels, and the microvesicles (MV) that they release in the circulation, have been implicated. Plasma MV numbers increase in CM patients and in the murine model, where blocking their release, genetically or pharmacologically, protects against brain pathology, suggesting a role of MV in CM neuropathogenesis. In this work, the microRNA (miRNA) cargo of MV is defined for the first time during experimental CM with the overarching hypothesis that this characterization could help understand CM pathogenesis. RESULTS: The change in abundance of miRNA was studied following infection of CBA mice with Plasmodium berghei ANKA strain (causing experimental CM), and Plasmodium yoelii, which causes severe malaria without cerebral complications, termed non-CM (NCM). miRNA expression was analyzed using microarrays to compare MV from healthy (NI) and CM mice, yielding several miRNA of interest. The differential expression profiles of these selected miRNA (miR-146a, miR-150, miR-193b, miR-205, miR-215, miR-467a, and miR-486) were analyzed in mouse MV, MV-free plasma, and brain tissue by quantitative reverse transcription PCR (RT-qPCR). Two miRNA-miR-146a and miR-193b-were confirmed as differentially abundant in MV from CM mice, compared with NCM and NI mice. These miRNA have been shown to play various roles in inflammation, and their dysregulation during CM may be critical for triggering the neurological syndrome via regulation of their potential downstream targets. CONCLUSIONS: These data suggest that, in the mouse model at least, miRNA may have a regulatory role in the pathogenesis of severe malaria.


Assuntos
Encéfalo/parasitologia , Micropartículas Derivadas de Células/parasitologia , Malária Cerebral/patologia , Malária Cerebral/fisiopatologia , Plasmodium berghei/fisiologia , Plasmodium yoelii/fisiologia , Animais , Encéfalo/patologia , Encéfalo/fisiopatologia , Malária/patologia , Malária/fisiopatologia , Camundongos , Camundongos Endogâmicos CBA , MicroRNAs/metabolismo
5.
PLoS Pathog ; 10(3): e1003839, 2014 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-24651155

RESUMO

In patients with cerebral malaria (CM), higher levels of cell-specific microparticles (MP) correlate with the presence of neurological symptoms. MP are submicron plasma membrane-derived vesicles that express antigens of their cell of origin and phosphatidylserine (PS) on their surface, facilitating their role in coagulation, inflammation and cell adhesion. In this study, the in vivo production, fate and pathogenicity of cell-specific MP during Plasmodium berghei infection of mice were evaluated. Using annexin V, a PS ligand, and flow cytometry, analysis of platelet-free plasma from infected mice with cerebral involvement showed a peak of MP levels at the time of the neurological onset. Phenotypic analyses showed that MP from infected mice were predominantly of platelet, endothelial and erythrocytic origins. To determine the in vivo fate of MP, we adoptively transferred fluorescently labelled MP from mice with CM into healthy or infected recipient mice. MP were quickly cleared following intravenous injection, but microscopic examination revealed arrested MP lining the endothelium of brain vessels of infected, but not healthy, recipient mice. To determine the pathogenicity of MP, we transferred MP from activated endothelial cells into healthy recipient mice and this induced CM-like brain and lung pathology. This study supports a pathogenic role for MP in the aggravation of the neurological lesion and suggests a causal relationship between MP and the development of CM.


Assuntos
Micropartículas Derivadas de Células/patologia , Malária Cerebral/sangue , Plasmodium berghei/patogenicidade , Transferência Adotiva , Animais , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Imunofluorescência , Camundongos , Camundongos Endogâmicos CBA , Virulência
6.
PLoS Pathog ; 10(7): e1004236, 2014 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-25033406

