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1.
Nat Commun ; 15(1): 2778, 2024 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-38555361

RESUMO

Bacterial genotoxins damage host cells by targeting their chromosomal DNA. In the present study, we demonstrate that a genotoxin of Salmonella Typhi, typhoid toxin, triggers the senescence-associated secretory phenotype (SASP) by damaging mitochondrial DNA. The actions of typhoid toxin disrupt mitochondrial DNA integrity, leading to mitochondrial dysfunction and disturbance of redox homeostasis. Consequently, it facilitates the release of damaged mitochondrial DNA into the cytosol, activating type I interferon via the cGAS-STING pathway. We also reveal that the GCN2-mediated integrated stress response plays a role in the upregulation of inflammatory components depending on the STING signaling axis. These SASP factors can propagate the senescence effect on T cells, leading to senescence in these cells. These findings provide insights into how a bacterial genotoxin targets mitochondria to trigger a proinflammatory SASP, highlighting a potential therapeutic target for an anti-toxin intervention.


Assuntos
Fenótipo Secretor Associado à Senescência , Febre Tifoide , Humanos , Febre Tifoide/metabolismo , Mutagênicos/metabolismo , Senescência Celular/fisiologia , Mitocôndrias/metabolismo , DNA Mitocondrial/metabolismo , Salmonella , Fenótipo
2.
J Biomed Sci ; 29(1): 96, 2022 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-36376874

RESUMO

In the past decade, single-cell technologies have revealed the heterogeneity of the tumor-immune microenvironment at the genomic, transcriptomic, and proteomic levels and have furthered our understanding of the mechanisms of tumor development. Single-cell technologies have also been used to identify potential biomarkers. However, spatial information about the tumor-immune microenvironment such as cell locations and cell-cell interactomes is lost in these approaches. Recently, spatial multi-omics technologies have been used to study transcriptomes, proteomes, and metabolomes of tumor-immune microenvironments in several types of cancer, and the data obtained from these methods has been combined with immunohistochemistry and multiparameter analysis to yield markers of cancer progression. Here, we review numerous cutting-edge spatial 'omics techniques, their application to study of the tumor-immune microenvironment, and remaining technical challenges.


Assuntos
Neoplasias , Proteômica , Humanos , Proteômica/métodos , Microambiente Tumoral/genética , Genômica/métodos , Neoplasias/metabolismo , Transcriptoma , Biomarcadores , Biomarcadores Tumorais/genética
4.
J Clin Invest ; 131(21)2021 11 01.
Artigo em Inglês | MEDLINE | ID: mdl-34720095

RESUMO

To explore how the immune system controls clearance of SARS-CoV-2, we used a single-cell, mass cytometry-based proteomics platform to profile the immune systems of 21 patients who had recovered from SARS-CoV-2 infection without need for admission to an intensive care unit or for mechanical ventilation. We focused on receptors involved in interactions between immune cells and virus-infected cells. We found that the diversity of receptor repertoires on natural killer (NK) cells was negatively correlated with the viral clearance rate. In addition, NK subsets expressing the receptor DNAM1 were increased in patients who more rapidly recovered from infection. Ex vivo functional studies revealed that NK subpopulations with high DNAM1 expression had cytolytic activities in response to target cell stimulation. We also found that SARS-CoV-2 infection induced the expression of CD155 and nectin-4, ligands of DNAM1 and its paired coinhibitory receptor TIGIT, which counterbalanced the cytolytic activities of NK cells. Collectively, our results link the cytolytic immune responses of NK cells to the clearance of SARS-CoV-2 and show that the DNAM1 pathway modulates host-pathogen interactions during SARS-CoV-2 infection.


