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Peroxiredoxin 3 (PRDX3), a mitochondrial hydrogen peroxide scavenger, is known to be upregulated during tumorigenesis and cancer progression. In this study, we provide evidence for the first time that PRDX3 could regulate cellular signaling pathways associated with Matrix Metalloproteinase-1 (MMP-1) expression and activity in breast cancer progression. We show that shRNA-mediated gene silencing of PRDX3 inhibits cell migration and invasion in two triple-negative breast cancer cell lines. Reciprocal experiments show that PRDX3 overexpression promotes invasion and migration of the cancer cells, processes which are important in the metastatic cascade. Notably, this phenomenon may be attributed to the activation of MMP-1, which is observed to be upregulated by PRDX3 in the breast cancer cells. Moreover, immunohistochemical staining of breast cancer tissues revealed a positive correlation between PRDX3 and MMP-1 expression in both epithelial and stromal parts of the tissues. Further pathway reporter array and luciferase assay demonstrated that activation of ERK signaling is responsible for the transcriptional activation of MMP-1 in PRDX3-overexpressed cells. These findings suggest that PRDX3 could mediate cancer spread via ERK-mediated activation of MMP-1. Targeted inhibition of ERK signaling may be able to inhibit tumor metastasis in triple-negative breast cancer.
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PURPOSE: Bladder cancer (BCa) is one of the most common cancer types worldwide and is characterized by a high rate of recurrence. In previous studies, we and others have described the functional influence of plasminogen activator inhibitor-1 (PAI1) in bladder cancer development. While polymorphisms in PAI1 have been associated with increased risk and worsened prognosis in some cancers, the mutational status of PAI1 in human bladder tumors has not been well defined. METHODS: In this study, we evaluated the mutational status of PAI1 in a series of independent cohorts, comprised of a total of 660 subjects. RESULTS: Sequencing analyses identified two clinically relevant 3' untranslated region (UTR) single nucleotide polymorphisms (SNPs) in PAI1 (rs7242; rs1050813). Somatic SNP rs7242 was present in human BCa cohorts (overall incidence of 72%; 62% in Caucasians and 72% in Asians). In contrast, the overall incidence of germline SNP rs1050813 was 18% (39% in Caucasians and 6% in Asians). Furthermore, Caucasian patients with at least one of the described SNPs had worse recurrence-free survival and overall survival (p = 0.03 and p = 0.03, respectively). In vitro functional studies demonstrated that SNP rs7242 increased the anti-apoptotic effect of PAI1, and SNP rs1050813 was related to a loss of contact inhibition associated with cellular proliferation when compared to wild type. CONCLUSION: Further investigation of the prevalence and potential downstream influence of these SNPs in bladder cancer is warranted.
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Inibidor 1 de Ativador de Plasminogênio , Polimorfismo de Nucleotídeo Único , Neoplasias da Bexiga Urinária , Humanos , Recidiva Local de Neoplasia , Inibidor 1 de Ativador de Plasminogênio/genética , Neoplasias da Bexiga Urinária/genéticaRESUMO
Gene Ontology (GO) analysis can provide a comprehensive function analysis for investigating genes, allowing us to identify the potential biological roles of genes. The present study conducted GO analysis to explore the biological function of IRAK2 and performed a case analysis to define its clinical role in disease progression and mediating tumor response to RT. Methods: We performed a GO enrichment analysis on the RNA-seq data to validate radiation-induced gene expression. A total of 172 I-IVB specimens from oral squamous cell carcinoma patients were collected for clinical analysis, from which IRAK2 expression was analyzed by immunohistochemistry. This was a retrospective study conducted between IRAK2 expression and the outcomes of oral squamous cell carcinoma patients after radiotherapy treatment. We conducted Gene Ontology (GO) analysis to explore the biological function of IRAK2 and performed a case analysis to define its clinical role in mediating tumor response to radiotherapy. GO enrichment analysis to validate radiation-induced gene expression was performed. Clinically, 172 stage I-IVB resected oral cancer patients were used to validate IRAK2 expression in predicting clinical outcomes. GO enrichment analysis showed that IRAK2 is involved in 10 of the 14 most enriched GO categories for post-irradiation biological processes, focusing on stress response and immune modulation. Clinically, high IRAK2 expression was correlated with adverse disease features, including pT3-4 status (p = 0.01), advanced overall stage (p = 0.02), and positive bone invasion (p = 0.01). In patients who underwent radiotherapy, the IRAK2-high group was associated with reduced post-irradiation local recurrence (p = 0.025) compared to the IRAK2-low group. IRAK2 plays a crucial role in the radiation-induced response. Patients with high IRAK2 expression demonstrated more advanced disease features but predicted higher post-irradiation local control in a clinical setting. These findings support IRAK2 as a potential predictive biomarker for radiotherapy response in non-metastatic and resected oral cancer patients.
