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BACKGROUND: Exogenous formaldehyde is classified by the IARC as a Category 1 known human carcinogen. Meanwhile, a significant amount of endogenous formaldehyde is produced in the human body; as such, formaldehyde-derived DNA and protein adducts have been detected in animals and humans in the absence of major exogenous formaldehyde exposure. However, the toxicological effects of endogenous formaldehyde on individuals with normal DNA damage repair functions are not well understood. In this study, we attempted to generate C57BL/6 mice deficient in both Adh5 and Aldh2, which encode two major enzymes that metabolize endogenous formaldehyde, in order to understand the effects of endogenous formaldehyde on mice with normal DNA repair function. RESULTS: Due to deficiencies in both ADH5 and ALDH2, few mice survived past post-natal day 21. In fact, the survival of pups within the first few days after birth was significantly decreased. Remarkably, two Aldh2 -/- /Adh5 -/- mice survived for 25 days after birth, and we measured their total body weight and organ weights. The body weight of Aldh2 -/- /Adh5 -/- mice decreased significantly by almost 37% compared to the Aldh2 -/- /Adh5 +/- and Aldh2 -/- /Adh5 +/+ mice of the same litter. In addition, the absolute weight of each organ was also significantly reduced. CONCLUSION: Mice deficient in both formaldehyde-metabolizing enzymes ADH5 and ALDH2 were found to develop partial synthetic lethality and mortality shortly after birth. This phenotype may be due to the accumulation of endogenous formaldehyde. No serious phenotype has been reported in people with dysfunctional, dominant-negative ALDH2*2 alleles, but it has been reported that they may be highly susceptible to osteoporosis and neurodegenerative diseases. It is important to further investigate these diseases in individuals with ALDH2*2 alleles, including an association with decreased metabolism, and thus accumulation, of formaldehyde.
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Irinotecan treats a range of solid tumors, but its effectiveness is severely limited by gastrointestinal (GI) tract toxicity caused by gut bacterial ß-glucuronidase (GUS) enzymes. Targeted bacterial GUS inhibitors have been shown to partially alleviate irinotecan-induced GI tract damage and resultant diarrhea in mice. Here, we unravel the mechanistic basis for GI protection by gut microbial GUS inhibitors using in vivo models. We use in vitro, in fimo, and in vivo models to determine whether GUS inhibition alters the anticancer efficacy of irinotecan. We demonstrate that a single dose of irinotecan increases GI bacterial GUS activity in 1 d and reduces intestinal epithelial cell proliferation in 5 d, both blocked by a single dose of a GUS inhibitor. In a tumor xenograft model, GUS inhibition prevents intestinal toxicity and maintains the antitumor efficacy of irinotecan. Remarkably, GUS inhibitor also effectively blocks the striking irinotecan-induced bloom of Enterobacteriaceae in immune-deficient mice. In a genetically engineered mouse model of cancer, GUS inhibition alleviates gut damage, improves survival, and does not alter gut microbial composition; however, by allowing dose intensification, it dramatically improves irinotecan's effectiveness, reducing tumors to a fraction of that achieved by irinotecan alone, while simultaneously promoting epithelial regeneration. These results indicate that targeted gut microbial enzyme inhibitors can improve cancer chemotherapeutic outcomes by protecting the gut epithelium from microbial dysbiosis and proliferative crypt damage.
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Microbioma Gastrointestinal/efeitos dos fármacos , Glucuronidase/antagonistas & inibidores , Glucuronidase/efeitos dos fármacos , Animais , Antineoplásicos Fitogênicos/farmacologia , Bactérias/efeitos dos fármacos , Modelos Animais de Doenças , Disbiose/tratamento farmacológico , Inibidores Enzimáticos/farmacologia , Feminino , Glucuronidase/metabolismo , Humanos , Irinotecano/farmacologia , Camundongos , Camundongos Nus , Neoplasias/tratamento farmacológicoRESUMO
In a recent Cell Reports article, Li et al. report that obesity is associated with altered fatty acid metabolism and DNA methylation in the colonic epithelium, which precede a tumor-prone gene-expression profile. Interestingly, obesity-associated methylation and transcriptome changes were reversed by weight loss, and the duration of weight loss correlated with the extent of restored gene expression. These findings have implications that are encouraging for weight loss and cancer prevention.
