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1.
Br J Cancer ; 128(4): 528-536, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-36418894

RESUMO

BACKGROUND: Distinct sets of microbes contribute to colorectal cancer (CRC) initiation and progression. Some occur due to the evolving intestinal environment but may not contribute to disease. In contrast, others may play an important role at particular times during the tumorigenic process. Here, we describe changes in the microbiota and host over the course of azoxymethane (AOM)-induced tumorigenesis. METHODS: Mice were administered AOM or PBS and were euthanised 8, 12, 24 and 48 weeks later. Samples were analysed using 16S rRNA gene sequencing, UPLC-MS and qRT-PCR. RESULTS: The microbiota and bile acid profile showed distinct changes at each timepoint. The inflammatory response became apparent at weeks 12 and 24. Moreover, significant correlations between individual taxa, cytokines and bile acids were detected. One co-abundance group (CAG) differed significantly between PBS- and AOM-treated mice at week 24. Correlation analysis also revealed significant associations between CAGs, bile acids and the bile acid transporter, ASBT. Aberrant crypt foci and adenomas were first detectable at weeks 24 and 48, respectively. CONCLUSION: The observed changes precede host hyperplastic transformation and may represent early therapeutic targets for the prevention or management of CRC at specific timepoints in the tumorigenic process.


Assuntos
Neoplasias do Colo , Microbioma Gastrointestinal , Camundongos , Animais , Azoximetano/efeitos adversos , Ácidos e Sais Biliares/efeitos adversos , RNA Ribossômico 16S , Cromatografia Líquida , Espectrometria de Massas em Tandem , Neoplasias do Colo/induzido quimicamente , Carcinogênese , Colo , Modelos Animais de Doenças
2.
Nat Commun ; 11(1): 1512, 2020 03 23.
Artigo em Inglês | MEDLINE | ID: mdl-32251296

RESUMO

Studies of inflammatory bowel disease (IBD) have been inconclusive in relating microbiota with distribution of inflammation. We report microbiota, host transcriptomics, epigenomics and genetics from matched inflamed and non-inflamed colonic mucosa [50 Crohn's disease (CD); 80 ulcerative colitis (UC); 31 controls]. Changes in community-wide and within-patient microbiota are linked with inflammation, but we find no evidence for a distinct microbial diagnostic signature, probably due to heterogeneous host-microbe interactions, and show only marginal microbiota associations with habitual diet. Epithelial DNA methylation improves disease classification and is associated with both inflammation and microbiota composition. Microbiota sub-groups are driven by dominant Enterbacteriaceae and Bacteroides species, representative strains of which are pro-inflammatory in vitro, are also associated with immune-related epigenetic markers. In conclusion, inflamed and non-inflamed colonic segments in both CD and UC differ in microbiota composition and epigenetic profiles.


Assuntos
Colite Ulcerativa/imunologia , Doença de Crohn/imunologia , Epigênese Genética/imunologia , Microbioma Gastrointestinal/imunologia , Interações entre Hospedeiro e Microrganismos/imunologia , Adulto , Idoso , Bacteroides/genética , Bacteroides/imunologia , Bacteroides/isolamento & purificação , Biópsia , Células CACO-2 , Estudos de Casos e Controles , Estudos de Coortes , Colite Ulcerativa/genética , Colite Ulcerativa/microbiologia , Colite Ulcerativa/patologia , Colo/diagnóstico por imagem , Colo/imunologia , Colo/microbiologia , Colo/patologia , Colonoscopia , Doença de Crohn/genética , Doença de Crohn/microbiologia , Doença de Crohn/patologia , DNA Bacteriano/isolamento & purificação , Enterobacteriaceae/genética , Enterobacteriaceae/imunologia , Enterobacteriaceae/isolamento & purificação , Epigenômica , Feminino , Microbioma Gastrointestinal/genética , Interações entre Hospedeiro e Microrganismos/genética , Humanos , Mucosa Intestinal/diagnóstico por imagem , Mucosa Intestinal/imunologia , Mucosa Intestinal/microbiologia , Mucosa Intestinal/patologia , Masculino , Pessoa de Meia-Idade , RNA Ribossômico 16S/genética , RNA-Seq , Adulto Jovem
3.
Mucosal Immunol ; 9(5): 1193-204, 2016 09.
Artigo em Inglês | MEDLINE | ID: mdl-26813344