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Carcinoma de Células Escamosas , Neoplasias de Cabeça e Pescoço , Neoplasias Bucais , Humanos , Neoplasias Bucais/genética , Neoplasias Bucais/radioterapia , Neoplasias Bucais/patologia , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/radioterapia , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas de Cabeça e Pescoço , Estudos RetrospectivosRESUMO
Secondary hyperparathyroidism (SHPT) is common in end-stage renal disease (ESRD) patients, and it can suppress erythropoiesis. We aimed to investigate the relationship between the consumption of erythropoiesis-stimulating agents (ESAs) and parathyroidectomy (PTX) in ESRD patients with SHPT and to determine the predictors for anemia improvement. The current standard of chronic kidney disease anemia therapy relies on the prescription of iron supplementation, and ESA. We retrospectively analyzed 81 ESRD patients with PTX at Ditmanson Medical Foundation Chiayi Christian Hospital from July 2004 to Dec 2018. The requirement of ESA therapy markedly declined from a dose of 41.6 (interquartile range [IQR], 0−91.2) to 10.3 (IQR, 0−59.5, p = 0.001) unit/kg/week. In addition, 63.7% of patients required iron replacement therapy preoperatively and the proportion reduced to 52.5% after PTX (p < 0.001). The hemoglobin (Hb) level showed an insignificant change from a median value of 10.7 g/dL (9.5−11.6 g/dL) before PTX to 10.5 g/dL (9.6−11.2 g/dL) at 6 months after PTX. A preoperative Hb level ≤ 10 mg/dL (odds ratio [OR], 20.1; 95% confidence interval [CI], 4.71−125, p < 0.001) and transferrin saturation (TSAT) < 25% (OR, 12.8; 95% CI, 2.51−129, p < 0.001) were predictors for anemia improvement. Our study demonstrated that PTX markedly decreased the requirement of ESA. Patients with a low preoperative Hb level or low TSAT showed an increase in the Hb level after PTX. PTX may be considered not only for SHPT with refractory anemia but also for high ESA-dependent patients.
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Anemia , Hematínicos , Hiperparatireoidismo Secundário , Falência Renal Crônica , Anemia/tratamento farmacológico , Eritropoese , Hematínicos/uso terapêutico , Hemoglobinas/metabolismo , Humanos , Hiperparatireoidismo Secundário/tratamento farmacológico , Hiperparatireoidismo Secundário/cirurgia , Ferro/uso terapêutico , Falência Renal Crônica/tratamento farmacológico , Falência Renal Crônica/terapia , Paratireoidectomia , Diálise Renal , Estudos Retrospectivos , Transferrinas/uso terapêuticoRESUMO
MicroRNAs (miRNAs) play an important role in gene regulation by degradation or translational inhibition of the targeted mRNAs. It has been experimentally shown that the way miRNAs interact with their targets can be used to explain the indirect interactions among their targets, i.e., competing endogenous RNA (ceRNA). However, whether the protein translated from the targeted mRNAs can play any role in this ceRNA network has not been explored. Here we propose a deterministic model to demonstrate that in a network of one miRNA interacting with multiple-targeted mRNAs, the competition between miRNA-targeted mRNAs is not sufficient for the significant change of those targeted mRNA levels, while dramatic changes of these miRNA-targeted mRNAs require transcriptional inhibition of miRNA by its target proteins. When applied to estrogen receptor signaling pathways, the miR-193a targets E2F6 (a target of estrogen receptor), c-KIT (a marker for cancer stemness), and PBX1 (a transcriptional activator for immunosuppressive cytokine, IL-10) in ovarian cancer, such that epigenetic silencing of miR-193a by E2F6 protein is required for the significant change of c-KIT and PBX1 mRNA level for cancer stemness and immunoevasion, respectively, in ovarian cancer carcinogenesis.