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Neoplasias Colorretais , Epigênese Genética , Animais , DNA , Dieta , Camundongos , Obesidade/genética , TranscriptomaRESUMO
BACKGROUND & AIMS: The successful culture of intestinal organoids has greatly enhanced our understanding of intestinal stem cell physiology and enabled the generation of novel intestinal disease models. Although of tremendous value, intestinal organoid culture systems have not yet fully recapitulated the anatomy or physiology of the in vivo intestinal epithelium. The aim of this work was to re-create an intestinal epithelium with a high density of polarized crypts that respond in a physiologic manner to addition of growth factors, metabolites, or cytokines to the basal or luminal tissue surface as occurs in vivo. METHODS: A self-renewing monolayer of human intestinal epithelium was cultured on a collagen scaffold microfabricated with an array of crypt-like invaginations. Placement of chemical factors in either the fluid reservoir below or above the cell-covered scaffolding created a gradient of that chemical across the growing epithelial tissue possessing the in vitro crypt structures. Crypt polarization (size of the stem/proliferative and differentiated cell zones) was assessed in response to gradients of growth factors, cytokines, and bacterial metabolites. RESULTS: Chemical gradients applied to the shaped human epithelium re-created the stem/proliferative and differentiated cell zones of the in vivo intestine. Short-chain fatty acids applied as a gradient from the luminal side confirmed long-standing hypotheses that butyrate diminished stem/progenitor cell proliferation and promoted differentiation into absorptive colonocytes. A gradient of interferon-γ and tumor necrosis factor-α significantly suppressed the stem/progenitor cell proliferation, altering crypt formation. CONCLUSIONS: The in vitro human colon crypt array accurately mimicked the architecture, luminal accessibility, tissue polarity, cell migration, and cellular responses of in vivo intestinal crypts.
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The intestinal epithelium provides a critical barrier that separates the gut microbiota from host tissues. Nonsteroidal anti-inflammatory drugs (NSAIDs) are efficacious analgesics and antipyretics and are among the most frequently used drugs worldwide. In addition to gastric damage, NSAIDs are toxic to the intestinal epithelium, causing erosions, perforations, and longitudinal ulcers in the gut. Here, we use a unique in vitro human primary small intestinal cell monolayer system to pinpoint the intestinal consequences of NSAID treatment. We found that physiologically relevant doses of the NSAID diclofenac (DCF) are cytotoxic because they uncouple mitochondrial oxidative phosphorylation and generate reactive oxygen species. We also find that DCF induces intestinal barrier permeability, facilitating the translocation of compounds from the luminal to the basolateral side of the intestinal epithelium. The results we outline here establish the utility of this novel platform, representative of the human small intestinal epithelium, to understand NSAID toxicity, which can be applied to study multiple aspects of gut barrier function including defense against infectious pathogens and host-microbiota interactions.
