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The respiratory effects of particulate matter (PM) in subway station platforms or tunnels have attracted considerable research attention. However, no studies have characterized the effects of subway PM on allergic immune responses. In this study, iron oxide (α-Fe2O3 and Fe3O4) particles-the main components of subway PM-were intratracheally administered to BALB/c mice where ovalbumin (OVA) induced allergic pulmonary inflammation. Iron oxide particles enhanced OVA-induced eosinophil recruitment around the bronchi and mucus production from airway epithelium. The concentrations of type 2 cytokines, namely, interleukin (IL)-5 and IL-13, in bronchial alveolar lavage fluids were increased by iron oxide particles. Iron oxide particles also increased the number of type 2 innate lymphoid cells and CD86+ cells in the lung. Moreover, phagocytosis of particles in lung cells was confirmed by Raman spectroscopy. In a subsequent in vitro study, bone marrow-derived antigen-presenting cells (APCs) isolated from NC/Nga mice were exposed to iron oxide particles and OVA. They were also exposed to outdoor ambient PM: Vehicle Exhaust Particulates (VEP) and Urban Aerosols (UA) as references. Iron oxide particles promoted the release of lactate dehydrogenase, C-X-C motif chemokine ligand 1 and IL-1α from APCs, which tended to be stronger than those of VEP. These results suggest that iron oxide particles enhance antigen presentation in the lungs, promoting allergic immune response in mice; iron oxide particles-induced death and inflammatory response of APCs can contribute to allergy exacerbation. Although iron oxide particles do not contain various compounds like VEP, iron oxide alone may have sufficient influence.
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Contaminantes Atmosféricos , Compuestos Férricos , Hipersensibilidad , Ratones Endogámicos BALB C , Material Particulado , Animales , Material Particulado/toxicidad , Ratones , Contaminantes Atmosféricos/toxicidad , Hipersensibilidad/inmunología , Pulmón/efectos de los fármacos , Pulmón/inmunología , Citocinas/metabolismo , Ovalbúmina , Líquido del Lavado Bronquioalveolar/química , Emisiones de Vehículos/toxicidad , FemeninoRESUMEN
Viruses infect and kill prokaryotic populations in a density- or frequency-dependent manner and affect carbon cycling. However, the effects of the stratification transition, including the stratified and de-stratified periods, on the changes in prokaryotic and viral communities and their interactions remain unclear. We conducted a monthly survey of the surface and deep layers of a large and deep freshwater lake (Lake Biwa, Japan) for a year and analyzed the prokaryotic production and prokaryotic and viral community composition. Our analysis revealed that, in the surface layer, 19 prokaryotic species, accounting for approximately 40% of the total prokaryotic abundance, could potentially contribute to the majority of prokaryotic production, which is the highest during the summer and is suppressed by viruses. This suggests that a small fraction of prokaryotes and phages were the key infection pairs during the peak period of prokaryotic activity in the freshwater lake. We also found that approximately 50% of the dominant prokaryotic and viral species in the deep layer were present throughout the study period. This suggests that the "kill the winner" model could explain the viral impact on prokaryotes in the surface layer, but other dynamics may be at play in the deep layer. Furthermore, we found that annual vertical mixing could result in a similar rate of community change between the surface and deep layers. These findings may be valuable in understanding how communities and the interaction among them change when freshwater lake stratification is affected by global warming in the future.IMPORTANCEViral infection associated with prokaryotic production occurs in a density- or frequency-dependent manner and regulates the prokaryotic community. Stratification transition and annual vertical mixing in freshwater lakes are known to affect the prokaryotic community and the interaction between prokaryotes and viruses. By pairing measurements of virome analysis and prokaryotic production of a 1-year survey of the depths of surface and deep layers, we revealed (i) the prokaryotic infection pairs associated with prokaryotic production and (ii) the reset in prokaryotic and viral communities through annual vertical mixing in a freshwater lake. Our results provide a basis for future work into changes in stratification that may impact the biogeochemical cycling in freshwater lakes.