Assuntos
COVID-19/imunologia , COVID-19/virologia , Células Matadoras Naturais/imunologia , Receptores de Células Matadoras Naturais/imunologia , SARS-CoV-2/imunologia , Adolescente , Adulto , Idoso , Animais , Antígenos de Diferenciação de Linfócitos T/imunologia , Moléculas de Adesão Celular/imunologia , Estudos de Coortes , Citotoxicidade Imunológica , Feminino , Xenoenxertos , Interações entre Hospedeiro e Microrganismos/imunologia , Humanos , Imunofenotipagem , Técnicas In Vitro , Ligantes , Masculino , Camundongos , Camundongos SCID , Pessoa de Meia-Idade , Subfamília D de Receptores Semelhantes a Lectina de Células NK/imunologia , Pandemias , Receptores Imunológicos/imunologia , Receptores Virais/imunologia , Carga Viral , Adulto Jovem
5.
Am J Cancer Res ; 11(10): 4994-5005, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-34765306

RESUMO

SARS-CoV-2 exploits the host cellular machinery for virus replication leading to the acute syndrome of coronavirus disease 2019 (COVID-19). Growing evidence suggests SARS-CoV-2 also exacerbates many chronic diseases, including cancers. As mutations on the spike protein (S) emerged as dominant variants that reduce vaccine efficacy, little is known about the relation between SARS-CoV-2 virus variants and cancers. Compared to the SARS-CoV-2 wild-type, the Gamma variant contains two additional NXT/S glycosylation motifs on the S protein. The hyperglycosylated S of Gamma variant is more stable, resulting in more significant epithelial-mesenchymal transition (EMT) potential. SARS-CoV-2 infection promoted NF-κB signaling activation and p65 nuclear translocation, inducing Snail expression. Pharmacologic inhibition of NF-κB activity by nature food compound, I3C suppressed viral replication and Gamma variant-mediated breast cancer metastasis, indicating that NF-κB inhibition can reduce chronic disease in COVID-19 patients. Our study revealed that the Gamma variant of SARS-CoV-2 activates NF-κB and, in turn, triggers the pro-survival function for cancer progression.

6.
Nat Commun ; 12(1): 2163, 2021 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-33846331

RESUMO

γδ T cells are a distinct subgroup of T cells that bridge the innate and adaptive immune system and can attack cancer cells in an MHC-unrestricted manner. Trials of adoptive γδ T cell transfer in solid tumors have had limited success. Here, we show that DNA methyltransferase inhibitors (DNMTis) upregulate surface molecules on cancer cells related to γδ T cell activation using quantitative surface proteomics. DNMTi treatment of human lung cancer potentiates tumor lysis by ex vivo-expanded Vδ1-enriched γδ T cells. Mechanistically, DNMTi enhances immune synapse formation and mediates cytoskeletal reorganization via coordinated alterations of DNA methylation and chromatin accessibility. Genetic depletion of adhesion molecules or pharmacological inhibition of actin polymerization abolishes the potentiating effect of DNMTi. Clinically, the DNMTi-associated cytoskeleton signature stratifies lung cancer patients prognostically. These results support a combinatorial strategy of DNMTis and γδ T cell-based immunotherapy in lung cancer management.


Assuntos
Citoesqueleto/metabolismo , Citotoxicidade Imunológica/genética , Epigênese Genética , Sinapses Imunológicas/genética , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/imunologia , Receptores de Antígenos de Linfócitos T gama-delta/metabolismo , Citoesqueleto de Actina/efeitos dos fármacos , Citoesqueleto de Actina/metabolismo , Animais , Linhagem Celular Tumoral , Citoesqueleto/efeitos dos fármacos , Citotoxicidade Imunológica/efeitos dos fármacos , DNA (Citosina-5-)-Metiltransferases/antagonistas & inibidores , DNA (Citosina-5-)-Metiltransferases/metabolismo , Decitabina/farmacologia , Inibidores Enzimáticos/farmacologia , Epigênese Genética/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Sinapses Imunológicas/efeitos dos fármacos , Marcação por Isótopo , Ativação Linfocitária/efeitos dos fármacos , Ativação Linfocitária/genética , Subpopulações de Linfócitos/efeitos dos fármacos , Subpopulações de Linfócitos/metabolismo , Masculino , Camundongos Endogâmicos NOD , Fosfotirosina/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Análise de Sobrevida , Proteína Supressora de Tumor p53/metabolismo , Regulação para Cima/efeitos dos fármacos , Ensaios Antitumorais Modelo de Xenoenxerto
7.
JCI Insight ; 5(20)2020 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-33055428