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Anti-Inflamatórios não Esteroides/efeitos adversos , Permeabilidade da Membrana Celular/efeitos dos fármacos , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Mucosa Intestinal/efeitos dos fármacos , Mucosa Intestinal/metabolismo , Humanos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismoRESUMO
BACKGROUND & AIMS: Three-dimensional organoid culture has fundamentally changed the in vitro study of intestinal biology enabling novel assays; however, its use is limited because of an inaccessible luminal compartment and challenges to data gathering in a three-dimensional hydrogel matrix. Long-lived, self-renewing 2-dimensional (2-D) tissue cultured from primary colon cells has not been accomplished. METHODS: The surface matrix and chemical factors that sustain 2-D mouse colonic and human rectal epithelial cell monolayers with cell repertoires comparable to that in vivo were identified. RESULTS: The monolayers formed organoids or colonoids when placed in standard Matrigel culture. As with the colonoids, the monolayers exhibited compartmentalization of proliferative and differentiated cells, with proliferative cells located near the peripheral edges of growing monolayers and differentiated cells predominated in the central regions. Screening of 77 dietary compounds and metabolites revealed altered proliferation or differentiation of the murine colonic epithelium. When exposed to a subset of the compound library, murine organoids exhibited similar responses to that of the monolayer but with differences that were likely attributable to the inaccessible organoid lumen. The response of the human primary epithelium to a compound subset was distinct from that of both the murine primary epithelium and human tumor cells. CONCLUSIONS: This study demonstrates that a self-renewing 2-D murine and human monolayer derived from primary cells can serve as a physiologically relevant assay system for study of stem cell renewal and differentiation and for compound screening. The platform holds transformative potential for personalized and precision medicine and can be applied to emerging areas of disease modeling and microbiome studies.
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Endogenous formaldehyde is abundantly present in our bodies, at around 100 µM under normal conditions. While such high steady state levels of formaldehyde may be derived by enzymatic reactions including oxidative demethylation/deamination and myeloperoxidation, it is unclear whether endogenous formaldehyde can initiate and/or promote diseases in humans. Here, we show that fluorescent malondialdehyde-formaldehyde (M2FA)-lysine adducts are immunogenic without adjuvants in mice. Natural antibody titers against M2FA are elevated in atherosclerosis-prone mice. Staining with an antibody against M2FA demonstrated that M2FA is present in plaque found on the aortic valve of ApoE -/- mice. To mimic inflammation during atherogenesis, human myeloperoxidase was incubated with glycine, H2O2, malondialdehyde, and a lysine analog in PBS at a physiological temperature, which resulted in M2FA generation. These results strongly suggest that the 1,4-dihydropyridine-type of lysine adducts observed in atherosclerosis lesions are likely produced by endogenous formaldehyde and malondialdehyde with lysine. These highly fluorescent M2FA adducts may play important roles in human inflammatory and degenerative diseases.
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Aterosclerose/imunologia , Aterosclerose/metabolismo , Epitopos/imunologia , Formaldeído/metabolismo , Animais , Apolipoproteínas E/deficiência , Cromatografia Líquida , Formaldeído/química , Humanos , Espectroscopia de Ressonância Magnética , Espectrometria de Massas , Camundongos , Camundongos Knockout , Estrutura Molecular , Peroxidase/metabolismo , Placa Aterosclerótica/imunologia , Placa Aterosclerótica/metabolismoRESUMO
Answer questions and earn CME/CNE The human body harbors enormous numbers of microbiota that influence cancer susceptibility, in part through their prodigious metabolic capacity and their profound influence on immune cell function. Microbial pathogens drive tumorigenesis in 15% to 20% of cancer cases. Even larger numbers of malignancies are associated with an altered composition of commensal microbiota (dysbiosis) based on microbiome studies using metagenomic sequencing. Although association studies cannot distinguish whether changes in microbiota are causes or effects of cancer, a causative role is supported by rigorously controlled preclinical studies using gnotobiotic mouse models colonized with one or more specific bacteria. These studies demonstrate that microbiota can alter cancer susceptibility and progression by diverse mechanisms, such as modulating inflammation, inducing DNA damage, and producing metabolites involved in oncogenesis or tumor suppression. Evidence is emerging that microbiota can be manipulated for improving cancer treatment. By incorporating probiotics as adjuvants for checkpoint immunotherapy or by designing small molecules that target microbial enzymes, microbiota can be harnessed to improve cancer care. CA Cancer J Clin 2017;67:326-344. © 2017 American Cancer Society.