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Bacteriófagos , Virus , Lagos/química , Células Procariotas , Virus/genética , Bacteriófagos/genética , JapónRESUMEN
BACKGROUND: DNA damage response (DDR) and repair are vital for safeguarding genetic information and ensuring the survival and accurate transmission of genetic material. DNA damage, such as DNA double-strand breaks (DSBs), triggers a response where sensor proteins recognize DSBs. Information is transmitted to kinases, initiating a sequence resulting in the activation of the DNA damage response and recruitment of other DDR and repair proteins to the DSB site in a highly orderly sequence. Research has traditionally focused on individual protein functions and their order, with limited quantitative analysis, prompting this study's attempt at absolute quantification of DNA damage response and repair proteins and capturing changes in protein chromatin affinity after DNA damage through biochemical fractionation methods. RESULTS: To assess the intracellular levels of proteins involved in DDR and repair, multiple proteins associated with different functions were quantified in EPC2-hTERT cells. H2AX had the highest intracellular abundance (1.93 × 106 molecules/cell). The components of the MRN complex were present at the comparable levels: 6.89 × 104 (MRE11), 2.17 × 104 (RAD50), and 2.35 × 104 (NBS1) molecules/cell. MDC1 was present at 1.27 × 104 molecules/cell. The intracellular levels of ATM and ATR kinases were relatively low: 555 and 4860 molecules/cell, respectively. The levels of cellular proteins involved in NHEJ (53BP1: 3.03 × 104; XRCC5: 2.62 × 104; XRCC6: 5.05 × 105 molecules/cell) were more than an order of magnitude higher than that involved in HR (RAD51: 2500 molecules/cell). Furthermore, we analyzed the dynamics of MDC1 and γH2AX proteins in response to DNA damage induced by the unstable agent neocarzinostatin (NCS). Using cell biochemical fractionation, cells were collected and analyzed at different time points after NCS exposure. Results showed that γH2AX in chromatin fraction peaked at 1 h post-exposure and gradually decreased, while MDC1 translocated from the isotonic to the hypertonic fraction, peaking at 1 hour as well. The study suggests increased MDC1 affinity for chromatin through binding to γH2AX induced by DNA damage. The γH2AX-bound MDC1 (in the hypertonic fraction) to γH2AX ratio at 1 h post-exposure was 1:56.4, with lower MDC1 levels which may attributed to competition with other proteins. CONCLUSIONS: The approach provided quantitative insights into protein dynamics in DNA damage response.
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Direct DNA sequencing can be used for characterizing mutagenicity in simple and complex biological models. Recently we described a method of whole-genome sequencing for detecting mutations in simple models of cultured bacteria, mammalian cells, and nematode. In the current proof-of-concept study, we expand and improve our method for evaluating a more complex mammalian biological model in outbred mice. We detail the method by applying it to a small set of animals treated with a mutagen with known mutagenicity profiles, N-ethyl-N-nitrosourea (ENU), for consistency with the known data. Whole-genome high-fidelity sequencing (HiFi Sequencing) showed frequencies and spectra of background mutations in tissues of untreated mice that were consistent with normal ageing and characterized by spontaneous or enzymatic deamination of 5-methylcytosine. In mice treated with a single 40 mg/kg dose of ENU, the frequency of mutations in the genomic DNA of solid tissues increased up to 7-fold, with the greatest increase observed in the spleen and the smallest increase in the liver. The most common mutations detected in ENU-treated mice were T > A transitions and T > C transversions, consistent with the types of mutations caused by alkylating agents. The data suggest that HiFi Sequencing may be useful for characterizing mutagenicity of novel compounds in various biological models.