RESUMO

A role for the IL-36 family of cytokines has been identified in the pathogenesis of psoriasis. Although significant mechanistic overlap can exist between psoriasis and inflammatory bowel disease (IBD), to date there have been no reports investigating the IL-36 family in gastrointestinal inflammation. Here we demonstrate that expression levels of IL-36α are specifically elevated in the colonic mucosa of ulcerative colitis patients. This elevated expression is mirrored in the inflamed colonic mucosa of mice, wherein IL-36 receptor deficiency confirmed this pathway as a mediator of mucosal inflammation. Il36r-/- mice exhibited reduced disease severity in an acute DSS-induced model of colitis in association with decreased innate inflammatory cell infiltration to the colon lamina propria. Consistent with these data, infection with the enteropathogenic bacteria Citrobacter rodentium, resulted in reduced innate inflammatory cell recruitment and increased bacterial colonization in the colons of il36r-/- mice. Il36r-/- mice also exhibited altered T helper cell responses in this model, with enhanced Th17 and reduced Th1 responses, demonstrating that IL-36R signaling also regulates intestinal mucosal T-cell responses. These data identify a novel role for IL-36 signaling in colonic inflammation and indicate that the IL-36R pathway may represent a novel target for therapeutic intervention in IBD.


Assuntos
Colite Ulcerativa/imunologia , Infecções por Enterobacteriaceae/imunologia , Imunidade nas Mucosas , Interleucina-1/imunologia , Mucosa Intestinal/imunologia , Receptores de Interleucina/imunologia , Adulto , Idoso , Animais , Criança , Citrobacter rodentium/crescimento & desenvolvimento , Citrobacter rodentium/imunologia , Colite/induzido quimicamente , Colite/genética , Colite/imunologia , Colite/patologia , Colite Ulcerativa/genética , Colite Ulcerativa/patologia , Colo/imunologia , Colo/patologia , Sulfato de Dextrana , Infecções por Enterobacteriaceae/genética , Infecções por Enterobacteriaceae/microbiologia , Infecções por Enterobacteriaceae/patologia , Feminino , Regulação da Expressão Gênica , Humanos , Interleucina-1/genética , Mucosa Intestinal/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Pessoa de Meia-Idade , Receptores de Interleucina/genética , Receptores de Interleucina-1/deficiência , Receptores de Interleucina-1/genética , Receptores de Interleucina-1/imunologia , Transdução de Sinais , Células Th1/imunologia , Células Th1/patologia , Células Th17/imunologia , Células Th17/patologia
4.
Clin Exp Immunol ; 173(2): 332-42, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23607276

RESUMO

Bcl-3 is a member of the IκB family of proteins and is an essential negative regulator of Toll-like receptor-induced responses. Recently, a single nucleotide polymorphism associated with reduced Bcl-3 gene expression has been identified as a potential risk factor for Crohn's disease. Here we report that in contrast to the predictions of single nucleotide polymorphism (SNP) analysis, patients with Crohn's disease and ulcerative colitis demonstrate elevated Bcl-3 mRNA expression relative to healthy individuals. To explore further the potential role of Bcl-3 in inflammatory bowel disease (IBD), we used the dextran-sodium sulphate (DSS)-induced model of colitis in Bcl-3(-/-) mice. We found that Bcl-3(-/-) mice were less sensitive to DSS-induced colitis compared to wild-type controls and demonstrated no significant weight loss following treatment. Histological analysis revealed similar levels of oedema and leucocyte infiltration between DSS-treated wild-type and Bcl-3(-/-) mice, but showed that Bcl-3(-/-) mice retained colonic tissue architecture which was absent in wild-type mice following DSS treatment. Analysis of the expression of the proinflammatory cytokines interleukin (IL)-1ß, tumour necrosis factor (TNF)-α and IL-6 revealed no significant differences between DSS-treated Bcl-3(-/-) and wild-type mice. Analysis of intestinal epithelial cell proliferation revealed enhanced proliferation in Bcl-3(-/-) mice, which correlated with preserved tissue architecture. Our results reveal that Bcl-3 has an important role in regulating intestinal epithelial cell proliferation and sensitivity to DSS-induced colitis which is distinct from its role as a negative regulator of inflammation.


Assuntos
Colite/metabolismo , Colo/imunologia , Doença de Crohn/genética , Proteínas Proto-Oncogênicas/metabolismo , Fatores de Transcrição/metabolismo , Animais , Proteína 3 do Linfoma de Células B , Processos de Crescimento Celular/genética , Células Cultivadas , Colite/induzido quimicamente , Colite/genética , Colo/patologia , Citocinas/metabolismo , Sulfato de Dextrana/administração & dosagem , Modelos Animais de Doenças , Células Epiteliais/patologia , Humanos , Mediadores da Inflamação/metabolismo , Leucócitos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Polimorfismo Genético , Proteínas Proto-Oncogênicas/genética , Fatores de Risco , Fatores de Transcrição/genética , Redução de Peso/genética
5.
J Control Release ; 168(1): 28-34, 2013 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-23500058