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Microbiota , Neoplasias/microbiologia , Neoplasias/terapia , Animais , Carcinogênese , Modelos Animais de Doenças , Progressão da Doença , Suscetibilidade a Doenças , Disbiose , Humanos , Metagenômica , Medicina de PrecisãoRESUMO
The association between obesity and breast cancer risk and prognosis is well established in estrogen receptor (ER)-positive disease but less clear in HER2-positive disease. Here, we report preclinical evidence suggesting weight maintenance through calorie restriction (CR) may limit risk of HER2-positive breast cancer. In female MMTV-HER2/neu transgenic mice, we found that ERα and ERß expression, mammary tumorigenesis, and survival are energy balance dependent in association with epigenetic reprogramming. Mice were randomized to receive a CR, overweight-inducing, or diet-induced obesity regimen (n = 27/group). Subsets of mice (n = 4/group/time point) were euthanized after 1, 3, and 5 months to characterize diet-dependent metabolic, transcriptional, and epigenetic perturbations. Remaining mice were followed up to 22 months. Relative to the overweight and diet-induced obesity regimens, CR decreased body weight, adiposity, and serum metabolic hormones as expected and also elicited an increase in mammary ERα and ERß expression. Increased DNA methylation accompanied this pattern, particularly at CpG dinucleotides located within binding or flanking regions for the transcriptional regulator CCCTC-binding factor of ESR1 and ESR2, consistent with sustained transcriptional activation of ERα and ERß. Mammary expression of the DNA methylation enzyme DNMT1 was stable in CR mice but increased over time in overweight and diet-induced obesity mice, suggesting CR obviates epigenetic alterations concurrent with chronic excess energy intake. In the survival study, CR elicited a significant suppression in spontaneous mammary tumorigenesis. Overall, our findings suggest a mechanistic rationale to prevent or reverse excess body weight as a strategy to reduce HER2-positive breast cancer risk. Cancer Res; 77(9); 2500-11. ©2017 AACR.
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Neoplasias da Mama/genética , Receptor alfa de Estrogênio/genética , Receptor beta de Estrogênio/genética , Neoplasias Mamárias Animais/genética , Obesidade/genética , Animais , Neoplasias da Mama/fisiopatologia , Restrição Calórica , Carcinogênese/genética , Metilação de DNA/genética , Metabolismo Energético/genética , Epigênese Genética/genética , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Neoplasias Mamárias Animais/etiologia , Neoplasias Mamárias Animais/fisiopatologia , Camundongos , Camundongos Transgênicos , Obesidade/complicações , Obesidade/fisiopatologia , Receptor ErbB-2/genética , Fatores de RiscoRESUMO
RATIONALE: The contractile dysfunction that underlies heart failure involves perturbations in multiple biological processes ranging from metabolism to electrophysiology. Yet the epigenetic mechanisms that are altered in this disease state have not been elucidated. SWI/SNF chromatin-remodeling complexes are plausible candidates based on mouse knockout studies demonstrating a combined requirement for the BRG1 and BRM catalytic subunits in adult cardiomyocytes. Brg1/Brm double mutants exhibit metabolic and mitochondrial defects and are not viable although their cause of death has not been ascertained. OBJECTIVE: To determine the cause of death of Brg1/Brm double-mutant mice, to test the hypothesis that BRG1 and BRM are required for cardiac contractility, and to identify relevant downstream target genes. METHODS AND RESULTS: A tamoxifen-inducible gene-targeting strategy utilizing αMHC-Cre-ERT was implemented to delete both SWI/SNF catalytic subunits in adult cardiomyocytes. Brg1/Brm double-mutant mice were monitored by echocardiography and electrocardiography, and they underwent rapidly progressive ventricular dysfunction including conduction defects and arrhythmias that culminated in heart failure and death within 3weeks. Mechanistically, BRG1/BRM repressed c-Myc expression, and enforced expression of a DOX-inducible c-MYC trangene in mouse cardiomyocytes phenocopied the ventricular conduction defects observed in Brg1/Brm double mutants. BRG1/BRM and c-MYC had opposite effects on the expression of cardiac conduction genes, and the directionality was consistent with their respective loss- and gain-of-function phenotypes. To support the clinical relevance of this mechanism, BRG1/BRM occupancy was diminished at the same target genes in human heart failure cases compared to controls, and this correlated with increased c-MYC expression and decreased CX43 and SCN5A expression. CONCLUSION: BRG1/BRM and c-MYC have an antagonistic relationship regulating the expression of cardiac conduction genes that maintain contractility, which is reminiscent of their antagonistic roles as a tumor suppressor and oncogene in cancer.