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Alquilantes , Mutágenos , Ratones , Animales , Mutágenos/toxicidad , Pruebas de Mutagenicidad , Mutagénesis , Mutación , Etilnitrosourea/toxicidad , ADN , MamíferosRESUMEN
BACKGROUND: Chronic inflammation induces DNA damage and promotes cell proliferation, thereby increasing the risk of cancer. DNA polymerase κ (Pol κ), involved in translesion DNA synthesis, counteracts mutagenesis induced by inflammation in the colon of mice. In the present study, we examined whether Pol κ suppressed inflammation-induced colon tumorigenesis by treating inactivated Polk knock-in (Polk-/-) mice with dextran sulfate sodium (DSS), an inducer of colon inflammation. RESULTS: Male and female Polk-/- and Polk+/+ mice were administered 2% DSS in drinking water for six consecutive days, succeeded via a recovery period of 16 days, followed by 2% DSS for another two days. DSS treatment strongly induced colitis, and the severity of colitis was higher in Polk-/- mice than in Polk+/+ mice. The mice were sacrificed after 19 weeks from the initiation of the first DSS treatment and subjected to pathological examination and mutation analysis. DSS treatment induced colonic dysplasia, and the multiplicity of dysplasia was higher in Polk-/- mice than in Polk+/+mice. Some of the dysplasias in Polk-/- mice exhibited ß-catenin-stained nucleus and/or cytoplasm. Mutation frequencies in the gpt reporter gene were increased by DSS treatment in Polk-/- mice, and were higher than those in Polk+/+ mice. CONCLUSIONS: Pol κ suppresses inflammation and inflammation-induced dysplasia as well as inflammation-induced mutagenesis. The possible mechanisms by which Pol κ suppresses colitis- and colitis-induced dysplasia are discussed.
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AIM: Mutation spectrum of TP53 in gastric cancer (GC) has been investigated world-widely, but a comparison of mutation spectrum among GCs from various regions in the world are still sparsely documented. In order to identify the difference of TP53 mutation spectrum in GCs in Eastern Europe and in East Asia, we sequenced TP53 in GCs from Eastern Europe, Lujiang (China), and Yokohama, Kanagawa (Japan) and identified the feature of TP53 mutations of GC in these regions. SUBJECTS AND METHOD: In total, 689 tissue samples of GC were analyzed: 288 samples from East European populations (25 from Hungary, 71 from Poland and 192 from Romania), 268 from Yokohama, Kanagawa, Japan and 133 from Lujiang, Anhui province, China. DNA was extracted from FFPE tissue of Chinese, East European cases; and from frozen tissue of Japanese GCs. PCR products were direct-sequenced by Sanger method, and in ambiguous cases, PCR product was cloned and up to 8 clones were sequenced. We used No. NC_000017.11(hg38) as the reference sequence of TP53. Mutation patterns were categorized into nine groups: six base substitutions, insertion, deletion and deletion-insertion. Within G:C > A:T mutations the mutations in CpG and non-CpG sites were divided. The Cancer Genome Atlas data (TCGA, ver.R20, July, 2019) having somatic mutation list of GCs from Whites, Asians, and other ethnicities were used as a reference for our data. RESULTS: The most frequent base substitutions were G:C > A:T transition in all the areas investigated. The G:C > A:T transition in non-CpG sites were prominent in East European GCs, compared with Asian ones. Mutation pattern from TCGA data revealed the same trend between GCs from White (TCGA category) vs Asian countries. Chinese and Japanese GCs showed higher ratio of G:C > A:T transition in CpG sites and A:T > G:C mutation was more prevalent in Asian countries. CONCLUSION: The divergence in mutation spectrum of GC in different areas in the world may reflect various pathogeneses and etiologies of GC, region to region. Diversified mutation spectrum in GC in Eastern Europe may suggest GC in Europe has different carcinogenic pathway of those from Asia.
