RESUMO
BACKGROUND: Colorectal cancer (CRC) is the third most lethal cancer in the world, and its incidence is steadily rising. In this study, we investigated the induction of humoral immunity by a phytogalactolipid enriched fraction (CRA) derived from the medicinal plant Crassocephalum rabens (Benth.) S. Moore to combat CRC. METHODS: Immunocompetent BALB/c mice were used to evaluate CRA's therapeutic effects in CRC. The phenotypes of B cell subsets in splenocytes and tumors from the CRA-treated mice were isolated and analyzed by flow cytometry. The titers, isotypes, specificity, antigen recognition, and cytotoxic activity of CRA-induced anti-tumor antibodies were determined. The mechanisms of CRA on B cell differentiation were determined by cell-based analyses, including co-cultural with T cells, cytokine analysis, gene expression by qPCR, and protein expression by western blotting. RESULTS: CRA efficiently inhibited tumor growth in colorectal tumor-bearing allograft mice. CRA treatment attracted an abundance of B cells into the tumor consequently enhancing the anti-tumor antibodies in sera and inducing a class-switch. CRA-induced antisera (designated CRA antisera) specifically recognized surface antigens on the plasma membrane of cancer cells. CRA antisera induced cytotoxicity including antibody-dependent cell cytotoxicity, phagocytosis, and complement-dependent cytotoxicity. CRA interacted with IL-6 receptor to activate STAT3 and cMaf, resulting in T cell secretion of IL-21, which, in turn induced B cell differentiation through the IL-21R/STAT3/Blimp-1 pathway. CONCLUSIONS: CRA regulated T cell activity resulting in B cell activation and triggering of anti-tumor antibodies to impede CRC progression.
Assuntos
Antineoplásicos , Neoplasias Colorretais , Camundongos , Animais , Imunidade Humoral , Neoplasias Colorretais/patologia , Antineoplásicos/farmacologia , Citocinas , Soros ImunesRESUMO
Medulloblastoma (MB) is the most common malignant brain tumor in children. It is classified into core molecular subgroups (wingless activated (WNT), sonic hedgehog activated (SHH), Group 3 (G3), and Group 4 (G4)). In this study, we analyzed the tumor-infiltrating immune cells and cytokine profiles of 70 MB patients in Taiwan using transcriptome data. In parallel, immune cell composition in tumors from the SickKids cohort dataset was also analyzed to confirm the findings. The clinical cohort data showed the WNT and G4 MB patients had lower recurrence rates and better 5-year relapse-free survival (RFP) compared with the SHH and G3 MB patients, among the four subgroups of MB. We found tumor-infiltrating B cells (TIL-Bs) enriched in the G4 subgroups in the Taiwanese MB patients and the SickKids cohort dataset. In the G4 subgroups, the patients with a high level of TIL-Bs had better 5-year overall survival. Mast cells presented in G4 MB tumors were positively correlated with TIL-Bs. Higher levels of CXCL13, IL-36γ, and CCL27 were found compared to other subgroups or normal brains. These three cytokines, B cells and mast cells contributed to the unique immune microenvironment in G4 MB tumors. Therefore, B-cell enrichment is a G4-subgroup-specific immune signature and the presence of B cells may be an indicator of a better prognosis in G4 MB patients.
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Neoplasias Cerebelares , Meduloblastoma , Neoplasias Cerebelares/genética , Neoplasias Cerebelares/patologia , Criança , Proteínas Hedgehog/genética , Humanos , Meduloblastoma/genética , Meduloblastoma/patologia , Recidiva Local de Neoplasia , Transcriptoma , Microambiente Tumoral/genéticaRESUMO
BACKGROUND: Inhibiting TNF-α is an effective therapy for inflammatory diseases such as rheumatoid arthritis. However, systemic, nondiscriminatory neutralization of TNF-α is associated with considerable adverse effects. METHODS: Here, we developed a trimeric chimeric TNF receptor by linking an N-terminal mouse Acrp30 trimerization domain and an MMP-2/9 substrate sequence to the mouse extracellular domain of TNF receptor 2 followed by a C-terminal mouse tetranectin coiled-coil domain (mouse Acrp-MMP-TNFR-Tn). RESULTS: Here, we show that the Acrp30 trimerization domain inhibited the binding activity of TNFR, possibly by closing the binding site of the trimeric receptor. Cleavage of the substrate sequence by MMP-9, an enzyme highly expressed in inflammatory sites, restored the binding activity of the mouse TNF receptor. We also constructed a recombinant human chimeric TNF receptor (human Acrp-MMP-TNFR-Tn) in which an MMP-13 substrate sequence was used to link the human Acrp and the human TNF receptor 2. Human Acrp-MMP-TNFR-Tn showed reduced binding activity, and MMP-13 digestion recovered its binding activity with TNF-α. CONCLUSION: Acrp-masked chimeric TNF receptors may be able to be used for inflammatory tissue-selective neutralization of TNF-α to reduce the adverse effects associated with systemic neutralization of TNF-α.