RESUMO

During experimental cerebral malaria (ECM) mice develop a lethal neuropathological syndrome associated with microcirculatory dysfunction and intravascular leukocyte sequestration. The precise spatio-temporal context in which the intravascular immune response unfolds is incompletely understood. We developed a 2-photon intravital microscopy (2P-IVM)-based brain-imaging model to monitor the real-time behaviour of leukocytes directly within the brain vasculature during ECM. Ly6C(hi) monocytes, but not neutrophils, started to accumulate in the blood vessels of Plasmodium berghei ANKA (PbA)-infected MacGreen mice, in which myeloid cells express GFP, one to two days prior to the onset of the neurological signs (NS). A decrease in the rolling speed of monocytes, a measure of endothelial cell activation, was associated with progressive worsening of clinical symptoms. Adoptive transfer experiments with defined immune cell subsets in recombinase activating gene (RAG)-1-deficient mice showed that these changes were mediated by Plasmodium-specific CD8(+) T lymphocytes. A critical number of CD8(+) T effectors was required to induce disease and monocyte adherence to the vasculature. Depletion of monocytes at the onset of disease symptoms resulted in decreased lymphocyte accumulation, suggesting reciprocal effects of monocytes and T cells on their recruitment within the brain. Together, our studies define the real-time kinetics of leukocyte behaviour in the central nervous system during ECM, and reveal a significant role for Plasmodium-specific CD8(+) T lymphocytes in regulating vascular pathology in this disease.


Assuntos
Linfócitos T CD8-Positivos , Células Endoteliais , Malária Cerebral , Monócitos , Plasmodium berghei/metabolismo , Animais , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/patologia , Células Endoteliais/metabolismo , Células Endoteliais/patologia , Malária Cerebral/metabolismo , Malária Cerebral/patologia , Malária Cerebral/fisiopatologia , Camundongos , Camundongos Knockout , Microscopia de Fluorescência , Monócitos/metabolismo , Monócitos/patologia
7.
Mult Scler ; 22(14): 1883-1887, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-26931477

RESUMO

BACKGROUND: No molecular marker can monitor disease progression and treatment efficacy in multiple sclerosis (MS). Circulating microparticles represent a potential snapshot of disease activity at the blood brain barrier. OBJECTIVES AND METHODS: To profile plasma microparticles by flow cytometry in MS and determine how fingolimod could impact endothelial microparticles production. RESULTS: In non-treated MS patients compared to healthy and fingolimod-treated patients, endothelial microparticles were higher, while B-cell-microparticle numbers were lower. Fingolimod dramatically reduced tumour necrosis factor (TNF)-induced endothelial microparticle release in vitro. CONCLUSION: Fingolimod restored dysregulated endothelial and B-cell-microparticle numbers, which could serve as a biomarker in MS.


Assuntos
Linfócitos B , Micropartículas Derivadas de Células , Células Endoteliais , Cloridrato de Fingolimode/farmacologia , Imunossupressores/farmacologia , Esclerose Múltipla/sangue , Esclerose Múltipla/tratamento farmacológico , Adulto , Endotélio Vascular/efeitos dos fármacos , Feminino , Cloridrato de Fingolimode/administração & dosagem , Humanos , Imunossupressores/administração & dosagem , Masculino , Pessoa de Meia-Idade
8.
J Immunol ; 193(7): 3378-87, 2014 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-25187656

RESUMO

Endothelial cells closely interact with circulating lymphocytes. Aggression or activation of the endothelium leads to an increased shedding of endothelial cell microparticles (MP). Endothelial MP (EMP) are found in high plasma levels in numerous immunoinflammatory diseases, such as atherosclerosis, sepsis, multiple sclerosis, and cerebral malaria, supporting their role as effectors and markers of vascular dysfunction. Given our recently described role for human brain microvascular endothelial cells (HBEC) in modulating immune responses, we investigated how HBEC-derived MP could interact with and support the proliferation of T cells. Like their mother cells, EMP expressed molecules important for Ag presentation and T cell costimulation, that is, ß2-microglobulin, MHC II, CD40, and ICOSL. HBEC were able to take up fluorescently labeled Ags with EMP also containing fluorescent Ags, suggestive of Ag carryover from HBEC to EMP. In cocultures, fluorescently labeled EMP from resting or cytokine-stimulated HBEC formed conjugates with both CD4(+) and CD8(+) subsets, with higher proportions of T cells binding EMP from cytokine-stimulated cells. The increased binding of EMP from cytokinestimulated HBEC to T cells was VCAM-1 and ICAM-1 dependent. Finally, in CFSE T cell proliferation assays using anti-CD3 mAb or T cell mitogens, EMP promoted the proliferation of CD4(+) T cells and that of CD8(+) T cells in the absence of exogenous stimuli and in the T cell mitogenic stimulation. Our findings provide novel evidence that EMP can enhance T cell activation and potentially ensuing Ag presentation, thereby pointing toward a novel role for MP in neuroimmunological complications of infectious diseases.