RESUMO

Inflammatory bowel disease (IBD) is a chronic relapsing inflammation of the gastrointestinal tract. The cytokine TNF-alpha (TNF-α) plays a pivotal role in mediating this inflammatory response. RNA interference (RNAi) holds great promise for the specific and selective silencing of aberrantly expressed genes, such as TNF-α in IBD. The aim of this study was to investigate the efficacy of an amphiphilic cationic cyclodextrin (CD) vector for effective TNF-α siRNA delivery to macrophage cells and to mice with induced acute-colitis. The stability of CD.siRNA was examined by gel electrophoresis in biorelevant media reflecting colonic fluids. RAW264.7 cells were transfected with CD.TNF-α siRNA, stimulated with lipopolysaccharide (LPS) and TNF-α and IL-6 responses were measured by PCR and ELISA. Female C57BL/6 mice were exposed to dextran sodium sulphate (DSS) and treated by intrarectal administration with either CD.siRNA TNF-α or a control solution. In vitro, siRNA in CD nanocomplexes remained intact and stable in both fed and fasted simulated colonic fluids. RAW264.7 cells transfected with CD.TNF-α siRNA and stimulated with LPS displayed a significant reduction in both gene and protein levels of TNF-α and IL-6. CD.TNF-α siRNA-treated mice revealed a mild amelioration in clinical signs of colitis, but significant reductions in total colon weight and colonic mRNA expression of TNF-α and IL-6 compared to DSS-control mice were detected. This data indicates the clinical potential of a local CD-based TNF-α siRNA delivery system for the treatment of IBD.


Assuntos
Colite/tratamento farmacológico , Inativação Gênica , RNA Interferente Pequeno/administração & dosagem , Fator de Necrose Tumoral alfa/genética , beta-Ciclodextrinas/química , Animais , Linhagem Celular , Colite/induzido quimicamente , Colite/metabolismo , Sulfato de Dextrana , Modelos Animais de Doenças , Feminino , Interleucina-6/metabolismo , Lipopolissacarídeos , Camundongos , Camundongos Endogâmicos C57BL , Polietilenoimina/química , RNA Interferente Pequeno/química , Fator de Necrose Tumoral alfa/metabolismo
6.
Mucosal Immunol ; 6(5): 1016-26, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23340823

RESUMO

Natural killer (NK) cells are traditionally considered in the context of tumor surveillance and infection defense but their role in chronic inflammatory disorders such as inflammatory bowel disease is less clear. Here, we investigated the role of NK cells in dextran sodium sulfate (DSS)-induced colitis in mice. Depletion of NK cells impairs the survival of mice with colitis and is linked with dramatic increases in colonic damage, leukocyte infiltration, and pro-inflammatory profiles. Mice depleted of NK cells had increased numbers of neutrophils in colons and mesenteric lymph nodes, compared with control mice, in addition to acquiring a hyper-activation status. In vitro and in vivo studies demonstrate that NK cells downregulate pro-inflammatory functions of activated neutrophils, including reactive oxygen species and cytokine production, by direct cell-to-cell contact involving the NK cell-inhibitory receptor NKG2A. Our results indicate an immunoregulatory mechanism of action of NK cells attenuating DSS-induced colitis neutrophil-mediated inflammation and tissue injury via NKG2A-dependent mechanisms.


Assuntos
Colite/imunologia , Colo/imunologia , Células Matadoras Naturais/imunologia , Subfamília C de Receptores Semelhantes a Lectina de Células NK/metabolismo , Neutrófilos/imunologia , Animais , Células Cultivadas , Colite/induzido quimicamente , Citocinas/metabolismo , Sulfato de Dextrana/administração & dosagem , Mediadores da Inflamação/metabolismo , Depleção Linfocítica , Camundongos , Camundongos Endogâmicos C57BL , Espécies Reativas de Oxigênio/metabolismo
7.
Clin Exp Immunol ; 162(1): 188-96, 2010 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-20718784

RESUMO

Inflammatory bowel disease (IBD) is associated with neutrophil infiltration into the mucosa and crypt abscesses. The chemokine interleukin (IL)-8 [murine homologues (KC) and macrophage inflammatory protein (MIP)-2] and its receptor CXCR2 are required for neutrophil recruitment; thus, blocking this engagement is a potential therapeutic strategy. In the present study, we developed a preclinical model of neutrophil migration suitable for investigating the biology of and testing new drugs that target neutrophil trafficking. Peritoneal exudate neutrophils from transgenic ß-actin-luciferase mice were isolated 12h after intraperitoneal injection with thioglycollate, and were assessed phenotypically and functionally. Exudate cells were injected intravenously into recipients with dextran sodium sulphate (DSS)-induced colitis followed by bioluminescence imaging of whole-body and ex vivo organs at 2, 4 and 16-22h post-transfer. Anti-KC antibody or an isotype control were administered at 20 µg/mouse 1h before transfer, followed by whole-body and organ imaging 4h post-transfer. The peritoneal exudate consisted of 80% neutrophils, 39% of which were CXCR2(+) . In vitro migration towards KC was inhibited by anti-KC. Ex vivo bioluminescent imaging showed that neutrophil trafficking into the colon of DSS recipients was inhibited by anti-KC 4h post-cell transfer. In conclusion, this study describes a new approach for investigating neutrophil trafficking that can be used in preclinical studies to evaluate potential inhibitors of neutrophil recruitment.