Assuntos
Adiponectina , Metaloproteinase 13 da Matriz , Metaloproteinase 2 da Matriz , Metaloproteinase 9 da Matriz , Multimerização Proteica , Receptores Tipo II do Fator de Necrose Tumoral , Proteínas Recombinantes de Fusão , Fator de Necrose Tumoral alfa , Adiponectina/química , Adiponectina/genética , Adiponectina/metabolismo , Animais , Linhagem Celular , Humanos , Metaloproteinase 13 da Matriz/química , Metaloproteinase 13 da Matriz/genética , Metaloproteinase 13 da Matriz/metabolismo , Metaloproteinase 2 da Matriz/química , Metaloproteinase 2 da Matriz/genética , Metaloproteinase 2 da Matriz/metabolismo , Metaloproteinase 9 da Matriz/química , Metaloproteinase 9 da Matriz/genética , Metaloproteinase 9 da Matriz/metabolismo , Camundongos , Especificidade de Órgãos , Ligação Proteica , Domínios Proteicos , Receptores Tipo II do Fator de Necrose Tumoral/química , Receptores Tipo II do Fator de Necrose Tumoral/genética , Receptores Tipo II do Fator de Necrose Tumoral/metabolismo , Fator de Necrose Tumoral alfa/química , Fator de Necrose Tumoral alfa/metabolismoRESUMO
BACKGROUND: Gastric cancer is currently the fourth leading cause of cancer-related death worldwide. Gastric cancer is often diagnosed at advanced stages and the outcome of the treatment is often poor. Therefore, identifying new therapeutic targets for this cancer is urgently needed. Integrin alpha 2 (ITGA2) subunit and the beta 1 subunit form a heterodimer for a transmembrane receptor for extracellular matrix, is an important molecule involved in tumor cell proliferation, survival and migration. Integrin α2ß1 is over-expressed on a variety of cancer cells, but is low or absent in most normal organs and resting endothelial cells. RESULTS: In this report, we assessed the ITGA2 as the potential therapeutic target with the bioinformatics tools from the TCGA dataset in which composed of 375 gastric cancer tissues and 32 gastric normal tissues. According to the information from the Cancer Cell Line Encyclopedia (CCLE) database, the AGS cell line with ITGA2 high expression and the SUN-1 cell line with low expression were chosen for the further investigation. Interestingly, the anti-ITGA2 antibody (at 3 µg/ml) inhibited approximately 50% survival of the AGS cells (over-expressed ITGA2), but had no effect in SNU-1 cells (ITGA2 negative). The extents of antibody-mediated cancer inhibition positively correlated with the expression levels of the ITGA2. We further showed that the anti-ITGA2 antibody induced apoptosis by up-regulating the RhoA-p38 MAPK signaling to promote the expressions of Bim, Apaf-1 and Caspase-9, whereas the expressions of Ras and Bax/Bcl-2 were not affected. Moreover, blocking ITGA2 by the specific antibody at lower doses also inhibited cell migration of gastric cancer cells. Blockade of ITGA2 by a specific antibody down-regulated the expression of N-WASP, PAK and LIMK to impede actin organization and cell migration of gastric cancer cells. CONCLUSIONS: Here, we showed that the mRNA expression levels of ITGA2 comparing to normal tissues significantly increased. In addition, the results revealed that targeting integrin alpha 2 subunit by antibodies did not only inhibit cell migration, but also induce apoptosis effect on gastric cancer cells. Interestingly, higher expression level of ITGA2 led to significant effects on apoptosis progression during anti-ITGA2 antibody treatment, which indicated that ITGA2 expression levels directly correlate with their functionality. Our findings suggest that ITGA2 is a potential therapeutic target for gastric cancer.