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BACKGROUND: Acrylamide (AA) is classified as "probably carcinogenic to humans (class 2A)" by the International Agency for Research on Cancer. AA causes cancer owing to its mutagenic and genotoxic metabolite, glycidamide (GA), and its effects on sex hormones. Both AA and GA can interact with hemoglobin to hemoglobin adducts (HbAA and HbGA, respectively), which are considered appropriate biomarkers of internal exposure of AA. However, few epidemiologic studies reported an association of HbAA and HbGA with breast cancer. METHODS: We conducted a nested case-control study within the Japan Public Health Center-based Prospective Study cohort (125 cases and 250 controls). Cases and controls were categorized into tertiles (lowest, middle, and highest) using the distribution of HbAA or HbGA levels in the control group and estimated ORs and 95% confidence intervals (CI) using conditional logistic regression, adjusting for potential confounders. RESULTS: No association was observed between HbAA (ORHighestvs.Lowest, 1.34; 95% CI, 0.69-2.59), HbGA (ORHighest vs. Lowest, 1.46; 95% CI, 0.79-2.69), their sum HbAA+HbGA (ORHighest vs. Lowest, 1.36; 95% CI, 0.72-2.58) and breast cancer; however, some evidence of positive association was observed between their ratio, HbGA/HbAA, and breast cancer (ORHighest vs. Lowest, 2.19; 95% CI, 1.11-4.31). CONCLUSIONS: There was no association between biomarkers of AA and breast cancer. IMPACT: It is unlikely that AA increases breast cancer risk; however, the association of AA with breast cancer may need to be evaluated, with a focus not only on the absolute amount of HbAA or HbGA but also on HbGA/HbAA and the activity of metabolic genes.
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Neoplasias de la Mama , Humanos , Femenino , Estudios de Casos y Controles , Acrilamida , Estudios Prospectivos , Pueblos del Este de Asia , Compuestos Epoxi/metabolismo , Hemoglobinas/análisis , Biomarcadores , Modelos LogísticosRESUMEN
Antibody maturation is the central function of the adaptive immune response. This process is driven by the repetitive selection of mutations that increase the affinity toward antigens. We hypothesized that a precise observation of this process by high-throughput sequencing along the time course of immunization will enable us to predict the antibodies reacting to the immunized antigen without any additional in vitro screening. An alpaca was immunized with IgG fragments using multiple antigen injections, and the antibody repertoire development was traced via high-throughput sequencing periodically for months. The sequences were processed into clusters, and the antibodies in the 16 most abundant clusters were generated to determine whether the clusters included antigen-binding antibodies. The sequences of most antigen-responsive clusters resembled those of germline cells in the early stages. These sequences were observed to accumulate significant mutations and also showed a continuous sequence turnover throughout the experimental period. The foregoing characteristics gave us >80% successful prediction of clusters composed of antigen-responding VHHs against IgG fragment. Furthermore, when the prediction method was applied to the data from other alpaca immunized with epidermal growth factor receptor, the success rate exceeded 80% as well, confirming the general applicability of the prediction method. Superior to previous studies, we identified the immune-responsive but very rare clusters or sequences from the immunized alpaca without any empirical screening data.
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Camélidos del Nuevo Mundo , Anticuerpos de Dominio Único , Animales , Inmunización , Vacunación , Antígenos , Inmunoglobulina GRESUMEN
NAD+ synthesis is a fundamental process in living cells. The effects of local metabolite production on chromatin influence the epigenetic status of chromatin in DNA metabolism. We have previously shown that K5 acetylation of H2AX by TIP60 is required for the ADP ribosylation activity of PARP-1, for histone H2AX exchange at DNA damage sites. However, the detailed molecular mechanism has remained unclear. Here, we identified de novo NAD synthetase 1 (NAD syn1) as a novel binding partner to H2AX. The enzymatic activity of NAD syn1 is crucial for the ADP ribosylation activity of PARP-1 for the H2AX dynamics at sites of DNA damage. Inhibition of the NAD synthetase activity in the cell nucleus decreased the overall cellular NAD+ concentration, leading to cellular senescence. Accordingly, the acetylation-dependent H2AX dynamics and homologous recombination repair were suppressed, leading to increased tumorigenesis. Our findings have revealed the importance of de novo NAD+ production in the cell nucleus for protection against the decreased DNA repair capacity caused by cellular senescence and thus against tumorigenesis.