Assuntos
Movimento Celular , Colite/metabolismo , Luminescência , Neutrófilos/citologia , Actinas/genética , Actinas/metabolismo , Animais , Quimiotaxia de Leucócito , Colite/induzido quimicamente , Colite/genética , Sulfato de Dextrana , Modelos Animais de Doenças , Feminino , Citometria de Fluxo , Cinética , Luciferases/genética , Luciferases/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Neutrófilos/metabolismo , Receptores de Interleucina-8B/metabolismo
8.
Stress ; 11(5): 348-62, 2008 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-18666024

RESUMO

Inflammatory bowel disease (IBD) is a chronic condition with alternating active and quiescent phases of inflammation. Stress has been suggested as a factor triggering a relapse of IBD. We investigated the role of repetitive psychological stress [water avoidance stress (WAS)] in reactivating colonic inflammation in a murine model of dextran sulfate sodium (DSS)-induced chronic colitis. Colitis was induced in C57BL/6 female mice by exposure to 3% DSS (5 days). During chronic inflammation(day 34), mice underwent repetitive WAS (1 h/day/7 days) and were given a sub-threshold concentration of DSS (1%, 5 days)or normal water to drink. At euthanasia (day 40), inflammatory parameters were assessed (colon inflammatory score, levels of inflammatory markers and histology). Mice with chronic colitis exposed to WAS had higher macroscopic and microscopic colonic inflammatory scores and levels of inflammatory markers (mainly IL-1beta, IL12p40 and CCL5) than non-stressed mice. Inflammatory responses were further enhanced by the presence of a sub-threshold concentration of DSS (1%). In mice without chronic inflammation, neither WAS nor 1% DSS, individually or in combination, elicited any inflammation. Hence stress, per se, reactivates a quiescent chronic inflammation, but does not initiate inflammation in healthy mice. Stress should be regarded as an environmental factor triggering IBD relapses in humans.


Assuntos
Colite/induzido quimicamente , Sulfato de Dextrana , Estresse Psicológico/complicações , Glândulas Suprarrenais/anatomia & histologia , Animais , Aprendizagem da Esquiva , Quimiocina CCL5/sangue , Quimiocina CXCL1/sangue , Colite/patologia , Corticosterona/sangue , Ciclo-Oxigenase 2/metabolismo , Modelos Animais de Doenças , Feminino , Haptoglobinas/metabolismo , Humanos , Doenças Inflamatórias Intestinais/etiologia , Interleucina-12/sangue , Interleucina-1beta/sangue , Interleucina-6/sangue , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico Sintase Tipo II/metabolismo , Tamanho do Órgão , Recidiva , Baço/anatomia & histologia , Timo/anatomia & histologia , Fator de Necrose Tumoral alfa/metabolismo
9.
Eur J Pain ; 12(7): 934-44, 2008 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-18280187

RESUMO

Tetrodotoxin-resistant voltage-gated sodium channels subtype 9 (Na(v)1.9) are expressed in small-diameter dorsal root ganglion neurons and have been involved in persistent somatic hyperalgesic responses associated with inflammation. We assessed the role of Na(v)1.9 channels on acute colonic inflammation-induced visceral hypersensitivity in conscious mice, using Na(v)1.9 knockout (KO) mice. Colorectal distension (CRD)-induced visceral pain was assessed in conscious wild-type and Na(v)1.9 KO mice (C57Bl/6 background). The mechanical activity of the abdominal muscles during isobaric colorectal distension was used as a measure of visceral pain. Acute colonic inflammation was induced by intracolonic administration of the toll-like receptor (TLR) 7 activator, R-848 (40mug/animal). CRD was performed 5h later, thereafter animals were euthanized and the colonic content of inflammatory mediators assessed. Normal pain responses were similar in Na(v)1.9 KO and wild-type mice. In wild-type mice, R-848 administration increased the response to phasic CRD by 62% compared with vehicle-treated animals (vehicle: 0.16+/-0.04, R-848: 0.26+/-0.03, n=6-7, P<0.05). However, in Na(v)1.9 KO mice, intracolonic R-848 did not affect the response to CRD (0.11+/-0.02, n=7) compared to animals treated with vehicle (0.17+/-0.03, n=5; P>0.05). After R-848 administration, the colonic content of pro-inflammatory cytokines was increased in similar proportion in wild type and Na(v)1.9 KO mice, suggesting the presence of a similar acute inflammatory reaction in both groups of animals. These results suggest that Na(v)1.9 channels do not significantly contribute to normal visceral pain responses to acute colonic mechanical stimulation but may be important for the development of inflammation-related acute visceral hyperalgesic responses.