Assuntos
Apresentação de Antígeno , Encéfalo/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Proliferação de Células , Micropartículas Derivadas de Células/imunologia , Células Endoteliais/imunologia , Animais , Encéfalo/irrigação sanguínea , Encéfalo/citologia , Linfócitos T CD4-Positivos/citologia , Antígenos CD40/imunologia , Linfócitos T CD8-Positivos/citologia , Células Endoteliais/citologia , Feminino , Antígenos de Histocompatibilidade Classe II/imunologia , Humanos , Ligante Coestimulador de Linfócitos T Induzíveis/imunologia , Masculino , Camundongos , Microglobulina beta-2/imunologia
9.
Korean J Parasitol ; 54(4): 455-60, 2016 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-27658597

RESUMO

Blastocystis is a common zoonotic enteric protozoan that has been classified into 17 distinct subtypes (STs). A cross-sectional study was conducted to determine the prevalence and subtype distributions of Blastocystis in villagers living along the Chao Phraya River, Ayutthaya Province, Thailand, and to assess the risk of zoonotic infection. In total, 220 stool samples were collected, and DNA was extracted. PCR and sequencing were performed with primers targeting the small-subunit ribosomal RNA (SSU rRNA) genes. Blastocystis was present in 5.9% (13/220) of samples, and ST3 (5.0%; 11/220) was the predominant subtype, followed by ST2 (0.45%; 1/220) and ST6 (0.45%; 1/220). Phylogenetic trees were constructed with the maximum-likelihood method based on the Hasegawa-Kishino-Yano + G + I model, neighbor-joining, and maximum parsimony methods. The percentage of bootstrapped trees in which the associated taxa clustered together was relatively high. All the sequences of the Blastocystis-positive samples (KU051524-KU051536) were closely related to those from animals (pig, cattle, and chicken), indicating a zoonotic risk. Therefore, the villagers require proper health education, especially regarding the prevention of parasitic infection, to improve their personal hygiene and community health. Further studies are required to investigate the Blastocystis STs in the animals living in these villages.


Assuntos
Infecções por Blastocystis/epidemiologia , Infecções por Blastocystis/parasitologia , Blastocystis/classificação , Blastocystis/genética , Variação Genética , Genótipo , Adolescente , Adulto , Idoso , Blastocystis/isolamento & purificação , Criança , Pré-Escolar , Análise por Conglomerados , Estudos Transversais , DNA de Algas/química , DNA de Algas/genética , DNA Ribossômico/química , DNA Ribossômico/genética , Fezes/parasitologia , Feminino , Humanos , Lactente , Masculino , Pessoa de Meia-Idade , Epidemiologia Molecular , Filogenia , Prevalência , RNA Ribossômico 18S/genética , Rios , Análise de Sequência de DNA , Tailândia/epidemiologia , Adulto Jovem
10.
J Immunol ; 190(2): 669-77, 2013 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-23241892

RESUMO

Mycobacterium tuberculosis infection is characterized by a strong inflammatory response whereby a few infected macrophages within the granuloma induce sustained cellular accumulation. The mechanisms coordinating this response are poorly characterized. We hypothesized that microparticles (MPs), which are submicron, plasma membrane-derived vesicles released by cells under both physiological and pathological conditions, are involved in this process. Aerosol infection of mice with M. tuberculosis increased CD45(+) MPs in the blood after 4 wk of infection, and in vitro infection of human and murine macrophages with mycobacteria enhanced MP release. MPs derived from mycobacteria-infected macrophages were proinflammatory, and when injected into uninfected mice they induced significant neutrophil, macrophage, and dendritic cell recruitment to the injection site. When incubated with naive macrophages, these MPs enhanced proinflammatory cytokine and chemokine release, and they aided in the disruption of the integrity of a respiratory epithelial cell monolayer, providing a mechanism for the egress of cells to the site of M. tuberculosis infection in the lung. In addition, MPs colocalized with the endocytic recycling marker Rab11a within macrophages, and this association increased when the MPs were isolated from mycobacteria-infected cells. M. tuberculosis-derived MPs also carried mycobacterial Ag and were able to activate M. tuberculosis-specific CD4(+) T cells in vivo and in vitro in a dendritic cell-dependent manner. Collectively, these data identify an unrecognized role for MPs in host response against M. tuberculosis by promoting inflammation, intercellular communication, and cell migration.