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Histonas , NAD , Humanos , Histonas/metabolismo , NAD/metabolismo , Inhibidores de Poli(ADP-Ribosa) Polimerasas , Reparación del ADN , Cromatina , Daño del ADN , Núcleo Celular/metabolismo , Senescencia Celular , CarcinogénesisRESUMEN
We isolated a phage infecting a member of the Sphingomonadaceae family from a freshwater lake. The phage has a DNA genome of 41,771 bp, with a GC content of 61.7%. The genome harbors 50 predicted protein-coding genes and an auxiliary metabolic gene, which encodes a protein belonging to the radical S-adenosylmethionine superfamily.
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We isolated two bacterial strains (Sphingomonadaceae family) from Lake Biwa, Japan. Based on whole-genome sequencing results, one strain (BSN-002) was assigned to the Sphingopyxis genus and the other (BSN-004) to Sphingomonas aquatilis.
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The XPC protein complex plays a central role in DNA lesion recognition for global genome nucleotide excision repair (GG-NER). Lesion recognition can be accomplished in either a UV-DDB-dependent or -independent manner; however, it is unclear how these sub-pathways are regulated in chromatin. Here, we show that histone deacetylases 1 and 2 facilitate UV-DDB-independent recruitment of XPC to DNA damage by inducing histone deacetylation. XPC localizes to hypoacetylated chromatin domains in a DNA damage-independent manner, mediated by its structurally disordered middle (M) region. The M region interacts directly with the N-terminal tail of histone H3, an interaction compromised by H3 acetylation. Although the M region is dispensable for in vitro NER, it promotes DNA damage removal by GG-NER in vivo, particularly in the absence of UV-DDB. We propose that histone deacetylation around DNA damage facilitates the recruitment of XPC through the M region, contributing to efficient lesion recognition and initiation of GG-NER.
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OBJECTIVES: Carbapenemase-producing Enterobacterales (CPE) pose serious threats to public health. Compared with clinical CPE, the genetic characteristics of environmental CPE are not well understood. This study aimed to characterize the genetic determinants of carbapenem resistance in CPE isolated from environmental waters in Japan. METHODS: Eighty-five water samples were collected from rivers and a lake in Japan. CPE were identified using selective media, and genome sequencing was performed for the obtained isolates (n = 21). RESULTS: Various rare/novel carbapenemases were identified: GES-5 in Raoultella planticola (n = 1), FRI-8 and FRI-11 in Enterobacter spp. (n = 8), IMI-22 and IMI-23 in Serratia ureilytica (n = 3), and SFC-1, SFC-2 and SFH-1 in Serratia fonticola (n = 9). Genomes of 11 isolates could be closed, allowing the elucidation of the genetic contexts of the carbapenemase genes. The blaGES-5 gene was located within a class 1 integron, In2071 (cassette array, blaGES-5-aacA3-aadA16), on a 33â kb IncP6 plasmid. The blaFRI-8 genes were carried on IncFII(Yp) plasmids ranging in size from 191â kb to 244â kb, and the blaFRI-11 genes were carried on 70â kb and 74â kb IncFII(pECLA)/IncR plasmids. The blaIMI-22 and blaIMI-23 genes were co-located on a 107â kb plasmid. The blaSFC and blaSFH-1 genes were found on putative genomic islands inserted at tRNA-Phe genes in chromosomes. CONCLUSIONS: This study revealed the presence of rare/novel carbapenemases among CPE in aquatic environments, suggesting that the environment may act as a potential reservoir of these minor carbapenemases.