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Local Treg responses are involved in Helicobacter pylori-related inflammation and clinical outcomes after infection, and H. pylori-derived HSP60 (HpHSP60) is an important virulence factor associated with gastric carcinogenesis. This study to investigate the role of HpHSP60 in immunosuppression, particularly with regard to whether it could induce the production of Treg cells. For this purpose, human peripheral blood mononuclear cells (PBMCs) were treated with or without HpHSP60 in the presence of an anti-CD3 mAb to determine the effect of HpHSP60 on cell proliferation. In this report, HpHSP60 decreased the expression of CDK4 to significantly arrest the proliferation of mitogen-stimulated T-cells, which correlated with the induction of Treg cells. Moreover, monocytic cells were essential for the induction of HpHSP60-induced Treg cells via the secretion of IL-10 and TGF-ß after treatment with HpHSP60. Blockage of HpHSP60 with specific monoclonal antibodies significantly reduced the colonization of H. pylori and the expression of Treg cells in vivo. Overall, our results suggest that HpHSP60 could act on macrophages to trigger the expression of IL-10 and TGF-ß, thereby leading to an increase in Treg cells and inhibition of T-cell proliferation.
Assuntos
Chaperonina 60/metabolismo , Chaperonina 60/farmacologia , Helicobacter pylori/metabolismo , Linfócitos T Reguladores/efeitos dos fármacos , Fatores de Virulência/imunologia , Fatores de Virulência/metabolismo , Animais , Complexo CD3/imunologia , Morte Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Chaperonina 60/genética , Chaperonina 60/imunologia , Quinase 4 Dependente de Ciclina/metabolismo , Citocinas/metabolismo , Feminino , Mucosa Gástrica/imunologia , Mucosa Gástrica/patologia , Regulação Bacteriana da Expressão Gênica , Infecções por Helicobacter/imunologia , Helicobacter pylori/genética , Helicobacter pylori/patogenicidade , Humanos , Terapia de Imunossupressão , Inflamação , Interleucina-10/metabolismo , Interleucina-8/metabolismo , Leucócitos Mononucleares/efeitos dos fármacos , Leucócitos Mononucleares/imunologia , Camundongos Endogâmicos BALB C , Monócitos/efeitos dos fármacos , Monócitos/metabolismo , Linfócitos T Reguladores/imunologia , Células THP-1 , Fator de Crescimento Transformador beta/metabolismoRESUMO
BACKGROUND: Tamoxifen is currently used for the treatment of both early and advanced estrogen receptor (ER) positive breast cancer in pre- and post-menopausal women. However, using tamoxifen routinely to inhibit endogenous or exogenous estrogen effects is occasionally difficult because of its potential side effects. OBJECTIVES: The aim of this study is to design a local drug delivery system to encapsulate tamoxifen for observing their efficacy of skin penetration, drug accumulation and cancer therapy. METHODS: A cationic liposome-PEG-PEI complex (LPPC) was used as a carrier for the encapsulation of tamoxifen and forming 'LPPC/TAM' for transdermal release. The cytotoxicity of LPPC/TAM was analyzed by MTT. The skin penetration, tumor growth inhibition and organ damages were measured in xenograft mice following transdermal treatment. RESULTS: LPPC/TAM had an average size less than 270 nm and a zeta-potential of approximately 40 mV. LPPC/TAM displayed dramatically increased the cytotoxic activity in all breast cancer cells, especially in ER-positive breast cancer cells. In vivo, LPPC drug delivery helped the fluorescent dye penetrating across the skim and accumulating rapidly in tumor area. Administration of LPPC/TAM by transdermal route inhibited about 86 % of tumor growth in mice bearing BT474 tumors. This local treatment of LPPC/TAM did not injury skin and any organs. CONCLUSION: LPPC-delivery system provided a better skin penetration and drug accumulation and therapeutic efficacy. Therefore, LPPC/TAM drug delivery maybe a useful transdermal tool of drugs utilization for breast cancer therapy.
