RESUMO
Evidence suggests that CD4+CD25+ regulatory T cells play a crucial role in the suppression of intestinal inflammation. However, their role in the suppression of inflammatory bowel disease has not yet been addressed. We examined the proportion of regulatory T cells in inflammatory bowel disease. First, we isolated CD4+CD45RO+CD25+ T cells from the peripheral blood of healthy persons and showed that these cells suppressed T cell proliferation profoundly and expressed FoxP3 abundantly, revealing that they are regulatory cells. Then the proportion of CD45RO+CD25+ in peripheral blood CD4+ T cells was analyzed in patients and healthy controls by flow cytometry. CD4+CD45RO+CD25+ T cell frequency was significantly lower in active ulcerative colitis than in the control and inactive ulcerative colitis. CD4+CD45RO+CD25+ T cell frequency was inversely correlated with the clinical and endoscopic severity of ulcerative colitis. These results suggest that a deficiency of regulatory T cells is associated with the progression of ulcerative colitis.
Assuntos
Colite Ulcerativa/sangue , Colite Ulcerativa/fisiopatologia , Linfócitos T Reguladores/imunologia , Adolescente , Adulto , Sequência de Bases , Biomarcadores/análise , Linfócitos T CD4-Positivos/imunologia , Estudos de Casos e Controles , Células Cultivadas , Progressão da Doença , Feminino , Humanos , Antígenos Comuns de Leucócito/imunologia , Masculino , Pessoa de Meia-Idade , Dados de Sequência Molecular , Probabilidade , Receptores de Interleucina-2/imunologia , Valores de Referência , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Sensibilidade e Especificidade , Índice de Gravidade de DoençaRESUMO
Mutations in the adenomatous polyposis coli (APC) gene and K-ras occur in the majority of human colorectal cancers. Loss of functional APC protein activates the Wnt signal transduction pathway, allowing the nuclear accumulation of beta-catenin, which then binds to T-cell factor-4 (Tcf-4), causing increased transcriptional activation of downstream target genes. We investigated the hypothesis that the activation of the WNT pathway regulates cyclooxygenase-2 (COX-2). COX-2 was down-regulated after the induction of full-length APC in the HT29-APC cell line. We identified a Tcf-4-binding element (TBE) in the COX-2 promoter that specifically bound to Tcf-4 in an electrophoretic mobility shift assay. COX-2 promoter luciferase activity is down-regulated by APC in a promoter reporter construct containing the, TBE but not with mutant TBE. Mutant beta-catenin expression up-regulated the COX-2 promoter activity and the endogenous COX-2 mRNA expression in HuH7, hepatocellular carcinoma cell line, which is partially abrogated by cotransfection with a dominant-negative Tcf-4 expression vector. Although beta-catenin alone did not increase COX-2 protein to detectable levels in HuH7 cells, coexpression of both mutant beta-catenin and mutant K-ras increased COX-2 protein expression, which is consistent with the previous reports that K-ras can stabilize COX-2 mRNA. Taken together, our data support the hypothesis that COX-2 is down-regulated by APC and up-regulated by nuclear beta-catenin accumulation, and additionally implicate the Wnt signal transduction pathway in colon and liver carcinogenesis.
Assuntos
Isoenzimas/biossíntese , Prostaglandina-Endoperóxido Sintases/biossíntese , Proteínas Proto-Oncogênicas/fisiologia , Proteínas de Peixe-Zebra , Proteínas ras/fisiologia , Proteína da Polipose Adenomatosa do Colo/biossíntese , Proteína da Polipose Adenomatosa do Colo/genética , Proteína da Polipose Adenomatosa do Colo/fisiologia , Ciclo-Oxigenase 2 , Proteínas do Citoesqueleto/fisiologia , Proteínas de Ligação a DNA/metabolismo , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Células HT29 , Humanos , Isoenzimas/genética , Luciferases/genética , Luciferases/metabolismo , Proteínas de Membrana , Regiões Promotoras Genéticas , Prostaglandina-Endoperóxido Sintases/genética , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Proteínas Recombinantes de Fusão/metabolismo , Transdução de Sinais/fisiologia , Fatores de Transcrição TCF , Transativadores/fisiologia , Proteína 2 Semelhante ao Fator 7 de Transcrição , Fatores de Transcrição/metabolismo , Transcrição Gênica , Regulação para Cima , Proteínas Wnt , beta CateninaRESUMO
Free radical-induced cellular stress contributes to cancer during chronic inflammation. Here, we investigated mechanisms of p53 activation by the free radical, NO. NO from donor drugs induced both ataxia-telangiectasia mutated (ATM)- and ataxia-telangiectasia mutated and Rad3-related-dependent p53 posttranslational modifications, leading to an increase in p53 transcriptional targets and a G(2)M cell cycle checkpoint. Such modifications were also identified in cells cocultured with NO-releasing macrophages. In noncancerous colon tissues from patients with ulcerative colitis (a cancer-prone chronic inflammatory disease), inducible NO synthase protein levels were positively correlated with p53 serine 15 phosphorylation levels. Immunostaining of HDM-2 and p21(WAF1) was consistent with transcriptionally active p53. Our study highlights a pivotal role of NO in the induction of cellular stress and the activation of a p53 response pathway during chronic inflammation.