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Epigênese Genética/genética , Estrogênios/genética , Redes Reguladoras de Genes/genética , MicroRNAs/genética , Neoplasias Ovarianas/genética , RNA Longo não Codificante/genética , RNA Mensageiro/genética , Carcinogênese/genética , Linhagem Celular Tumoral , Epigenômica/métodos , Feminino , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Transdução de Sinais/genéticaRESUMO
BACKGROUND: Urothelial carcinoma (UC) is the second most common malignancy of the urinary system with high rate of recurrence, UC patients therefore needed to be treated with surgery followed by chemotherapy. Development of novel therapeutics with minimal side-effect is an urgent issue. Our previous study showed that cyproheptadine (CPH), an anti-histamine, exhibited antitumor activity in UC in vitro and in an xenograft model. However, the molecular mechanism of how CPH inhibits tumor progression is not fully understood. METHODS: Genes that were upregulated after treatment with CPH in UC cells, were examined by RNA-Seq. Real-time quantitative PCR (RT-qPCR) was employed to detect IRF6 expression while COBRA assay and bisulphite pyrosequencing were used to examine promoter methylation of IRF6. Enrichment of total H3K27 acetylation and H3K4 mono-methylation were detected by western blotting. Colony formation and flow cytometry were used to examine proliferation and apoptosis in UC cells overexpressed or depleted with IRF6. Nude mice xenograft model was used to examine the effect of IRF6 in UC. RESULTS: Our result showed that several genes, including IRF6 were upregulated after treatment with CPH in BFTC905 UC cells. Further experiments found that treatment of CPH could restore the expression of IRF6 in several other UC cell lines, probably due to promoter hypomethylation and enrichment of H3K27 acetylation and H3K4 mono-methylation. These results may be due to the fact that CPH could alter the activity, but not the expression of epigenetic modifiers. Finally, re-expression of IRF6 in UC inhibited tumor growth in vitro and in an xenograft mouse model, by inducing apoptosis. CONCLUSION: In conclusion, our results suggested that CPH may be an epigenetic modifier, modulating the expression of the potential tumor suppressor IRF6, in inhibiting tumor growth in UC.
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Dendritic cells (DCs) play key roles in regulating T cell proliferation and differentiation, and epigenetic modification involves in this process. In the periphery, programmed death ligand-1 (PD-L1) expressed on antigen-presenting cells interacts with programmed death-1 (PD-1) on T cells to negatively regulate T cell responses. In this study, we investigate whether DNA demethylation in DCs, downmodulates CD4+ T cell activation, to halt progression of experimental autoimmune encephalomyelitis (EAE). These results showed that during the development of bone marrow-derived DCs (BMDCs), DNA hypomethylation by 0.1⯵M and 1⯵M 5-aza-2'-deoxycytidine (5-aza) upregulated PD-L1, but not CD40, CD80, or CD86, with surprising downregulation of PD-L2. In co-culture, 5-aza-treated BMDCs, as well as CD11c+ cells from 5-aza-treated EAE mice, inhibited EAE CD4+ T cell proliferation and cytokine secretion. Additionally, in vivo 5-aza pretreatment arrested disease progression, inflammatory cell infiltration, and CNS demyelination, in EAE mice. Compared to DCs from vehicle control-treated EAE rodents, DCs from 5-aza-treated EAE mice upregulated PD-L1, in correlation with hypomethylation of the Cd274 promoter. Furthermore, antibody-mediated blockage of PD-L1 rescued EAE progression from 5-aza treatment, in vivo, while also disinhibiting EAE CD4+ T cell proliferation, by 5-aza-treated DCs, in vitro. Consequently, we conclude that PD-L1 is upregulated via DNA hypomethylation in DCs, resulting in downregulation of autoimmune effector T cell functions, thereby halting progression of EAE.
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Antígeno B7-H1/genética , Desmetilação do DNA , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Encefalomielite Autoimune Experimental/etiologia , Linfócitos T/imunologia , Linfócitos T/metabolismo , Animais , Antígeno B7-H1/metabolismo , Biomarcadores , Citocinas/metabolismo , Modelos Animais de Doenças , Suscetibilidade a Doenças , Mediadores da Inflamação/metabolismo , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , CamundongosRESUMO
Gastric cancer (GC) is among the most treatment-refractory epithelial malignancies. Aberrant activation of Wnt/ß-catenin-signaling has been implicated in a variety of human cancers, including gastric cancer. Here we report that the elevated expression of lymphoid enhancer binding factor 1 (Lef1) is associated with the TNM (tumor- node-metastasis) stage of gastric cancer. Subsequently, 2,4-diamino-quinazoline (2,4-DAQ), a selective inhibitor of Lef1, was identified to suppress the expression of Wnt/ß-catenin target genes such as AXIN2, MYC and LGR5 and result in the suppression of gastric cancer cell growth through the apoptotic pathway. The 2,4-DAQ also exhibited an inhibitory effect on the migration/invasion of gastric cancer cells. Importantly, the treatment of human gastric tumor xenograft with 2,4-DAQ suppressed tumor growth in a nude mouse model. Furthermore, 2,4-DAQ appears effective on patient-derived organoids (PDOs). Transcriptome sequencing analysis also revealed that 2,4-DAQ are more effective on the gastric cancers that exhibit higher expression levels of Wnt-signaling pathway-related genes than their adjacent normal gastric tissues.