Assuntos
Colite/fisiopatologia , Hiperalgesia/fisiopatologia , Neuropeptídeos/fisiologia , Canais de Sódio/fisiologia , Doença Aguda , Animais , Cateterismo/efeitos adversos , Colite/induzido quimicamente , Colo/química , Feminino , Hiperalgesia/etiologia , Imidazóis/toxicidade , Mediadores da Inflamação/análise , Masculino , Glicoproteínas de Membrana/antagonistas & inibidores , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Canal de Sódio Disparado por Voltagem NAV1.9 , Neuropeptídeos/deficiência , Neuropeptídeos/genética , Reto/química , Recidiva , Canais de Sódio/deficiência , Canais de Sódio/genética , Estresse Mecânico , Receptor 7 Toll-Like/antagonistas & inibidores
10.
Scand J Immunol ; 65(5): 421-9, 2007 May.
Artigo em Inglês | MEDLINE | ID: mdl-17444952

RESUMO

One of the most widely used animal models for inflammatory bowel disease (IBD) is the dextran sulfate sodium (DSS)-induced colitis. We have previously reported that 5 days administration of DSS in C57Bl/6J mice induces a colonic inflammation that progresses into chronicity after DSS removal, whereas in BALB/cJ mice the inflammation resolves within 4 weeks post-DSS. Here we show that both thymic size and thymocyte numbers dramatically decreased in the acute phase of inflammation in C57Bl/6 mice, 7 days after DSS withdrawal. Mature, CD4(+) and CD8(+) single positive (SP) CD69(lo) CD62L(hi) thymocytes were enriched in these mice, accompanied by a major decrease in the number of immature double positive (DP) thymocytes. However, the different maturation stages within the DP thymocyte subset were unchanged between healthy and inflamed C57Bl/6J mice. Interestingly, as the inflammation progressed into the chronic phase, the thymus recovered and 2 weeks after the acute inflammatory phase all the thymic parameters investigated in this study were restored to normal. In contrast, BALB/cJ mice only develop mild thymic alterations. Nevertheless, we found that within the double negative (DN) thymocytes an increased frequency and also total numbers of CD44(+) CD25(-) (DN1) cells correlated with the severity of colitis, and that the frequency of CD44(-) CD25(-) (DN4) thymocytes decreased proportionally in the acute phase in BALB/cJ mice. Our observations suggest that the thymic effects are intimately connected to the intestinal inflammatory response in colitis regardless of the inflammatory stimuli.


Assuntos
Diferenciação Celular/imunologia , Colite/patologia , Sulfato de Dextrana/administração & dosagem , Subpopulações de Linfócitos/patologia , Subpopulações de Linfócitos T/patologia , Timo/patologia , Doença Aguda , Animais , Doença Crônica , Colite/induzido quimicamente , Colite/imunologia , Imunofenotipagem , Subpopulações de Linfócitos/efeitos dos fármacos , Subpopulações de Linfócitos/imunologia , Linfopenia/imunologia , Linfopenia/patologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Índice de Gravidade de Doença , Subpopulações de Linfócitos T/efeitos dos fármacos , Subpopulações de Linfócitos T/imunologia , Timo/efeitos dos fármacos , Timo/imunologia
11.
Scand J Immunol ; 60(1-2): 167-77, 2004.
Artigo em Inglês | MEDLINE | ID: mdl-15238086

RESUMO

Cell-mediated lymphocyte cytotoxicity in ileum and colon of patients with ulcerative colitis (UC), Crohn's disease (CD) and controls was investigated. Frequencies of cells expressing perforin and Fas-ligand (FasL) were determined by immunomorphometry. mRNA expression of perforin, granzyme B and FasL in T cells and subsets was assayed by reverse transcriptase-polymerase chain reaction. Cytotoxicity of intraepithelial and lamina propria lymphocytes was analysed without ex vivo activation in three functional assays: (1) anti-CD3-dependent T-cell receptor (TCR)-/CD3-mediated redirected cytotoxicity, (2) Fas-/FasL-mediated TCR-/CD3-independent cytotoxicity and (3) natural killer (NK) cell cytotoxicity. Inflammation in ileum of CD patients caused increased frequency of perforin-expressing cells and enhanced perforin-dependent TCR-/CD3-mediated cytotoxicity. In contrast, lymphocytes in the inflamed colon of UC or Crohn's colitis patients did not display this cytotoxicity nor did lymphocytes of normal colon. Normal colon lymphocytes showed spontaneous Fas-/FasL-mediated cytotoxicity. This activity was retained but not enhanced in inflamed UC colon. In contrast, a significant increase of FasL-expressing cells was seen in situ. Inflammation did not induce NK cell activity in colonic lymphocytes. Intestinal lymphocytes comprise effectors active in two different cytolytic processes. 'Classical' cytotoxic T lymphocytes in small intestine and lymphocytes executing TCR-/CD3-independent FasL-/Fas-mediated killing of unknown biological role present throughout the intestinal mucosa. Ongoing normal cytolytic processes seem to be enhanced by chronic inflammation.