Assuntos
Micropartículas Derivadas de Células/imunologia , Macrófagos/imunologia , Macrófagos/microbiologia , Mycobacterium tuberculosis/imunologia , Tuberculose/imunologia , Animais , Antígenos de Bactérias/imunologia , Transporte Biológico , Linfócitos T CD4-Positivos/imunologia , Comunicação Celular/imunologia , Linhagem Celular , Movimento Celular/imunologia , Micropartículas Derivadas de Células/metabolismo , Quimiocinas/biossíntese , Quimiocinas/imunologia , Citocinas/biossíntese , Citocinas/imunologia , Células Dendríticas/imunologia , Células Dendríticas/microbiologia , Endossomos/metabolismo , Humanos , Mediadores da Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Leucócitos/imunologia , Leucócitos/metabolismo , Ativação Linfocitária/imunologia , Macrófagos/metabolismo , Camundongos , Camundongos Transgênicos , Tuberculose/metabolismo , Proteínas rab de Ligação ao GTP/metabolismo
11.
Antimicrob Agents Chemother ; 58(1): 602-5, 2014.
Artigo em Inglês | MEDLINE | ID: mdl-24165175

RESUMO

Cerebral malaria (CM) is characterized by a dysregulated immune response that results in endothelial membrane destabilization and increased microparticle (MP) production. Citicoline (CTC) is a membrane stabilizer used for the treatment of neurological disorders. We evaluated the efficacy of CTC as adjunct therapy to aid recovery from experimental CM. We show that CTC reduces MP production in vitro; in combination with artesunate in vivo, confers partial protection against CM; and prolongs survival.


Assuntos
Antimaláricos/uso terapêutico , Citidina Difosfato Colina/uso terapêutico , Malária Cerebral/tratamento farmacológico , Animais , Artemisininas/uso terapêutico , Artesunato , Feminino , Humanos , Camundongos
12.
FASEB J ; 27(2): 672-83, 2013 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-23159932

RESUMO

Elevated endothelial microparticle (MP) levels are observed in numerous diseases, increasingly supporting roles as effectors and valuable markers of vascular dysfunction. While a contractile role for the actin cytoskeleton has been implicated in vesiculation, i.e., MP production, the precise interactions and mechanisms of its constituents, ß- and γ-cytoplasmic actins, is unknown. Human cerebral microvascular endothelial cells were stimulated with known agonists, and vesiculation development was monitored by scanning electron microscopy (SEM) and flow cytometry. These data in combination provide new insight into the kinetics, patterns of vesiculating cell recruitment, and degrees of response specific to stimuli. Reorganization of ß- and γ-actins, F-actin, vinculin, and talin accompanied significant MP release. ß-Actin redistribution into basal stress fibers following stimulation was associated with increased apically situated actin-rich particulate structures, which in turn directly correlated with electron-lucent membrane protrusions observed by SEM. Y-27632 Rho-kinase inhibition abolished basal ß-actin fiber formation, minimizing apically associated actin-rich structures, significantly reducing membrane protrusions and MP release to near basal levels. Cytoskeletal protein expression and distribution varied between MPs and mother cells, as determined by Western blot. These data strongly suggest that ß-actin plays an active facilitative role in agonist-induced protuberance formation, through mechanical interactions with newly described actin-rich structures.


Assuntos
Actinas/fisiologia , Micropartículas Derivadas de Células/fisiologia , Células Endoteliais/fisiologia , Actinas/ultraestrutura , Amidas/farmacologia , Fenômenos Biomecânicos , Calcimicina/farmacologia , Linhagem Celular , Micropartículas Derivadas de Células/efeitos dos fármacos , Micropartículas Derivadas de Células/ultraestrutura , Citoesqueleto/efeitos dos fármacos , Citoesqueleto/fisiologia , Citoesqueleto/ultraestrutura , Células Endoteliais/efeitos dos fármacos , Células Endoteliais/ultraestrutura , Inibidores Enzimáticos/farmacologia , Humanos , Interferon gama/farmacologia , Cinética , Lipopolissacarídeos/farmacologia , Microscopia Eletrônica de Varredura , Modelos Biológicos , Fragmentos de Peptídeos/farmacologia , Piridinas/farmacologia , Fator de Necrose Tumoral alfa/farmacologia , Quinases Associadas a rho/antagonistas & inibidores
14.
Infect Immun ; 81(11): 3984-91, 2013 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-23940206