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DNA Helicases/metabolismo , Sistema de Condução Cardíaco , Contração Miocárdica , Miócitos Cardíacos/metabolismo , Proteínas Nucleares/metabolismo , Proteínas Proto-Oncogênicas c-myc/metabolismo , Fatores de Transcrição/metabolismo , Animais , DNA Helicases/genética , Eletrocardiografia , Perfilação da Expressão Gênica , Regulação da Expressão Gênica , Insuficiência Cardíaca/diagnóstico , Insuficiência Cardíaca/genética , Insuficiência Cardíaca/metabolismo , Insuficiência Cardíaca/fisiopatologia , Humanos , Camundongos , Camundongos Transgênicos , Mutação , Contração Miocárdica/genética , Proteínas Nucleares/genética , Ligação Proteica , Proteínas Proto-Oncogênicas c-myc/genética , Fatores de Transcrição/genéticaRESUMO
Despite the success of colonoscopy screening, colorectal cancer (CRC) remains one of the most common and deadly cancers, and CRC incidence is rising in some countries where screening is not routine and populations have recently switched from traditional diets to western diets. Diet and energy balance influence CRC by multiple mechanisms. They modulate the composition and function of gut microbiota, which have a prodigious metabolic capacity and can produce oncometabolites or tumor-suppressive metabolites depending, in part, on which dietary factors and digestive components are present in the GI tract. Gut microbiota also have a profound effect on immune cells in the lamina propria, which influences inflammation and subsequently CRC. Nutrient availability, which is an outcome of diet and energy balance, determines the abundance of certain energy metabolites that are essential co-factors for epigenetic enzymes and therefore impinges upon epigenetic regulation of gene expression. Aberrant epigenetic marks accumulate during CRC, and epimutations that are selected for drive tumorigenesis by causing transcriptome profiles to diverge from the cell of origin. In some instances, the above mechanisms are intertwined as exemplified by dietary fiber being metabolized by colonic bacteria into butyrate, which is both a short-chain fatty acid (SCFA) and a histone deacetylase (HDAC) inhibitor that epigenetically upregulates tumor-suppressor genes in CRC cells and anti-inflammatory genes in immune cells.
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Neoplasias Colorretais/genética , Neoplasias Colorretais/microbiologia , Epigênese Genética , Microbioma Gastrointestinal , Animais , Butiratos/metabolismo , Neoplasias Colorretais/prevenção & controle , Dieta , Fibras na Dieta/farmacologia , Microbioma Gastrointestinal/genética , Humanos , Prebióticos , Probióticos/farmacologiaRESUMO
It is becoming increasingly clear that microbiota inhabiting our bodies influence cancer predisposition and etiology. In addition to pathogens with oncogenic properties, commensal and symbiotic microbiota have tumor-suppressive properties. Diet and other environmental factors can modulate the abundance of certain members of microbial communities within the gastrointestinal tract and at other anatomical sites. Furthermore, some dietary factors are metabolized by commensal/symbiotic gut microbiota into bioactive food components believed to prevent cancer. For example, dietary fiber undergoes bacterial fermentation in the colon to yield butyrate, which is a short-chain fatty acid and histone deacetylase (HDAC) inhibitor that suppresses the viability and growth of colorectal cancer cell lines. A recent study using gnotobiotic mouse models demonstrates that fiber can protect against colorectal tumorigenesis in a microbiota- and butyrate-dependent manner that involves the Warburg effect. This and other examples suggest that some of the inter-individual variation observed in epidemiology and intervention studies that have investigated associations between diet and cancer risk might be explained by differences in microbiota among the participants. Data from basic research studies also support the idea that probiotics and prebiotics could be plausible chemoprevention strategies that may be utilized to a greater extent in the future.