Assuntos
Colite Ulcerativa/imunologia , Colo/imunologia , Doença de Crohn/imunologia , Íleo/imunologia , Linfócitos T/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Colo/citologia , Testes Imunológicos de Citotoxicidade , Proteína Ligante Fas , Feminino , Granzimas , Humanos , Íleo/citologia , Imuno-Histoquímica , Células Jurkat , Células K562 , Masculino , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/imunologia , Pessoa de Meia-Idade , Perforina , Proteínas Citotóxicas Formadoras de Poros , RNA/química , RNA/genética , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Serina Endopeptidases/genética , Serina Endopeptidases/imunologia
12.
Clin Exp Immunol ; 134(1): 127-37, 2003 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-12974765

RESUMO

Ulcerative colitis (UC), a chronic inflammatory bowel disease, exhibits pronounced increase of T lymphocytes in the inflamed mucosa. To understand the role of intestinal T lymphocytes in the pathogenesis of UC their cytokine production in the mucosa was analysed. Intestinal T lymphocytes of UC, Crohn's disease and control patients were analysed for cytokine mRNA levels by real-time quantitative reverse transcription-polymerase chain reaction (RT-PCR) directly after isolation without in vitro stimulation. Frequencies of cytokine positive cells were determined in UC and control colon by immunomorphometry. T lymphocytes in normal colon expressed interleukin (IL)-2, interferon (IFN)-gamma, tumour necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta1, but not IL-4, IL-5 or IL-10. In UC, a highly significant increase in IL-10 mRNA levels in T lymphocytes and an increased frequency of IL-10 positive cells was seen in colon. IL-10 mRNA levels were also elevated in T lymphocytes of the non-inflamed ileum and correlated with disease activity at both locations. CD4+ T lymphocytes were the major source of IL-10 mRNA. IL-2, IFN-gamma and TNF-alpha mRNA levels were decreased in colonic T lymphocytes, and virtually no IL-2, IFN-gamma, TNF-alpha or TGF-beta positive cells were detected in basal lymphoid aggregates. However, scattered IL-10 positive cells were found here. Lamina propria outside the aggregates contained IL-10-, IFN-gamma, TNF-alpha and TGF-beta but not IL-2 positive cells. T cells of UC patients did not express IL-4 or IL-5. Taken, together the data suggest a generalized activation of IL-10 producing CD4+ T cells along the intestine of UC patients. The local environment seems to determine the biological consequences of elevated IL-10.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Colite Ulcerativa/imunologia , Colo/imunologia , Interleucina-10/imunologia , Mucosa Intestinal/imunologia , Doença Aguda , Adulto , Idoso , Biópsia , Estudos de Casos e Controles , Feminino , Humanos , Imuno-Histoquímica/métodos , Interferon gama/genética , Interleucina-10/genética , Interleucina-2/genética , Interleucina-4/genética , Interleucina-5/genética , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/análise , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fator de Crescimento Transformador beta/genética , Fator de Necrose Tumoral alfa/genética
13.
Gut ; 47(2): 215-27, 2000 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-10896913

RESUMO

BACKGROUND AND AIMS: A histopathological feature considered indicative of ulcerative colitis (UC) is the so-called basal lymphoid aggregates. Their relevance in the pathogenesis of UC is, however, unknown. We have performed a comprehensive analysis of the immune cells in these aggregates most likely corresponding to the lymphoid follicular hyperplasia also described in other colitides. METHODS: Resection specimens of UC and normal colon were analysed by immunomorphometry, immunoflow cytometry, and immunoelectron microscopy, using a large panel of monoclonal antibodies. RESULTS: (1) In all cases of UC, colonic lamina propria contained numerous basal aggregates composed of lymphocytes, follicular dendritic cells, and CD80/B7.1 positive dendritic cells. (2) CD4(+)CD28(-) alphabeta T cells and B cells were the dominant cell types in the aggregates. (3) The aggregates contained a large fraction of cells that are normally associated with the epithelium: that is, gammadelta T cells (11 (7)%) and alpha(E)beta(7)(+) cells (26 (13)%). The gammadelta T cells used Vdelta1 and were CD4(-)CD8(-). Immunoelectron microscopy analysis demonstrated TcR-gammadelta internalisation and surface downregulation, indicating that the gammadelta T cells were activated and engaged in the disease process. (4) One third of cells in the aggregates expressed the antiapoptotic protein bcl-2. CONCLUSIONS: Basal lymphoid aggregates in UC colon are a consequence of anomalous lymphoid follicular hyperplasia, characterised by abnormal follicular architecture and unusual cell immunophenotypes. The aggregates increase in size with severity of disease, and contain large numbers of apoptosis resistant cells and activated mucosal gammadelta T cells. The latter probably colonise the aggregates as an immunoregulatory response to stressed lymphocytes or as a substitute for defective T helper cells in B cell activation. gammadelta T cells in the aggregates may be characteristic of UC.