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Proteínas Bacterianas , beta-Lactamasas , Antibacterianos/farmacología , Proteínas Bacterianas/genética , Japón , Pruebas de Sensibilidad Microbiana , Plásmidos , beta-Lactamasas/genéticaRESUMEN
Lysine N-pyrrolation, a posttranslational modification, which converts lysine residues to Nε-pyrrole-L-lysine, imparts electronegative properties to proteins, causing them to mimic DNA. Apolipoprotein E (apoE) has been identified as a soluble receptor for pyrrolated proteins (pyrP), and accelerated lysine N-pyrrolation has been observed in apoE-deficient (apoE-/-) hyperlipidemic mice. However, the impact of pyrP accumulation consequent to apoE deficiency on the innate immune response remains unclear. Here, we investigated B-1a cells known to produce germline-encoded immunoglobulin M (IgM) from mice deficient in apoE and identified a particular cell population that specifically produces IgM antibodies against pyrP and DNA. We demonstrated an expansion of B-1a cells involved in IgM production in the peritoneal cavity of apoE-/- mice compared with wild-type mice, consistent with a progressive increase of IgM response in the mouse sera. We found that pyrP exhibited preferential binding to B-1a cells and facilitated the production of IgM. B cell receptor analysis of pyrP-specific B-1a cells showed restricted usage of gene segments selected from the germline gene set; most sequences contained high levels of non-templated-nucleotide additions (N-additions) that could contribute to junctional diversity of B cell receptors. Finally, we report that a subset of monoclonal IgM antibodies against pyrP/DNA established from the apoE-/- mice also contained abundant N-additions. These results suggest that the accumulation of pyrP due to apoE deficiency may influence clonal diversity in the pyrP-specific B cell repertoire. The discovery of these unique B-1a cells for pyrP/DNA provides a key link connecting covalent protein modification, lipoprotein metabolism, and innate immunity.
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Apolipoproteínas E , Subgrupos de Linfocitos B , ADN , Animales , Apolipoproteínas E/deficiencia , Apolipoproteínas E/genética , Apolipoproteínas E/metabolismo , Subgrupos de Linfocitos B/metabolismo , ADN/genética , ADN/metabolismo , Inmunoglobulina M/metabolismo , Lisina/metabolismo , Ratones , Receptores de Antígenos de Linfocitos BRESUMEN
We have recently discovered Japanese children with a novel Fanconi anemia-like inherited bone marrow failure syndrome (IBMFS). This disorder is likely caused by the loss of a catabolic system directed toward endogenous formaldehyde due to biallelic variants in ADH5 combined with a heterozygous ALDH2*2 dominant-negative allele (rs671), which is associated with alcohol-induced Asian flushing. Phytohemagglutinin-stimulated lymphocytes from these patients displayed highly increased numbers of spontaneous sister chromatid exchanges (SCEs), reflecting homologous recombination repair of formaldehyde damage. Here, we report that, in contrast, patient-derived fibroblasts showed normal levels of SCEs, suggesting that different cell types or conditions generate various amounts of formaldehyde. To obtain insights about endogenous formaldehyde production and how defects in ADH5/ALDH2 affect human hematopoiesis, we constructed disease model cell lines, including induced pluripotent stem cells (iPSCs). We found that ADH5 is the primary defense against formaldehyde, and ALDH2 provides a backup. DNA repair capacity in the ADH5/ALDH2-deficient cell lines can be overwhelmed by exogenous low-dose formaldehyde, as indicated by higher levels of DNA damage than in FANCD2-deficient cells. Although ADH5/ALDH2-deficient cell lines were healthy and showed stable growth, disease model iPSCs displayed drastically defective cell expansion when stimulated into hematopoietic differentiation in vitro, displaying increased levels of DNA damage. The expansion defect was partially reversed by treatment with a new small molecule termed C1, which is an agonist of ALDH2, thus identifying a potential therapeutic strategy for the patients. We propose that hematopoiesis or lymphocyte blastogenesis may entail formaldehyde generation that necessitates elimination by ADH5/ALDH2 enzymes.