RESUMO

Loss-of-function variants of protein tyrosine phosphatase non-receptor type 2 (PTPN2) enhance risk of inflammatory bowel disease and rheumatoid arthritis; however, whether the association between PTPN2 and autoimmune arthritis depends on gut inflammation is unknown. Here we demonstrate that induction of subclinical intestinal inflammation exacerbates development of autoimmune arthritis in SKG mice. Ptpn2-haploinsufficient SKG mice - modeling human carriers of disease-associated variants of PTPN2 - displayed enhanced colitis-induced arthritis and joint accumulation of Tregs expressing RAR-related orphan receptor γT (RORγt) - a gut-enriched Treg subset that can undergo conversion into FoxP3-IL-17+ arthritogenic exTregs. SKG colonic Tregs underwent higher conversion into arthritogenic exTregs when compared with peripheral Tregs, which was exacerbated by haploinsufficiency of Ptpn2. Ptpn2 haploinsufficiency led to selective joint accumulation of RORγt-expressing Tregs expressing the colonic marker G protein-coupled receptor 15 (GPR15) in arthritic mice and selectively enhanced conversion of GPR15+ Tregs into exTregs in vitro and in vivo. Inducible Treg-specific haploinsufficiency of Ptpn2 enhanced colitis-induced SKG arthritis and led to specific joint accumulation of GPR15+ exTregs. Our data validate the SKG model for studies at the interface between intestinal and joint inflammation and suggest that arthritogenic variants of PTPN2 amplify the link between gut inflammation and arthritis through conversion of colonic Tregs into exTregs.


Assuntos
Artrite/genética , Doenças Autoimunes/genética , Proteínas de Ligação a DNA/genética , Proteína Tirosina Fosfatase não Receptora Tipo 2/genética , Receptores Acoplados a Proteínas G/genética , Receptores de Peptídeos/genética , Animais , Artrite/induzido quimicamente , Artrite/patologia , Doenças Autoimunes/induzido quimicamente , Doenças Autoimunes/patologia , Colite/induzido quimicamente , Colite/genética , Colite/patologia , Colo/efeitos dos fármacos , Colo/metabolismo , Fatores de Transcrição Forkhead/genética , Regulação da Expressão Gênica , Haploinsuficiência/genética , Humanos , Inflamação/induzido quimicamente , Inflamação/genética , Inflamação/patologia , Interleucina-17/genética , Intestinos/patologia , Articulações/metabolismo , Articulações/patologia , Mananas/toxicidade , Camundongos , Camundongos Knockout , Dodecilsulfato de Sódio/toxicidade , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia
8.
Sci Signal ; 13(623)2020 03 17.
Artigo em Inglês | MEDLINE | ID: mdl-32184287

RESUMO

The hematopoietic-specific protein tyrosine phosphatase nonreceptor type 22 (PTPN22) is encoded by a major autoimmunity risk gene. PTPN22 inhibits T cell activation by dephosphorylating substrates involved in proximal T cell receptor (TCR) signaling. Here, we found by mass spectrometry that PTPN22 was phosphorylated at Ser751 by PKCα in Jurkat and primary human T cells activated with phorbol ester/ionomycin or antibodies against CD3/CD28. The phosphorylation of PTPN22 at Ser751 prolonged its half-life by inhibiting K48-linked ubiquitination and impairing recruitment of the phosphatase to the plasma membrane, which is necessary to inhibit proximal TCR signaling. Additionally, the phosphorylation of PTPN22 at Ser751 enhanced the interaction of PTPN22 with the carboxyl-terminal Src kinase (CSK), an interaction that is impaired by the PTPN22 R620W variant associated with autoimmune disease. The phosphorylation of Ser751 did not affect the recruitment of PTPN22 R620W to the plasma membrane but protected this mutant from degradation. Together, out data indicate that phosphorylation at Ser751 mediates a reciprocal regulation of PTPN22 stability versus translocation to TCR signaling complexes by CSK-dependent and CSK-independent mechanisms.