Assuntos
Linfócitos B , Colite Ulcerativa/imunologia , Regulação da Expressão Gênica , Receptores de Retorno de Linfócitos/genética , Linfócitos T , Adulto , Idoso , Idoso de 80 Anos ou mais , Linfócitos B/citologia , Linfócitos B/imunologia , Colite Ulcerativa/patologia , Células Dendríticas Foliculares/citologia , Regulação para Baixo , Feminino , Citometria de Fluxo , Genes Codificadores da Cadeia delta de Receptores de Linfócitos T/genética , Genes bcl-2 , Humanos , Imunidade Celular , Imunofenotipagem , Masculino , Microscopia Eletrônica , Pessoa de Meia-Idade , Fenótipo , Receptores de Antígenos de Linfócitos T gama-delta/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia
14.
Clin Exp Immunol ; 120(3): 424-33, 2000 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-10844519

RESUMO

Neuroendocrine peptides have a variety of physiological functions in the gastrointestinal tract. This study was carried out to investigate the impact of IL-2 deficiency on the neuroendocrine system in normal colon, and the neuroendocrine changes during colonic inflammation. Mice with homozygous disrupted IL-2 gene (IL-2-/-) spontaneously developed a bowel disease with similarities to human ulcerative colitis. Different types of colonic endocrine cells and myenteric nerves were analysed in the IL-2-/- mice using immunomorphometry. The neuropeptide contents in the colonic tissues were determined by radioimmunoassay. Age-matched healthy IL-2+/- and IL-2+/+ mice served as controls and the colonic IL-2 levels were compared between these two groups of mice by ELISA. Our data showed that less than half the amount of IL-2 was synthesized in the colon of IL-2+/- mice compared with the IL-2+/+ wild-type mice. Two major differences in the neuroendocrine colon were found between the mice with an intact and disrupted IL-2 gene. One was age-related. The frequencies of various endocrine cells and myenteric nerves increased with age in the IL-2+/+ mice. However, no such increases were seen in the mice with a disrupted IL-2 gene. Instead, the volume densities of enteroglucagon, serotonin cells and substance P (SP), vasoactive intestinal polypeptide (VIP) and total myenteric nerves were lower in the older IL-2+/- and IL-2-/- mice compared with the wild type. The other was disease-related. Polypeptide YY (PYY) cells and tissue levels of PYY, SP and VIP were significantly decreased in the IL-2-/- mice during the course of bowel inflammation compared with the healthy IL-2+/- and IL-2+/+ controls. These findings indicate that colonic neuroendocrine alterations did occur in the mice with a disrupted IL-2 gene and diminished local IL-2 level, suggesting a role of IL-2 in the regulation of the neuroendocrine system and a prevalent interaction between the immune and neuroendocrine systems in normal colon. On the other hand, there were some changes that seemed to correlate with the bowel inflammatory process. They might be associated with the impaired function in inflamed gut and contribute to the development and/or prolongation of disease.


Assuntos
Colo/patologia , Interleucina-2/genética , Interleucina-2/fisiologia , Sistemas Neurossecretores/crescimento & desenvolvimento , Animais , Colite/fisiopatologia , Ensaio de Imunoadsorção Enzimática , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Radioimunoensaio
15.
Am J Trop Med Hyg ; 60(5): 740-5, 1999 May.
Artigo em Inglês | MEDLINE | ID: mdl-10344645

RESUMO

For effective control programs, accurate assessment of Trypanosoma cruzi infection in vectors is essential and has traditionally been performed by microscopic examination. For particular vectors and not others, polymerase chain reaction (PCR) analysis of fecal samples recently has been shown to be an effective means of detection. The sensitivities of the PCR and microscopy for detection of T. cruzi in different anatomic sites were compared in the two major vectors of Guatemala, Triatoma dimidiata and Rhodnius prolixus. Preliminary studies established that T. cruzi can be detected by the PCR in the presence of 90% T. rangeli. One hundred thirty-five vectors were collected, and samples were obtained from the rectum, intestines, and stomach and analyzed by microscopy and the PCR. For Triatoma dimidiata rectal samples, the PCR sensitivity (39.1% T. cruzi positive) and the microscopic sensitivity (24.6% positive) was not significantly different. However, in R. prolixus, the PCR proved significantly more sensitive than microscopy: 57.6% positive by PCR compared with 22.7% by microscopy. Rectal samples showed the highest rates of infection followed by intestine and stomach samples. However, 10.5% of the Rhodnius infections would have been missed if only the rectal sample had been analyzed. Thus, the PCR is significantly more sensitive than microscopy for detection of T. cruzi in R. prolixus. Analysis of anatomic sites in addition to the rectal sample may be necessary for accurate assessment of infection in particular vectors.