RESUMO

Sequestration of infected red blood cells (iRBC) within the cerebral and pulmonary microvasculature is a hallmark of human cerebral malaria (hCM). The interaction between iRBC and the endothelium in hCM has been studied extensively and is linked to the severity of malaria. Experimental CM (eCM) caused by Plasmodium berghei ANKA reproduces most features of hCM, although the sequestration of RBC infected by P. berghei ANKA (PbA-iRBC) has not been completely delineated. The role of PbA-iRBC sequestration in the severity of eCM is not well characterized. Using static and flow cytoadherence assays, we provide the first direct in vitro evidence for the binding of PbA-iRBC to murine brain and lung microvascular endothelial cells (MVEC). We found that basal PbA-iRBC cytoadherence to MVECs was significantly higher than that of normal red blood cells (NRBC) and of RBC infected with P. berghei K173 (PbK173-iRBC), a strain that causes noncerebral malaria (NCM). MVEC prestimulation with tumor necrosis factor (TNF) failed to promote any further significant increase in mixed-stage iRBC adherence. Interestingly, enrichment of the blood for mature parasites significantly increased PbA-iRBC binding to the MVECs prestimulated with TNF, while blockade of VCAM-1 reduced this adhesion. Our study provides evidence for the firm, flow-resistant binding to endothelial cells of iRBC from strain ANKA-infected mice, which develop CM, and for less binding of iRBC from strain K173-infected mice, which develop NCM. An understanding of P. berghei cytoadherence may help elucidate the importance of sequestration in the development of CM and aid the development of antibinding therapies to help reduce the burden of this syndrome.


Assuntos
Adesão Celular , Células Endoteliais/fisiologia , Eritrócitos/fisiologia , Eritrócitos/parasitologia , Plasmodium berghei/patogenicidade , Animais , Encéfalo/citologia , Células Cultivadas , Feminino , Pulmão/citologia , Camundongos , Camundongos Endogâmicos CBA
15.
Microb Pathog ; 61-62: 43-50, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23665198

RESUMO

Aggregatibacter actinomycetemcomitans is a human pathogen that produces leukotoxin (LtxA) as a major virulence factor. In this study the effect of LtxA on microvascular endothelial cell viability and phenotype was studied. High doses of single LtxA treatment (500 ng/ml to 5 µg/ml) significantly and irreversibly decreased cell proliferation and induced apoptosis, as assessed by tetrazolium salt and annexin V assay, respectively. Apoptosis was partially inhibited by the pan-caspase inhibitor, z-VAD-fmk. LtxA caused a cell cycle arrest in the G2/M phase after 72 h. Between 500 ng/ml and 5 µg/ml, after long- or short-term stimulation LtxA increased the expression of ICAM-1 and VCAM-1, as well as the percentages of endothelial cells expressing these adhesion molecules. Thus, A. actinomycetemcomitans LtxA has substantial pro-inflammatory effects on human brain endothelial cells by upregulation of ICAM-1 and VCAM-1. Furthermore, LtxA in higher concentration was found to decrease proliferation and induces apoptosis in microvascular endothelial cells.


Assuntos
Aggregatibacter actinomycetemcomitans/metabolismo , Aggregatibacter actinomycetemcomitans/patogenicidade , Células Endoteliais/efeitos dos fármacos , Exotoxinas/farmacologia , Apoptose/efeitos dos fármacos , Toxinas Bacterianas/metabolismo , Toxinas Bacterianas/farmacologia , Proliferação de Células/efeitos dos fármacos , Células Endoteliais/citologia , Células Endoteliais/metabolismo , Exotoxinas/metabolismo , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Molécula 1 de Adesão Intercelular/efeitos dos fármacos , Molécula 1 de Adesão Intercelular/metabolismo , Pontos de Checagem da Fase M do Ciclo Celular/efeitos dos fármacos , Molécula 1 de Adesão de Célula Vascular/efeitos dos fármacos , Molécula 1 de Adesão de Célula Vascular/metabolismo , Fatores de Virulência/metabolismo , Fatores de Virulência/farmacologia
16.
FASEB J ; 26(1): 420-9, 2012 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-21965597