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Butiratos/metabolismo , Neoplasias Colorretais/prevenção & controle , Fibras na Dieta/metabolismo , Microbioma Gastrointestinal/fisiologia , Animais , Fibras na Dieta/administração & dosagem , Humanos , Interações Microbianas , Prebióticos , ProbióticosRESUMO
Mammalian SWI/SNF chromatin-remodeling complexes utilize either BRG1 or Brm as alternative catalytic subunits to alter the position of nucleosomes and regulate gene expression. Genetic studies have demonstrated that SWI/SNF complexes are required during cardiac development and also protect against cardiovascular disease and cancer. However, Brm constitutive null mutants do not exhibit a cardiomyocyte phenotype and inducible Brg1 conditional mutations in cardiomyocyte do not demonstrate differences until stressed with transverse aortic constriction, where they exhibit a reduction in cardiac hypertrophy. We recently demonstrated the overlapping functions of Brm and Brg1 in vascular endothelial cells and sought here to test if this overlapping function occurred in cardiomyocytes. Brg1/Brm double mutants died within 21 days of severe cardiac dysfunction associated with glycogen accumulation and mitochondrial defects based on histological and ultrastructural analyses. To determine the underlying defects, we performed nontargeted metabolomics analysis of cardiac tissue by GC/MS from a line of Brg1/Brm double-mutant mice, which lack both Brg1 and Brm in cardiomyocytes in an inducible manner, and two groups of controls. Metabolites contributing most significantly to the differences between Brg1/Brm double-mutant and control-group hearts were then determined using the variable importance in projection analysis. Increased cardiac linoleic acid and oleic acid suggest alterations in fatty acid utilization or intake are perturbed in Brg1/Brm double mutants. Conversely, decreased glucose-6-phosphate, fructose-6-phosphate, and myoinositol suggest that glycolysis and glycogen formation are impaired. These novel metabolomics findings provide insight into SWI/SNF-regulated metabolic pathways and will guide mechanistic studies evaluating the role of SWI/SNF complexes in homeostasis and cardiovascular disease prevention.
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Although the two catalytic subunits of the SWI/SNF chromatin-remodeling complex--Brahma (Brm) and Brg1--are almost invariably co-expressed, their mutually exclusive incorporation into distinct SWI/SNF complexes predicts that Brg1- and Brm-based SWI/SNF complexes execute specific functions. Here, we show that Brg1 and Brm have distinct functions at discrete stages of muscle differentiation. While Brg1 is required for the activation of muscle gene transcription at early stages of differentiation, Brm is required for Ccnd1 repression and cell cycle arrest prior to the activation of muscle genes. Ccnd1 knockdown rescues the ability to exit the cell cycle in Brm-deficient myoblasts, but does not recover terminal differentiation, revealing a previously unrecognized role of Brm in the activation of late muscle gene expression independent from the control of cell cycle. Consistently, Brm null mice displayed impaired muscle regeneration after injury, with aberrant proliferation of satellite cells and delayed formation of new myofibers. These data reveal stage-specific roles of Brm during skeletal myogenesis, via formation of repressive and activatory SWI/SNF complexes.