Assuntos
Proteína Tirosina Fosfatase não Receptora Tipo 22/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Serina/metabolismo , Transdução de Sinais , Doenças Autoimunes/genética , Doenças Autoimunes/metabolismo , Proteína Tirosina Quinase CSK/metabolismo , Células Cultivadas , Células HEK293 , Humanos , Células Jurkat , Espectrometria de Massas/métodos , Mutação de Sentido Incorreto , Fosforilação , Proteína Tirosina Fosfatase não Receptora Tipo 22/genética , Serina/genética , Linfócitos T/metabolismo
9.
Nat Commun ; 9(1): 463, 2018 01 31.
Artigo em Inglês | MEDLINE | ID: mdl-29386580

RESUMO

X-linked lymphoproliferative syndrome type-2 (XLP-2) is a primary immunodeficiency disease attributed to XIAP mutation and is triggered by infection. Here, we show that mouse Xiap-/- regulatory T (Treg) cells and human XIAP-deficient Treg cells are defective in suppressive function. The Xiap-/- Treg cell defect is linked partly to decreased SOCS1 expression. XIAP binds SOCS1 and promotes SOCS1 stabilization. Foxp3 stability is reduced in Xiap-/- Treg cells. In addition, Xiap-/- Treg cells are prone to IFN-γ secretion. Transfer of wild-type Treg cells partly rescues infection-induced inflammation in Xiap-/- mice. Notably, inflammation-induced reprogramming of Xiap-/- Treg cells can be prevented by blockade of the IL-6 receptor (IL-6R), and a combination of anti-IL-6R and Xiap-/- Treg cells confers survival to inflammatory infection in Xiap-/- mice. Our results suggest that XLP-2 can be corrected by combination treatment with autologous iTreg (induced Treg) cells and anti-IL-6R antibody, bypassing the necessity to transduce Treg cells with XIAP.


Assuntos
Doenças Genéticas Ligadas ao Cromossomo X/imunologia , Transtornos Linfoproliferativos/imunologia , Receptores de Interleucina-6/antagonistas & inibidores , Linfócitos T Reguladores/imunologia , Animais , Fatores de Transcrição Forkhead/metabolismo , Humanos , Interferon gama/metabolismo , Camundongos , Camundongos Knockout , Estabilidade Proteica , Proteína 1 Supressora da Sinalização de Citocina/metabolismo , Linfócitos T Reguladores/metabolismo , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/genética
10.
Nat Commun ; 7: 11904, 2016 06 17.
Artigo em Inglês | MEDLINE | ID: mdl-27312851

RESUMO

Death-associated protein kinase (DAPK) is a tumour suppressor. Here we show that DAPK also inhibits T helper 17 (Th17) and prevents Th17-mediated pathology in a mouse model of autoimmunity. We demonstrate that DAPK specifically downregulates hypoxia-inducible factor 1α (HIF-1α). In contrast to the predominant nuclear localization of HIF-1α in many cell types, HIF-1α is located in both the cytoplasm and nucleus in T cells, allowing for a cytosolic DAPK-HIF-1α interaction. DAPK also binds prolyl hydroxylase domain protein 2 (PHD2) and increases HIF-1α-PHD2 association. DAPK thereby promotes the proline hydroxylation and proteasome degradation of HIF-1α. Consequently, DAPK deficiency leads to excess HIF-1α accumulation, enhanced IL-17 expression and exacerbated experimental autoimmune encephalomyelitis. Additional knockout of HIF-1α restores the normal differentiation of Dapk(-/-) Th17 cells and prevents experimental autoimmune encephalomyelitis development. Our results reveal a mechanism involving DAPK-mediated degradation of cytoplasmic HIF-1α, and suggest that raising DAPK levels could be used for treatment of Th17-associated inflammatory diseases.