RESUMO

Drug resistance is a major cause of cancer treatment failure, with multidrug resistance (MDR) being the most serious, whereby cancer cells display cross-resistance to structurally and functionally unrelated drugs. MDR is caused by overexpression of the efflux transporters P-glycoprotein (P-gp) and multidrug resistance-associated protein 1 (MRP1). These transporters act to maintain sublethal intracellular drug concentrations within the cancer cell, making the population treatment unresponsive. Recently, we discovered a novel nongenetic basis to MDR whereby microparticles (MPs) transfer P-gp intercellularly from MDR donor cells to drug-sensitive recipient cells. MPs isolated from MDR leukemia and breast cancer cells were cocultured with their drug-sensitive counterparts. P-gp transfer was assessed by direct immunolabeling, and acquired transcripts and regulatory microRNAs by quantitative real-time PCR. We show that MDR MPs incorporate nucleic acids; MPs change recipient cells' transcriptional environment to reflect donor MDR phenotype, and distinct pathways exist among cancers of different origin that may be dependent on donor cells' ABCB1 overexpression. We demonstrate that this pathway exists for both hematological and nonhematological malignancies. By conferring MDR and "retemplating" the transcriptional landscape of recipient cells, MPs provide a novel pathway, having implications in the dissemination and acquisition of deleterious traits in clinical oncology.


Assuntos
Adenocarcinoma/patologia , Neoplasias da Mama/patologia , Micropartículas Derivadas de Células/patologia , Resistencia a Medicamentos Antineoplásicos/fisiologia , Proteínas Associadas à Resistência a Múltiplos Medicamentos/genética , Ácidos Nucleicos/metabolismo , Leucemia-Linfoma Linfoblástico de Células Precursoras/patologia , Adenocarcinoma/tratamento farmacológico , Neoplasias da Mama/tratamento farmacológico , Comunicação Celular/fisiologia , Linhagem Celular Tumoral , Micropartículas Derivadas de Células/ultraestrutura , Técnicas de Cocultura , Resistência a Múltiplos Medicamentos/fisiologia , Feminino , Humanos , MicroRNAs/metabolismo , Microscopia Eletrônica de Varredura , Proteínas Associadas à Resistência a Múltiplos Medicamentos/metabolismo , Fenótipo , Leucemia-Linfoma Linfoblástico de Células Precursoras/tratamento farmacológico , Transporte Proteico/fisiologia
17.
RNA Biol ; 10(8): 1333-44, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23807490

RESUMO

Interactions between glioma cells and their local environment are critical determinants of brain tumor growth, infiltration and neovascularisation. Communication with host cells and stroma via microvesicles represents one pathway by which tumors can modify their surroundings to achieve a tumor-permissive environment. Here we have taken an unbiased approach to identifying RNAs in glioma-derived microvesicles, and explored their potential to regulate gene expression in recipient cells. We find that glioma microvesicles are predominantly of exosomal origin and contain complex populations of coding and noncoding RNAs in proportions that are distinct from those in the cells from which they are derived. Microvesicles show a relative depletion in microRNA compared with their cells of origin, and are enriched in unusual or novel noncoding RNAs, most of which have no known function. Short-term exposure of brain microvascular endothelial cells to glioma microvesicles results in many gene expression changes in the endothelial cells, most of which cannot be explained by direct delivery of transcripts. Our data suggest that the scope of potential actions of tumor-derived microvesicles is much broader and more complex than previously supposed, and highlight a number of new classes of small RNA that remain to be characterized.


Assuntos
Células Endoteliais/metabolismo , Exossomos/metabolismo , Regulação Neoplásica da Expressão Gênica , Glioma/fisiopatologia , RNA Mensageiro/metabolismo , RNA não Traduzido/metabolismo , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patologia , Linhagem Celular Tumoral , Células Endoteliais/patologia , Exossomos/genética , Perfilação da Expressão Gênica , Glioma/metabolismo , Humanos , Microvasos/citologia , Neovascularização Patológica , Transporte de RNA , RNA Neoplásico/genética , RNA Neoplásico/metabolismo
18.
Microorganisms ; 11(9)2023 Sep 14.
Artigo em Inglês | MEDLINE | ID: mdl-37764162

RESUMO

Blood and tissue protozoan infections are responsible for an enormous burden in tropical and subtropical regions, even though they can also affect people living in high-income countries, mainly as a consequence of migration and travel. These pathologies are responsible for heavy socio-economic issues in endemic countries, where the lack of proper therapeutic interventions and effective vaccine strategies is still hampering their control. Moreover, the pathophysiological mechanisms associated with the establishment, progression and outcome of these infectious diseases are yet to be fully described. Among all the players, extracellular vesicles (EVs) have raised significant interest during the last decades due to their capacity to modulate inter-parasite and host-parasite interactions. In the present manuscript, we will review the state of the art of circulating host-derived EVs in clinical samples or in experimental models of human blood and tissue protozoan diseases (i.e., malaria, leishmaniasis, Chagas disease, human African trypanosomiasis and toxoplasmosis) to gain novel insights into the mechanisms of pathology underlying these conditions and to identify novel potential diagnostic markers.