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Pontos de Checagem do Ciclo Celular/genética , DNA Helicases/metabolismo , Expressão Gênica , Desenvolvimento Muscular/genética , Proteínas Nucleares/metabolismo , Fatores de Transcrição/metabolismo , Animais , Ciclina D1/deficiência , Ciclina D1/genética , DNA Helicases/genética , Técnicas de Silenciamento de Genes , Camundongos , Células Musculares , Proteínas Nucleares/genética , Fatores de Transcrição/genéticaRESUMO
SWI/SNF complexes utilize BRG1 (also known as SMARCA4) or BRM (also known as SMARCA2) as alternative catalytic subunits with ATPase activity to remodel chromatin. These chromatin-remodeling complexes are required for mammalian development and are mutated in ~20% of all human primary tumors. Yet our knowledge of their tumor-suppressor mechanism is limited. To investigate the role of SWI/SNF complexes in the DNA-damage response (DDR), we used shRNAs to deplete BRG1 and BRM and then exposed these cells to a panel of 6 genotoxic agents. Compared to controls, the shRNA knockdown cells were hypersensitive to certain genotoxic agents that cause double-strand breaks (DSBs) associated with stalled/collapsed replication forks but not to ionizing radiation-induced DSBs that arise independently of DNA replication. These findings were supported by our analysis of DDR kinases, which demonstrated a more prominent role for SWI/SNF in the activation of the ATR-Chk1 pathway than the ATM-Chk2 pathway. Surprisingly, γH2AX induction was attenuated in shRNA knockdown cells exposed to a topoisomerase II inhibitor (etoposide) but not to other genotoxic agents including IR. However, this finding is compatible with recent studies linking SWI/SNF with TOP2A and TOP2BP1. Depletion of BRG1 and BRM did not result in genomic instability in a tumor-derived cell line but did result in nucleoplasmic bridges in normal human fibroblasts. Taken together, these results suggest that SWI/SNF tumor-suppressor activity involves a role in the DDR to attenuate replicative stress and genomic instability. These results may also help to inform the selection of chemotherapeutics for tumors deficient for SWI/SNF function.
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Dano ao DNA , DNA Helicases/genética , Proteínas Nucleares/genética , Fatores de Transcrição/genética , Neoplasias do Colo do Útero/genética , Linhagem Celular Tumoral , Sobrevivência Celular/genética , DNA Helicases/deficiência , Feminino , Técnicas de Silenciamento de Genes , Células HeLa , Humanos , Proteínas Nucleares/deficiência , RNA Interferente Pequeno/genética , Fatores de Transcrição/deficiênciaRESUMO
UNLABELLED: Whether dietary fiber protects against colorectal cancer is controversial because of conflicting results from human epidemiologic studies. However, these studies and mouse models of colorectal cancer have not controlled the composition of gut microbiota, which ferment fiber into short-chain fatty acids such as butyrate. Butyrate is noteworthy because it has energetic and epigenetic functions in colonocytes and tumor-suppressive properties in colorectal cancer cell lines. We used gnotobiotic mouse models colonized with wild-type or mutant strains of a butyrate-producing bacterium to demonstrate that fiber does have a potent tumor-suppressive effect but in a microbiota- and butyrate-dependent manner. Furthermore, due to the Warburg effect, butyrate was metabolized less in tumors where it accumulated and functioned as a histone deacetylase (HDAC) inhibitor to stimulate histone acetylation and affect apoptosis and cell proliferation. To support the relevance of this mechanism in human cancer, we demonstrate that butyrate and histone-acetylation levels are elevated in colorectal adenocarcinomas compared with normal colonic tissues. SIGNIFICANCE: These results, which link diet and microbiota to a tumor-suppressive metabolite, provide insight into conflicting epidemiologic findings and suggest that probiotic/prebiotic strategies can modulate an endogenous HDAC inhibitor for anticancer chemoprevention without the adverse effects associated with synthetic HDAC inhibitors used in chemotherapy.