Assuntos
Proteínas Quinases Associadas com Morte Celular/genética , Encefalomielite Autoimune Experimental/genética , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Prolina Dioxigenases do Fator Induzível por Hipóxia/genética , Células Th17/imunologia , Animais , Proteínas Quinases Associadas com Morte Celular/deficiência , Proteínas Quinases Associadas com Morte Celular/imunologia , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/patologia , Regulação da Expressão Gênica , Células HEK293 , Células HeLa , Humanos , Hidroxilação , Subunidade alfa do Fator 1 Induzível por Hipóxia/imunologia , Prolina Dioxigenases do Fator Induzível por Hipóxia/antagonistas & inibidores , Prolina Dioxigenases do Fator Induzível por Hipóxia/imunologia , Interleucina-17/genética , Interleucina-17/imunologia , Células Jurkat , Camundongos , Camundongos Knockout , Glicoproteína Mielina-Oligodendrócito/administração & dosagem , Fragmentos de Peptídeos/administração & dosagem , Toxina Pertussis/administração & dosagem , Prolina/metabolismo , Complexo de Endopeptidases do Proteassoma , Proteólise , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais , Linfócitos T Reguladores/efeitos dos fármacos , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia , Células Th17/efeitos dos fármacos , Células Th17/patologia
11.
Mol Microbiol ; 97(6): 1128-41, 2015 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-26082024

RESUMO

Helicobacter pylori inhabits the gastric mucosa where it senses and responds to various stresses via a two-component systems (TCSs) that enable its persistent colonization. The aim of this study was to investigate whether any of the three paired TCSs (ArsRS, FleRS and CrdRS) in H. pylori respond to nitrosative stress. The results showed that the expression of crdS was significantly increased upon exposure to nitric oxide (NO). crdS-knockout (ΔcrdS) and crdR/crdS-knockout (ΔcrdRS) H. pylori, but not arsS-knockout (ΔarsS) or fleS-knockout (ΔfleS) H. pylori, showed a significant loss of viability upon exposure to NO compared with wild-type strain. Knockin crdS (ΔcrdS-in) significantly restored viability in the presence of NO. Global transcriptional profiling analysis of wild-type and ΔcrdS H. pylori in the presence or absence of NO showed that 101 genes were differentially expressed, including copper resistance determinant A (crdA), transport, binding and envelope proteins. The CrdR binding motifs were investigated by competitive electrophoretic mobility shift assay, which revealed that the two AC-rich regions in the crdA promoter region are required for binding. These results demonstrate that CrdR-crdA interaction enables H. pylori to survive under nitrosative stress.


Assuntos
Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Helicobacter pylori/metabolismo , Óxido Nítrico/metabolismo , Estresse Fisiológico , Sequência de Bases , Cobre/metabolismo , Proteínas de Ligação a DNA/metabolismo , Perfilação da Expressão Gênica , Técnicas de Inativação de Genes , Helicobacter pylori/genética , Dados de Sequência Molecular , Regiões Promotoras Genéticas
12.
Nat Commun ; 6: 6353, 2015 Feb 19.
Artigo em Inglês | MEDLINE | ID: mdl-25695215

RESUMO

Application of regulatory T cells (Tregs) in transplantation, autoimmunity and allergy has been extensively explored, but how Foxp3 and Treg stability is regulated in vivo is incompletely understood. Here, we identify a requirement for Deltex1 (DTX1), a contributor to T-cell anergy and Foxp3 protein level maintenance in vivo. Dtx1(-/-) Tregs are as effective as WT Tregs in the inhibition of CD4(+)CD25(-) T-cell activation in vitro. However, the suppressive ability of Dtx1(-/-) Tregs is greatly impaired in vivo. We find that Foxp3 expression is diminished when Dtx1(-/-) Tregs are co-transferred with effector T cells in vivo. DTX1 promotes the degradation of HIF-1α. Knockout of HIF-1α restores the Foxp3 stability and rescues the defective suppressive activity in Dtx1(-/-) Treg cells in vivo. Our results suggest that DTX1 exerts another level of control on Treg stability in vivo by sustaining the expression of Foxp3 protein in Tregs.