Assuntos
Doença de Chagas/transmissão , Insetos Vetores/parasitologia , Reação em Cadeia da Polimerase/métodos , Rhodnius/parasitologia , Triatoma/parasitologia , Trypanosoma cruzi/isolamento & purificação , Animais , DNA de Protozoário/análise , Fezes/parasitologia , Guatemala , Intestinos/parasitologia , Reto/parasitologia , Sensibilidade e Especificidade , Estômago/parasitologia , Trypanosoma cruzi/genética
16.
FEMS Microbiol Lett ; 172(1): 29-34, 1999 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-10079524

RESUMO

Representative strains of the different diarrheagenic Escherichia coli virotypes were tested for their potential cytotoxicity in the J774 macrophage cell line. All the seven virotypes of E. coli were cytotoxic to J774 macrophages, and in most cases the bacteria induced an apoptotic response. With the exception of the enterotoxigenic E. coli (ETEC) strain, all the other six virotypes caused induction of apoptosis as evidenced by quantitative analysis of the characteristic DNA fragmentation at the individual cell level. These results suggest that apoptosis could be one of the mechanisms contributing to the diarrheal disease development.


Assuntos
Apoptose , Escherichia coli/patogenicidade , Macrófagos/microbiologia , Animais , Linhagem Celular , Fragmentação do DNA , Diarreia/microbiologia , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Humanos , Macrófagos/fisiologia , Camundongos
17.
J Immunol ; 157(5): 1926-34, 1996 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-8757311

RESUMO

The functional properties of intraepithelial lymphocytes (IEL) in normal human jejunum, ileum, and colon were investigated. Cytokine mRNA expression in IEL and enterocytes was determined by reverse transcriptase-PCR and IFN-gamma+ IEL by immunohistochemistry. Polyclonal activators were used to study proliferation and IFN-gamma secretion of IEL, and an anti-CD3-mediated redirected cytotoxicity assay was used to determine the lytic potential of IEL. Freshly isolated IEL at all three gut levels expressed mRNA for IL-1 beta, IL-2, IL-8, IFN-gamma, and TNF-alpha. Approximately 10% of IEL produced IFN-gamma, suggesting that IEL are immunologically active in vivo, performing similar functions along the intestine. IEL could be stimulated further in vitro to express IL-10, TNF-beta, and TGF-beta 1, while no Th2-type cytokines were induced, suggesting suppressive and cytolytic functions for IEL. All three jejunal IEL subpopulations (CD4-CD8-TCR-gamma delta+, CD4+TCR-alpha beta+, CD8+TCR-alpha beta+) expressed the same four cytokines, IL-2, IL-8, IFN-gamma, and TNF-alpha, indicating that CD4+TCR-alpha beta+ IEL are Th1 cells and that TCR-gamma delta+ IEL and CD8+TCR-alpha beta+ IEL include cytotoxic effector cells. Indeed, freshly isolated jejunal IEL displayed cytolytic activity. IEL were induced to proliferation by anti-CD3/TCR complex mAbs and leukoagglutinin, but not by Con A. There was no correlation between the magnitude of the proliferative response and the amounts of secreted IFN-gamma. Enterocytes expressed IL-1 beta and IL-8, and sometimes TNF-alpha. Although jejunal enterocytes express HLA-DR and hsp60, Ag presentation by these cells may induce anergy since their cytokine profile is different from that of classical APCs.


Assuntos
Citocinas/biossíntese , Citotoxicidade Imunológica , Mucosa Intestinal/imunologia , Linfócitos T Citotóxicos/imunologia , Células Th1/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Células Cultivadas , Colo/imunologia , Colo/metabolismo , Citocinas/genética , Células Epiteliais , Epitélio/imunologia , Epitélio/metabolismo , Feminino , Humanos , Íleo/imunologia , Íleo/metabolismo , Interferon gama/genética , Interferon gama/metabolismo , Interleucina-1/genética , Interleucina-10/genética , Interleucina-2/genética , Interleucina-8/genética , Mucosa Intestinal/citologia , Mucosa Intestinal/metabolismo , Jejuno/imunologia , Jejuno/metabolismo , Lectinas/farmacologia , Linfotoxina-alfa/genética , Masculino , Pessoa de Meia-Idade , RNA Mensageiro/biossíntese , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T Citotóxicos/metabolismo , Células Th1/metabolismo , Fator de Crescimento Transformador beta/genética , Células Tumorais Cultivadas , Fator de Necrose Tumoral alfa/genética
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