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Antineoplásicos/uso terapêutico , Fator 1 de Ligação ao Facilitador Linfoide/antagonistas & inibidores , Quinazolinas/uso terapêutico , Neoplasias Gástricas/tratamento farmacológico , Via de Sinalização Wnt/efeitos dos fármacos , Idoso , Animais , Antineoplásicos/farmacologia , Apoptose , Proteína Axina/genética , Proteína Axina/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Células Cultivadas , Feminino , Humanos , Fator 1 de Ligação ao Facilitador Linfoide/genética , Fator 1 de Ligação ao Facilitador Linfoide/metabolismo , Masculino , Camundongos , Camundongos Nus , Pessoa de Meia-Idade , Metástase Neoplásica , Proteínas Proto-Oncogênicas c-myc/genética , Proteínas Proto-Oncogênicas c-myc/metabolismo , Quinazolinas/farmacologia , Receptores Acoplados a Proteínas G/genética , Receptores Acoplados a Proteínas G/metabolismo , Neoplasias Gástricas/metabolismo , Neoplasias Gástricas/patologiaRESUMO
Serous ovarian cancer is the most frequent type of epithelial ovarian cancer. Despite the use of surgery and platinum-based chemotherapy, many patients suffer from recurrence within 6 months, termed platinum resistance. Currently, the lack of relevant molecular biomarkers for the prediction of the early recurrence of serous ovarian cancers is linked to the poor prognosis. To identify an effective biomarker for early recurrence, we analyzed the genome-wide DNA methylation status characteristic of early recurrence after treatment. The patients in The Cancer Genome Atlas (TCGA) dataset who showed a complete response after the first therapy were categorized into 2 groups: early recurrence serous ovarian cancer (ERS, recurrence ≤12 months, n = 51) and late recurrence serous ovarian cancer (LRS, recurrence >12 months, n = 158). Among the 12 differently methylated probes identified between the 2 groups, we found that ZNF671 was the most significantly methylated gene in the early recurrence group. A validation cohort of 78 serous ovarian cancers showed that patients with ZNF671 DNA methylation had a worse prognosis (P < .05). The multivariate analysis revealed that the methylation status of ZNF671 was an independent factor for predicting the recurrence of serous ovarian cancer patients both in the TCGA dataset and our cohort (P = .049 and P = .021, respectively). Functional analysis revealed that the depletion of ZNF671 expression conferred a more migratory and invasive phenotype to the ovarian cancer cells. Our data indicate that ZNF671 functions as a tumor suppressor in ovarian cancer and that the DNA methylation status of ZNF671 might be an effective biomarker for the recurrence of serous ovarian cancer after platinum-based adjuvant chemotherapy.
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Carcinoma Epitelial do Ovário/genética , Cistadenocarcinoma Seroso/genética , Metilação de DNA/genética , Recidiva Local de Neoplasia/genética , Proteínas Supressoras de Tumor/genética , Biomarcadores Tumorais/genética , Carcinoma Epitelial do Ovário/tratamento farmacológico , Carcinoma Epitelial do Ovário/patologia , Linhagem Celular Tumoral , Cistadenocarcinoma Seroso/patologia , Metilação de DNA/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/genética , Feminino , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/genética , Humanos , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/patologia , Platina/uso terapêutico , PrognósticoRESUMO
Ovarian cancer is the most lethal cancer of the female reproductive system. In that regard, several epidemiological studies suggest that long-term exposure to estrogen could increase ovarian cancer risk, although its precise role remains controversial. To decipher a mechanism for this, we previously generated a mathematical model of how estrogen-mediated upregulation of the transcription factor, E2F6, upregulates the ovarian cancer stem/initiating cell marker, c-Kit, by epigenetic silencing the tumor suppressor miR-193a, and a competing endogenous (ceRNA) mechanism. In this study, we tested that previous mathematical model, showing that estrogen treatment of immortalized ovarian surface epithelial cells upregulated both E2F6 and c-KIT, but downregulated miR-193a. Luciferase assays further confirmed that microRNA-193a targets both E2F6 and c-Kit. Interestingly, ChIP-PCR and bisulphite pyrosequencing showed that E2F6 also epigenetically suppresses miR-193a, through recruitment of EZH2, and by a complex ceRNA mechanism in ovarian cancer cell lines. Importantly, cell line and animal experiments both confirmed that E2F6 promotes ovarian cancer stemness, whereas E2F6 or EZH2 depletion derepressed miR-193a, which opposes cancer stemness, by alleviating DNA methylation and repressive chromatin. Finally, 118 ovarian cancer patients with miR-193a promoter hypermethylation had poorer survival than those without hypermethylation. These results suggest that an estrogen-mediated E2F6 ceRNA network epigenetically and competitively inhibits microRNA-193a activity, promoting ovarian cancer stemness and tumorigenesis.