Assuntos
Proteínas de Ligação a DNA/metabolismo , Fatores de Transcrição Forkhead/metabolismo , Hipersensibilidade/imunologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Linfócitos T Reguladores/fisiologia , Animais , Feminino , Doenças Inflamatórias Intestinais/imunologia , Ativação Linfocitária , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ubiquitina-Proteína Ligases
13.
Blood ; 124(18): 2847-57, 2014 Oct 30.
Artigo em Inglês | MEDLINE | ID: mdl-25190756

RESUMO

Emerging evidence indicates that innate immunodeficiency syndromes are linked to mutations in innate receptors and to specific infections. X-linked lymphoproliferative syndrome type-2 (XLP-2) is associated with deficiency in X-linked inhibitor of apoptosis protein (XIAP), with poorly understood molecular mechanisms. Here we showed that XIAP deficiency selectively impaired B-cell chronic lymphocytic leukemia/lymphoma 10 (BCL10)-mediated innate responses to dectin-1 ligands but did not affect responses to various Toll-like receptor agonists. Consequently, Xiap(-/-) mice became highly vulnerable on Candida albicans infection. The compromised early innate responses led to the persistent presence of C albicans and inflammatory cytokines in Xiap(-/-) mice. Furthermore, priming of Xiap(-/-) mice with the dectin-1 ligand curdlan alone resulted in XLP-2-like syndromes. Restoration of dectin-1-induced Rac1 activation and phagocytosis by resolvin D1, but not up-regulation of nuclear factor-κB, rescued Xiap(-/-) mice from C albicans lethal infection. Therefore, development of XLP-2 in XIAP-deficient patients could be partly due to sustained inflammation as a consequence of defective BCL10-dependent innate immunity toward specific pathogens. Importantly, our results suggest the potential therapeutic value of resolvin D1 in the treatment of XLP-2 and innate immunodeficiency syndromes.


Assuntos
Candidíase/imunologia , Candidíase/patologia , Imunidade Inata , Proteínas Inibidoras de Apoptose/deficiência , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Animais , Proteína 10 de Linfoma CCL de Células B , Candida albicans/efeitos dos fármacos , Candida albicans/fisiologia , Candidíase/microbiologia , Receptores ErbB/metabolismo , Doenças Genéticas Ligadas ao Cromossomo X/imunologia , Doenças Genéticas Ligadas ao Cromossomo X/patologia , Humanos , Imidazóis/farmacologia , Imunidade Inata/efeitos dos fármacos , Proteínas Inibidoras de Apoptose/metabolismo , Lectinas Tipo C/agonistas , Lectinas Tipo C/metabolismo , Lipopeptídeos/farmacologia , Lipopolissacarídeos/farmacologia , Transtornos Linfoproliferativos/imunologia , Transtornos Linfoproliferativos/patologia , Lisina/metabolismo , Lisofosfolipídeos/metabolismo , Macrófagos/efeitos dos fármacos , Macrófagos/metabolismo , Camundongos , NF-kappa B/metabolismo , Fagocitose/efeitos dos fármacos , Poli I-C/farmacologia , Ligação Proteica/efeitos dos fármacos , Receptores de Antígenos de Linfócitos T/metabolismo , Receptores Toll-Like/metabolismo , Fator de Necrose Tumoral alfa/farmacologia , Ubiquitinação/efeitos dos fármacos , beta-Glucanas
14.
Molecules ; 18(6): 6521-31, 2013 Jun 03.
Artigo em Inglês | MEDLINE | ID: mdl-23736788

RESUMO

This study develops a new solvent-compatible microfluidic chip based on phenol formaldehyde resin (PFR). In addition to its solvent-resistant characteristics, this microfluidic platform also features easy fabrication, organization, decomposition for cleaning, and reusability compared with conventional chips. Both solvent-dependent (e.g., polycaprolactone) and nonsolvent-dependent (e.g., chitosan) microparticles were successfully prepared. The size of emulsion droplets could be easily adjusted by tuning the flow rates of the dispersed/continuous phases. After evaporation, polycaprolactone microparticles ranging from 29.3 to 62.7 µm and chitosan microparticles ranging from 215.5 to 566.3 µm were obtained with a 10% relative standard deviation in size. The proposed PFR microfluidic platform has the advantages of active control of the particle size with a narrow size distribution as well as a simple and low cost process with a high throughput.