19.
Sci Rep ; 13(1): 22553, 2023 12 18.
Artigo em Inglês | MEDLINE | ID: mdl-38110522

RESUMO

The use of megakaryoblastic leukemia MEG-01 cells can help reveal the mechanisms of thrombopoiesis. However, conventional in vitro activation of platelet release from MEG-01 cells requires thrombopoietin, which is costly. Here, we aim to develop a more straightforward and affordable method. Synchronization of the MEG-01 cells was initially performed using serum-free culture, followed by spontaneous cell differentiation in the presence of serum. Different stages of megakaryoblast differentiation were classified based on cell morphology, DNA content, and cell cycle. The MEG-01 cells released platelet-like particles at a level comparable to that of the thrombopoietin-activated MEG-01 cells. The platelet-like particles were distinguishable from PLP-derived extracellular vesicles and could express P-selectin following ADP activation. Importantly, the platelet-like particles induced fibrin clotting in vitro using platelet-poor plasma. Therefore, this thrombopoietin-independent cell synchronization method is an effective and straightforward method for studying megakaryopoiesis and thrombopoiesis.


Assuntos
Megacariócitos , Trombopoetina , Megacariócitos/metabolismo , Trombopoetina/farmacologia , Trombopoetina/metabolismo , Células Progenitoras de Megacariócitos , Plaquetas , Trombopoese
20.
J Cell Mol Med ; 16(8): 1731-8, 2012 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21883894

RESUMO

Platelet-derived microparticles (PMP) bind and modify the phenotype of many cell types including endothelial cells. Recently, we showed that PMP were internalized by human brain endothelial cells (HBEC). Here we intend to better characterize the internalization mechanisms of PMP and their intracellular fate. Confocal microscopy analysis of PKH67-labelled PMP distribution in HBEC showed PMP in early endosome antigen 1 positive endosomes and in LysoTracker-labelled lysosomes, confirming a role for endocytosis in PMP internalization. No fusion of calcein-loaded PMP with HBEC membranes was observed. Quantification of PMP endocytosis using flow cytometry revealed that it was partially inhibited by trypsin digestion of PMP surface proteins and by extracellular Ca(2+) chelation by EDTA, suggesting a partial role for receptor-mediated endocytosis in PMP uptake. This endocytosis was independent of endothelial receptors such as intercellular adhesion molecule-1 and vascular cell adhesion molecule-1 and was not increased by tumour necrosis factor stimulation of HBEC. Platelet-derived microparticle internalization was dramatically increased in the presence of decomplemented serum, suggesting a role for PMP opsonin-dependent phagocytosis. Platelet-derived microparticle uptake was greatly diminished by treatment of HBEC with cytochalasin D, an inhibitor of microfilament formation required for both phagocytosis and macropinocytosis, with methyl-ß-cyclodextrin that depletes membrane cholesterol needed for macropinocytosis and with amiloride that inhibits the Na(+)/H(+) exchanger involved in macropinocytosis. In conclusion, PMP are taken up by active endocytosis in HBEC, involving mechanisms consistent with both phagocytosis and macropinocytosis. These findings identify new processes by which PMP could modify endothelial cell phenotype and functions.


Assuntos
Plaquetas/citologia , Plaquetas/metabolismo , Encéfalo/citologia , Micropartículas Derivadas de Células/metabolismo , Endocitose , Células Endoteliais/metabolismo , Espaço Intracelular/metabolismo , Endossomos/metabolismo , Células Endoteliais/citologia , Fluoresceínas/metabolismo , Humanos , Lisossomos/metabolismo , Fusão de Membrana , Frações Subcelulares/metabolismo
SELEÇÃO DE REFERÊNCIAS
DETALHE DA PESQUISA