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Fator de Transcrição E2F6/genética , Células-Tronco Neoplásicas/patologia , Neoplasias Ovarianas/genética , RNA/genética , Transcrição Gênica/genética , Animais , Linhagem Celular Tumoral , Metilação de DNA/efeitos dos fármacos , Metilação de DNA/genética , Epigênese Genética/efeitos dos fármacos , Epigênese Genética/genética , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/patologia , Estrogênios/efeitos adversos , Feminino , Genes Supressores de Tumor/fisiologia , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Camundongos SCID , MicroRNAs/genética , Células-Tronco Neoplásicas/efeitos dos fármacos , Neoplasias Ovarianas/etiologia , Transcrição Gênica/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genéticaRESUMO
DNA methylation contributes to tumor formation, development and metastasis. Epigenetic dysregulation of stem cells is thought to predispose to malignant development. The clinical significance of DNA methylation in ovarian tumor-initiating cells (OTICs) remains unexplored. We analyzed the methylomic profiles of OTICs (CP70sps) and their derived progeny using a human methylation array. qRT-PCR, quantitative methylation-specific PCR (qMSP) and pyrosequencing were used to verify gene expression and DNA methylation in cancer cell lines. The methylation status of genes was validated quantitatively in cancer tissues and correlated with clinicopathological factors. ATG4A and HIST1H2BN were hypomethylated in OTICs. Methylation analysis of ATG4A and HIST1H2BN by qMSP in 168 tissue samples from patients with ovarian cancer showed that HIST1H2BN methylation was a significant and independent predictor of progression-free survival (PFS) and overall survival (OS). Multivariate Cox regression analysis showed that patients with a low level of HIST1H2BN methylation had poor PFS (hazard ratio (HR), 4.5; 95% confidence interval (CI), 1.4-14.8) and OS (HR, 4.3; 95% CI, 1.3-14.0). Hypomethylation of both ATG4A and HIST1H2BN predicted a poor PFS (HR, 1.8; 95% CI, 1.0-3.6; median, 21 months) and OS (HR, 1.7; 95% CI, 1.0-3.0; median, 40 months). In an independent cohort of ovarian tumors, hypomethylation predicted early disease recurrence (HR, 1.7; 95% CI, 1.1-2.5) and death (HR, 1.4; 95% CI, 1.0-1.9). The demonstration that expression of ATG4A in cells increased their stem properties provided an indication of its biological function. Hypomethylation of ATG4A and HIST1H2BN in OTICs predicts a poor prognosis for ovarian cancer patients.
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Cisteína Endopeptidases/genética , Metilação de DNA/genética , Histonas/genética , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/mortalidade , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas Relacionadas à Autofagia , Sequência de Bases , Biomarcadores Tumorais/genética , Cisteína Endopeptidases/biossíntese , Intervalo Livre de Doença , Feminino , Humanos , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/genética , Recidiva Local de Neoplasia/mortalidade , Células-Tronco Neoplásicas , Prognóstico , Regiões Promotoras Genéticas , Interferência de RNA , RNA Interferente Pequeno , Análise de Sequência de DNA , Esferoides Celulares , Células Tumorais Cultivadas , Adulto JovemRESUMO
Alternative intravesical agents are required to overcome the side effects currently associated with the treatment of bladder cancer. This study used an orthotopic bladder cancer mouse model to evaluate Guizhi Fuling Wan (GFW) as an intravesical agent. The effects of GFW were compared with those of mitomycin-C (Mito-C) and bacille Calmette-Guérin (BCG). We began by evaluating the response of the mouse bladder cancer cell line MB49 to GFW treatment, with regard to cell viability, cell cycle progression and apoptosis. MB49 cells were subsequently implanted into the urothelial walls of the bladder in female C57BL/6 mice. The success of the model was confirmed by the appearance of hematuria and tumor growth in the bladder. Intravesical chemotherapy was administered in accordance with a published protocol. In vitro data revealed that GFW arrested MB49 cell cycle in the G0/G1 phase, resulting in the suppression of cell proliferation and induced apoptosis. One possible mechanism underlying these effects is an increase in intracellular reactive oxygen species (ROS) levels leading to the activation of ataxia telangiectasia-mutated (ATM)/checkpoint kinase 2 (CHK2) and ATM/P53 pathways, thereby mediating cell cycle progression and apoptosis, respectively. This mouse model demonstrates the effectiveness of GFW in the tumor growth, with results comparable to those achieved by using BCG and Mito-C. Furthermore, GFW was shown to cause only mild hematuria. The low toxicity of the compound was confirmed by a complete lack of lesions on bladder tissue, even after 10 consecutive treatments using high concentrations of GFW. These results demonstrate the potential of GFW for the intravesical therapy of bladder cancer.