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Neoplasias da Mama/tratamento farmacológico , Lipossomos/administração & dosagem , Tamoxifeno/administração & dosagem , Administração Cutânea , Animais , Mama/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sistemas de Liberação de Medicamentos/métodos , Feminino , Células HEK293 , Humanos , Células MCF-7 , Camundongos , Camundongos Nus , Polietilenoglicóis/administração & dosagem , Polietilenoimina/administração & dosagem , Polietilenoimina/análogos & derivadosRESUMO
MicroRNAs (miRNAs) are small non-coding RNAs of approximately 22 nucleotides, which negatively regulate the gene expression at the post-transcriptional level. This study describes an update of the miRTarBase (http://miRTarBase.mbc.nctu.edu.tw/) that provides information about experimentally validated miRNA-target interactions (MTIs). The latest update of the miRTarBase expanded it to identify systematically Argonaute-miRNA-RNA interactions from 138 crosslinking and immunoprecipitation sequencing (CLIP-seq) data sets that were generated by 21 independent studies. The database contains 4966 articles, 7439 strongly validated MTIs (using reporter assays or western blots) and 348 007 MTIs from CLIP-seq. The number of MTIs in the miRTarBase has increased around 7-fold since the 2014 miRTarBase update. The miRNA and gene expression profiles from The Cancer Genome Atlas (TCGA) are integrated to provide an effective overview of this exponential growth in the miRNA experimental data. These improvements make the miRTarBase one of the more comprehensively annotated, experimentally validated miRNA-target interactions databases and motivate additional miRNA research efforts.
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Bases de Dados de Ácidos Nucleicos , MicroRNAs/metabolismo , RNA Mensageiro/metabolismo , Doença/genética , Perfilação da Expressão Gênica , Humanos , RNA Mensageiro/química , Análise de Sequência de RNARESUMO
MicroRNAs (miRNAs) are small non-coding RNA molecules capable of negatively regulating gene expression to control many cellular mechanisms. The miRTarBase database (http://mirtarbase.mbc.nctu.edu.tw/) provides the most current and comprehensive information of experimentally validated miRNA-target interactions. The database was launched in 2010 with data sources for >100 published studies in the identification of miRNA targets, molecular networks of miRNA targets and systems biology, and the current release (2013, version 4) includes significant expansions and enhancements over the initial release (2010, version 1). This article reports the current status of and recent improvements to the database, including (i) a 14-fold increase to miRNA-target interaction entries, (ii) a miRNA-target network, (iii) expression profile of miRNA and its target gene, (iv) miRNA target-associated diseases and (v) additional utilities including an upgrade reminder and an error reporting/user feedback system.
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Bases de Dados de Ácidos Nucleicos , MicroRNAs/metabolismo , RNA Mensageiro/metabolismo , Doença/genética , Regulação da Expressão Gênica , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Internet , MicroRNAs/química , RNA Mensageiro/química , Análise de Sequência de RNA , TranscriptomaRESUMO
The increased adhesion of monocytes to injured endothelial layers is a critical early event in atherogenesis. Under inflammatory conditions, there is increased expression of specific cell adhesion molecules on activated vascular endothelial cells, which increases monocyte adhesion. In our current study, we demonstrate a putative mechanism for the anti-inflammatory effects of carnosol, a diterpene derived from the herb rosemary. Our results show that both carnosol and rosemary essential oils inhibit the adhesion of TNFalpha-induced monocytes to endothelial cells and suppress the expression of ICAM-1 at the transcriptional level. Moreover, carnosol was found to exert its inhibitory effects by blocking the degradation of the inhibitory protein IkappaBalpha in short term pretreatments but not in 12 h pretreatments. Our data show that carnosol reduces IKK-beta phosphorylation in pretreatments of less than 3 h. In TNFalpha-treated ECs, NF-kappaB nuclear translocation and transcriptional activity was abolished by up to 12 h of carnosol pretreatment and this was blocked by Nrf-2 siRNA. The long-term inhibitory effects of carnosol thus appear to be mediated through its induction of Nrf-2-related genes. The inhibition of ICAM-1 expression and p65 translocation is reversed by HO-1 siRNA. Carnosol also upregulates the Nrf-2-related glutathione synthase gene and thereby increases the GSH levels after 9 h of exposure. Treating ECs with a GSH synthesis inhibitor, BSO, blocks the inhibitory effects of carnosol. In addition, carnosol increases p65 glutathionylation. Hence, our present findings indicate that carnosol suppresses TNFalpha-induced singling pathways through the inhibition of IKK-beta activity or the upregulation of HO-1 expression. The resulting GSH levels are dependent, however, on the length of the carnosol pretreatment period.