Assuntos
Quitosana/química , Formaldeído/química , Microfluídica/métodos , Microesferas , Fenóis/química , Poliésteres/química , Polímeros/química , Emulsões , Técnicas Analíticas Microfluídicas , Tamanho da Partícula
15.
Sensors (Basel) ; 12(2): 1455-67, 2012.
Artigo em Inglês | MEDLINE | ID: mdl-22438719

RESUMO

This study presents the development of a robust aluminum-based microfluidic chip fabricated by conventional mechanical micromachining (computer numerical control-based micro-milling process). It applied the aluminum-based microfluidic chip to form poly(lactic-co-glycolic acid) (PLGA) microparticles encapsulating CdSe/ZnS quantum dots (QDs). A cross-flow design and flow-focusing system were employed to control the oil-in-water (o/w) emulsification to ensure the generation of uniformly-sized droplets. The size of the droplets could be tuned by adjusting the flow rates of the water and oil phases. The proposed microfluidic platform is easy to fabricate, set up, organize as well as program, and is valuable for further applications under harsh reaction conditions (high temperature and/or strong organic solvent systems). The proposed method has the advantages of actively controlling the droplet diameter, with a narrow size distribution, good sphericity, as well as being a simple process with a high throughput. In addition to the fluorescent PLGA microparticles in this study, this approach can also be applied to many applications in the pharmaceutical and biomedical area.


Assuntos
Alumínio/química , Materiais Revestidos Biocompatíveis/síntese química , Ácido Láctico/química , Ácido Láctico/síntese química , Técnicas Analíticas Microfluídicas/instrumentação , Ácido Poliglicólico/química , Ácido Poliglicólico/síntese química , Pontos Quânticos , Espectrometria de Fluorescência/métodos , Desenho de Equipamento , Análise de Falha de Equipamento , Microesferas , Copolímero de Ácido Poliláctico e Ácido Poliglicólico
16.
FEMS Microbiol Lett ; 323(2): 155-63, 2011 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22092715

RESUMO

Upon infection of the gastric epithelial cells, the Helicobacter pylori cytotoxin-associated gene A (CagA) virulence protein is injected into the epithelial cells via the type IV secretion system (TFSS), which is dependent on cholesterol. Translocated CagA is targeted by the membrane-recruited c-Src family kinases in which a tyrosine residue in the Glu-Pro-Ile-Tyr-Ala (EPIYA)-repeat region, which can be phosphorylated, induces cellular responses, including interleukin-8 (IL-8) secretion and hummingbird phenotype formation. In this study, we explored the role of EPIYA-containing C-terminal domain (CTD) in CagA tethering to the membrane lipid rafts and in IL-8 activity. We found that disruption of the lipid rafts reduced the level of CagA translocation/phosphorylation as well as CagA-mediated IL-8 secretion. By CagA truncated mutagenesis, we identified that the CTD, rather than the N-terminal domain, was responsible for CagA tethering to the plasma membrane and association with detergent-resistant membranes, leading to CagA-induced IL-8 promoter activity. Our results suggest that CagA CTD-containing EPIYAs directly interact with cholesterol-rich microdomains that induce efficient IL-8 secretion in the epithelial cells.


Assuntos
Antígenos de Bactérias/imunologia , Proteínas de Bactérias/imunologia , Colesterol/metabolismo , Células Epiteliais/imunologia , Células Epiteliais/microbiologia , Helicobacter pylori/imunologia , Interleucina-8/metabolismo , Células Cultivadas , Humanos , Estrutura Terciária de Proteína , Transporte Proteico
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