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Aberrant Janus kinase (JAK)/signal transducer and activator of transcription (STAT) signaling is crucial to the development of gastric cancer. In this study, we examined the role of STAT3 in the expression and methylation of its targets in gastric cancer patients. Results from RNA sequencing identified an inverse correlation between the expression of STAT3 and GATA6 in 23 pairs of gastric cancer patient samples. We discovered that the expression of GATA6 is epigenetically silenced through promoter methylation in gastric cancer cell lines. Interestingly, the inhibition of STAT3 using a novel STAT3 inhibitor restored the expression of GATA6 and its targets, trefoil factors 1 and 2 (TFF1/2). Moreover, disruption of STAT3 binding to GATA6 promoter by small hairpin RNA restored GATA6 expression in AGS cells. A clinically significant correlation was also observed between the expression of GATA6 and TFF1/2 among tissue samples from 60 gastric cancer patients. Finally, bisulfite pyrosequencing revealed GATA6 methylation in 65% (39/60) of the patients, and those with higher GATA6 methylation tended to have shorter overall survival. In conclusion, we demonstrated that aberrant JAK/STAT signaling suppresses TFF1/2 partially through the epigenetic silencing of GATA6. Therapeutic intervention of STAT3 in reversing the epigenetic status of GATA6 could benefit the treatment of gastric cancer and is worthy of further investigation.
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Fator de Transcrição GATA6/metabolismo , Inativação Gênica , Transdução de Sinais , Neoplasias Gástricas/metabolismo , Fator Trefoil-1/metabolismo , Fator Trefoil-2/metabolismo , Idoso , Idoso de 80 Anos ou mais , Linhagem Celular Tumoral , Feminino , Fator de Transcrição GATA6/genética , Humanos , Janus Quinases/genética , Janus Quinases/metabolismo , Masculino , Pessoa de Meia-Idade , Fator de Transcrição STAT3/genética , Fator de Transcrição STAT3/metabolismo , Neoplasias Gástricas/genética , Fator Trefoil-1/genética , Fator Trefoil-2/genéticaRESUMO
Traumatic brain injury (TBI) is one of the most prevalent causes of worldwide mortality and morbidity. We previously had evidenced that TBI induced Na-K-2Cl co-transporter (NKCC1) upregulation in hippocampus. Here, we aim to investigate the role of NKCC1 in TBI-induced neurogenesis and the detailed mechanisms. The TBI-associated alternations in the expression of NKCC1, HIF-1α, VEGF, MAPK cascade, and CREB phosphorylation were analyzed by Western blot. TBI-induced neurogenesis was determined by immuno-fluorescence labeling. Chromatin immunoprecipitation was used to elucidate whether HIF-1α would activate VEGF gene after TBI. We found that the level of hippocampal NKCC1 and VEGF began to rise 8 h after TBI, and both of them reached maxima at day 7. Along with the upregulation of NKCC1 and VEGF, MAPK cascade was activated and hippocampal neurogenesis was promoted. Administration of CREB antisense oligonucleotide significantly attenuated the expression of HIF-1α, while HIF-1α antisense oligonucleotide exhibited little effect on the expression of CREB. However, HIF-1α antisense oligonucleotide administration did effectively suppress the expression of VEGF. Our results of the chromosome immunoprecipitation also indicated that HIF-1α could directly act on the VEGF promoter and presumably would elevate the VEGF expression after TBI. All these results have illustrated the correlation between NKCC1 upregulation and TBI-associated neurogenesis. The pathway involves the activation of Raf/MEK/ERK cascade, CREB phosphorylation, and HIF-1α upregulation, and finally leads to the stimulation of VEGF expression and the induction of neurogenesis.
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Lesões Encefálicas/metabolismo , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/metabolismo , Hipocampo/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Neurogênese , Neurônios/metabolismo , Membro 2 da Família 12 de Carreador de Soluto/metabolismo , Animais , Sítios de Ligação , Lesões Encefálicas/genética , Lesões Encefálicas/patologia , Lesões Encefálicas/fisiopatologia , Proteína de Ligação ao Elemento de Resposta ao AMP Cíclico/genética , Modelos Animais de Doenças , Regulação da Expressão Gênica , Hipocampo/patologia , Hipocampo/fisiopatologia , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Sistema de Sinalização das MAP Quinases , Masculino , Neurônios/patologia , Oligonucleotídeos Antissenso/genética , Oligonucleotídeos Antissenso/metabolismo , Fosforilação , Regiões Promotoras Genéticas , Ratos Wistar , Fatores de Tempo , Transcrição Gênica , Fator A de Crescimento do Endotélio Vascular/metabolismoRESUMO
Forkhead box P3 (Foxp3) is the major transcription factor controlling the development and function of regulatory T (Treg) cells. Previous studies have indicated epigenetic regulation of Foxp3 expression. Here, we investigated whether the deoxyribonucleic acid (DNA) methyltransferase inhibitor 5-aza-2'-deoxycytidine (5-Aza) applied peripherally could modulate central nervous system (CNS) inflammation, by using a mouse experimental autoimmune encephalomyelitis (EAE) model. We found that disease activity was inhibited in a myelin oligodendrocyte glycoprotein (MOG) peptide-induced EAE mouse briefly pretreated with low-dose (0.15 mg/kg) 5-Aza, ameliorating significant CNS inflammatory responses, as indicated by greatly decreased proinflammatory cytokines. On the contrary, control EAE mice expressed high levels of IFN-γ and interleukin (IL)-17. In addition, 5-Aza treatment in vitro increased GFP expression in CD4(+)GFP(-) T cells isolated from GFP knock-in Foxp3 transgenic mice. Importantly, 5-Aza treatment increased Treg cell numbers, in EAE mice, at both disease onset and peak. However, Treg inhibition assays showed 5-Aza treatment did not enhance per-cell Treg inhibitory function, but did maintain a lower activation threshold for effector cells in EAE mice. In conclusion, 5-Aza treatment prevented EAE development and suppressed CNS inflammation, by increasing the number of Treg cells and inhibiting effector cells in the periphery.