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Butiratos/metabolismo , Transformação Celular Neoplásica , Neoplasias Colorretais/etiologia , Fibras na Dieta , Vida Livre de Germes , Microbiota , Animais , Carcinógenos/administração & dosagem , Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Humanos , Mucosa Intestinal/patologia , Camundongos , Gradação de TumoresRESUMO
Dietary factors, microbial composition, and metabolism are intimately intertwined into a complex network whose activities influence important intestinal functions. In a recent issue of Cell, Belcheva et al. (2014) show that microbial-derived butyrate promotes proliferation of cancer-initiated intestinal epithelial cells, suggesting that it can act as an oncometabolite.
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Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/patologia , Carboidratos da Dieta/metabolismo , Proteína 2 Homóloga a MutS/metabolismo , AnimaisRESUMO
Gene-environment interactions underlie cancer susceptibility and progression. Yet, we still have limited knowledge of which environmental factors are important and how they function during tumorigenesis. In this respect, the microbial communities that inhabit our gastrointestinal tract and other body sites have been unappreciated until recently. However, our microbiota are environmental factors that we are exposed to continuously, and human microbiome studies have revealed significant differences in the relative abundance of certain microbes in cancer cases compared with controls. To characterize the function of microbiota in carcinogenesis, mouse models of cancer have been treated with antibiotics. They have also been maintained in a germfree state or have been colonized with specific bacteria in specialized (gnotobiotic) facilities. These studies demonstrate that microbiota can increase or decrease cancer susceptibility and progression by diverse mechanisms such as by modulating inflammation, influencing the genomic stability of host cells and producing metabolites that function as histone deacetylase inhibitors to epigenetically regulate host gene expression. One might consider microbiota as tractable environmental factors because they are highly quantifiable and relatively stable within an individual compared with our exposures to external agents. At the same time, however, diet can modulate the composition of microbial communities within our gut, and this supports the idea that probiotics and prebiotics can be effective chemoprevention strategies. The trajectory of where the current work is headed suggests that microbiota will continue to provide insight into the basic mechanisms of carcinogenesis and that microbiota will also become targets for therapeutic intervention.
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Antibacterianos/uso terapêutico , Trato Gastrointestinal/microbiologia , Microbiota/efeitos dos fármacos , Neoplasias/microbiologia , Animais , Trato Gastrointestinal/efeitos dos fármacos , Humanos , Camundongos , Neoplasias/tratamento farmacológicoRESUMO
Mammalian SWI/SNF-related complexes are recruited to the promoters of numerous target genes, and the BRG1 catalytic subunit confers ATPase activity necessary to slide or evict nucleosomes and to regulate transcription. Based on gene-targeting experiments in mice, BRG1 is essential for early embryonic development. However, Brg1 null mutants have provided limited insight into gene-dosage considerations and structure-function relationships. To extend our knowledge of BRG1 function, we describe the genetic and biochemical characteristics of an ENU-induced hypomorphic mutation that encodes a protein with a single amino-acid substitution (E1083G) within the bilobal ATPase/chromatin-remodeling domain. Brg1(ENU1/ENU1) mice have ~50% genetic activity and survive embryogenesis but exhibit a postnatal developmental phenotype associated with runting and incompletely penetrant lethality. The E1083G mutant protein is stable, and experiments with recombinant FLAG-tagged BRG1 proteins demonstrated that it retains full ATPase activity. Yet the biochemical activity of the mutant protein is diminished to ~50% of normal in chromatin-remodeling assays. Consistent with these findings, the E1083G substitution is predicted to disrupt a structurally conserved α-helix within the lobe that participates in DNA translocation but does not contain the ATPase catalytic site. We propose that this α-helix participates in the DNA translocation cycle by mechanistically linking DNA interaction surfaces at the DNA entry/anchor point to those within the Helicase C domain of lobe 2 of the bilobal ATPase motor. Taken together, these results demonstrate that BRG1 genetic and biochemical activities are tightly correlated. They also indicate that BRG1 ATPase activity is necessary but not sufficient for chromatin remodeling.