Assuntos
Azacitidina/análogos & derivados , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/prevenção & controle , Linfócitos T Reguladores/imunologia , Animais , Azacitidina/farmacologia , Azacitidina/uso terapêutico , Metilases de Modificação do DNA/antagonistas & inibidores , Decitabina , Encefalomielite Autoimune Experimental/patologia , Feminino , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/imunologia , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mycobacterium tuberculosis , Glicoproteína Mielina-Oligodendrócito , Medula Espinal/patologia , Baço/patologiaRESUMO
BACKGROUND: The treatment of oral squamous cell carcinoma (OSCC) following early detection is associated with good outcomes. Therefore, the survival and prognosis of OSCC patients could be hugely improved by identifying reliable biomarkers for the early diagnosis of the disease. Our previous methylation microarray analysis results have suggested that the gene encoding tissue factor pathway inhibitor-2 (TFPI-2) is a potential clinical predictor as well as a key regulator involved in OSCC malignancy. METHODS: Methylation of the TFPI-2 promoter in oral tissue specimens was evaluated by bisulfite sequencing assay, quantitative methylation-specific PCR, and pyrosequencing assay. The differences in methylation levels among the groups were compared using the Mann-Whitney U test. The area under the receiver operating characteristic curve (AUROC) was used to evaluate the discrimination ability for detecting OSCC. Cellular TFPI-2 expression was analyzed by quantitative reverse-transcription PCR before and after treatment with 5'-aza-2'-deoxycytidine and trichostatin A, to confirm whether TFPI-2 was epigenetically silenced in OSCC cells. We investigated whether TFPI-2 plays a role as a tumor suppressor by establishing TFPI-2-overexpressing OSCC cells and subjecting them to in vitro cellular proliferation, migration, and invasion assays, as well as an in vivo metastasis assay. RESULTS: TFPI-2 was hypermethylated in OSCC tissues versus normal oral tissues (P < 0.0001), with AUROC = 0.91, when using a pyrosequencing assay to quantify the methylation level. TFPI-2 silencing in OSCC was regulated by both DNA methylation and chromatin histone modification. Restoration of TFPI-2 counteracted the invasiveness of OSCC by inhibiting the enzymatic activity of matrix metalloproteinase-2, and consequently interfered with OSCC metastasis in vivo. CONCLUSIONS: Our data suggest strongly that TFPI-2 is a down-regulated tumor suppressor gene in OSCC, probably involving epigenetic silencing mechanisms. The loss of TFPI-2 expression is a key event for oral tumorigenesis, especially in the process of tumor metastasis.
Assuntos
Carcinoma de Células Escamosas/genética , Metilação de DNA , Inativação Gênica , Glicoproteínas/genética , Neoplasias Bucais/genética , Regiões Promotoras Genéticas , Animais , Biomarcadores Tumorais/genética , Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/diagnóstico , Carcinoma de Células Escamosas/patologia , Feminino , Regulação Neoplásica da Expressão Gênica , Glicoproteínas/metabolismo , Células HEK293 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Bucais/diagnóstico , Neoplasias Bucais/patologia , PrognósticoRESUMO
In this study, we aim to determine the relationship between methylation level of an inflammatory-related gene, SOCS-1 in serum samples of patients with ankylosing spondylitis (AS) and their degree of inflammation as well as serum cytokine level. Quantitative real time methylation specific PCR was performed to examine the promoter methylation of SOCS-1 in serum samples of 43 HLA-B27+ AS patients and 6 B27+ healthy controls. Degree of inflammation was accessed by spondylopathy, sacroiliitis as well as acute phase reactant, erythrocyte sedimentation rate and C-reactive protein (CRP). Serum IL-6 and TNF-α level was determined by ELISA assay. SOCS-1 methylation can only be found in serums samples from patients but not normal control. Methylation of SOCS-1 significantly associated with severity of patient's spondylopathy (P < 0.005), sacroiliitis (P < 0.005) and acute phase reactant CRP (P = 0.0278). AS patients also exhibited higher serum IL-6 (P < 0.001) and TNF-α level (P < 0.001). Importantly, patients with high serum IL-6 or TNF-α level demonstrated a significantly higher SOCS-1 methylation (P < 0.001). In conclusion, this proof-of-principle study suggested that methylation of SOCS-1 can be detected in serum of HLA-B27+ AS patients but not in B27+ controls. The pathogenic potential of SOCS-1 methylation in AS deserves further investigation.
Assuntos
Metilação de DNA/genética , Interleucina-6/sangue , Espondilite Anquilosante/genética , Proteínas Supressoras da Sinalização de Citocina/genética , Fator de Necrose Tumoral alfa/sangue , Adulto , Idoso , Idoso de 80 Anos ou mais , Epigênese Genética/genética , Feminino , Estudos de Associação Genética , Humanos , Inflamação/sangue , Inflamação/genética , Masculino , Pessoa de Meia-Idade , Regiões Promotoras Genéticas , Espondilite Anquilosante/sangue , Espondilite Anquilosante/patologia , Proteína 1 Supressora da Sinalização de CitocinaRESUMO
We previously reported that the sustained exposure of human urothelial cells (HUCs) to low-dose sodium arsenite induces changes in the gene expression profile and neoplastic transformation. In this study, we used the HumanMethylation27 BeadChip to analyze genome-wide methylation profiles and 5-aza-2'-deoxycytidine to examine the involvement of promoter methylation in gene expression. Because the expression of lipocalin-2 (LCN2) was highly enhanced by promoter hypomethylation in inorganic arsenic (iAs)-HUCs cells as well as bladder cancer tissues, we further showed that mutations at the binding sequences for NF-κB and C/EBP-α significantly reduced LCN2 promoter activity. By chromatin immunoprecipitation assay, we demonstrated the significantly increased binding of RelA (p65) and NF-κB1 (p50) to the hypomethylated promoter of LCN2 in the iAs-HUCs. Furthermore, we also demonstrated that LCN2 overexpression was crucial for the neoplastic characteristics of the iAs-HUCs, such as enhanced anchorage-independent growth, resistance to serum deprivation and activation of NF-κB signaling. In addition, our results indicated that enhanced NF-κB activity in iAs-HUCs was via LCN2-mediated increase in intracellular iron and reactive oxygen species levels. Taken together, our results show that sustained low-dose arsenic exposure results in epigenetic changes and enhanced oncogenic potential via LCN2 overexpression.
Assuntos
Proteínas de Fase Aguda/genética , Arsenitos/toxicidade , Metilação de DNA , Poluentes Ambientais/toxicidade , Lipocalinas/genética , Regiões Promotoras Genéticas , Proteínas Proto-Oncogênicas/genética , Compostos de Sódio/toxicidade , Urotélio/efeitos dos fármacos , Azacitidina/análogos & derivados , Azacitidina/farmacologia , Técnicas de Cultura de Células , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Imunoprecipitação da Cromatina , Meios de Cultura Livres de Soro , Metilação de DNA/efeitos dos fármacos , Metilação de DNA/genética , Decitabina , Relação Dose-Resposta a Droga , Inativação Gênica , Estudo de Associação Genômica Ampla , Humanos , Lipocalina-2 , Mutagênese Sítio-Dirigida , NF-kappa B/metabolismo , Ligação Proteica , Fatores de Tempo , Neoplasias da Bexiga Urinária/genética , Neoplasias da Bexiga Urinária/metabolismo , Neoplasias da Bexiga Urinária/patologia , Urotélio/citologia , Urotélio/metabolismoRESUMO
Ovarian cancer, as well as other cancers, is primarily caused by methylation at cytosines in CpG islands, but the current marker for ovarian cancer is low in sensitivity and failed in early-stage detection. Fourier transform infrared (FT-IR) spectroscopy is powerful in analysis of functional groups within molecules, and infrared microscopy illustrates the location of specific groups within single cells. In this study, we applied HPLC and FT-IR microspectrometry to study normal epithelial ovarian cell line immortalized ovarian surface epithelium (IOSE), two epithelial ovarian cell lines (A2780 and CP70) with distinct properties, and the effect of a cancer drug 5-aza-2'-deoxycytidine (5-aza) without labeling. Our results reveal that inhibition of methylation on cytosine with 5-aza initiates the protein expression. Furthermore, paraffin-adsorption kinetic study allows us to distinguish hypermethylated and hypomethyated cells, and this assay can be a potential diagnosis method for cancer screening.