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Aldehído Deshidrogenasa Mitocondrial/genética , Síndromes Congénitos de Insuficiencia de la Médula Ósea/genética , Anemia de Fanconi/genética , Células Madre Pluripotentes Inducidas/patología , Sistemas CRISPR-Cas , Línea Celular , Células Cultivadas , Síndromes Congénitos de Insuficiencia de la Médula Ósea/diagnóstico , Síndromes Congénitos de Insuficiencia de la Médula Ósea/patología , Daño del ADN , Anemia de Fanconi/diagnóstico , Anemia de Fanconi/patología , Eliminación de Gen , Humanos , Células Madre Pluripotentes Inducidas/metabolismo , MutaciónRESUMEN
The levels of hemoglobin adducts of acrylamide (AA-Hb), a biomarker of acrylamide exposure, have not been reported for Japanese subjects. Herein, we determined the AA-Hb levels in a Japanese population and compared them with the estimated dietary intake from the duplicate diet method (DM) and a food frequency questionnaire (FFQ). One-day DM samples, FFQ, and blood samples were collected from 89 participants and analyzed for acrylamide. AA-Hb was analyzed using liquid chromatography tandem mass spectrometry and the N-alkyl Edman method. Participants were divided into tertiles of estimated acrylamide intake and geometric means (GMs) of AA-Hb adjusted for sex and smoking status. A stratified analysis according to smoking status was also performed. The average AA-Hb levels for all participants, never, past, and current smokers were 46, 38, 65, and 86 pmol/g Hb, respectively. GMs of AA-Hb levels in all participants were significantly associated with tertiles of estimated acrylamide intake from DM (p for trend = 0.02) and FFQ (p for trend = 0.04), although no association with smokers was observed. AA-Hb levels reflected smoking status, which were similar to values reported in Western populations, and they were associated with estimated dietary intake of acrylamide when adjusted for sex and smoking status.
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Acrilamida/sangre , Dieta/métodos , Exposición a Riesgos Ambientales/estadística & datos numéricos , Hemoglobinas/análisis , Adulto , Biomarcadores/sangre , Femenino , Humanos , Japón , Masculino , Persona de Mediana Edad , Encuestas y Cuestionarios , Adulto JovenRESUMEN
Alcohol consumption is the key risk factor for the development of esophageal squamous cell carcinoma (ESCC), and acetaldehyde, a metabolite of alcohol, is an alcohol-derived major carcinogen that causes DNA damage. Aldehyde dehydrogenase2 (ALDH2) is an enzyme that detoxifies acetaldehyde, and its activity is reduced by ALDH2 gene polymorphism. Reduction in ALDH2 activity increases blood, salivary and breath acetaldehyde levels after alcohol intake, and it is deeply associated with the development of ESCC. Heavy alcohol consumption in individuals with ALDH2 gene polymorphism significantly elevates the risk of ESCC; however, effective prevention has not been established yet. In this study, we investigated the protective effects of Alda-1, a small molecule ALDH2 activator, on alcohol-mediated esophageal DNA damage. Here, we generated novel genetically engineered knock-in mice that express the human ALDH2*1 (wild-type allele) or ALDH2*2 gene (mutant allele). Those mice were crossed, and human ALDH2*1/*1, ALDH2*1/*2 and ALDH2*2/*2 knock-in mice were established. They were given 10% ethanol for 7 days in the presence or absence of Alda-1, and we measured the levels of esophageal DNA damage, represented by DNA adduct (N2-ethylidene-2'-deoxyguanosine). Alda-1 significantly increased hepatic ALDH2 activity both in human ALDH2*1/*2 and/or ALDH2*2/*2 knock-in mice and reduced esophageal DNA damage levels after alcohol drinking. Conversely, cyanamide, an ALDH2-inhibitor, significantly exacerbated esophageal DNA adduct level in C57BL/6N mice induced by alcohol drinking. These results indicate the protective effects of ALDH2 activation by Alda-1 on esophageal DNA damage levels in individuals with ALDH2 gene polymorphism, providing a new insight into acetaldehyde-mediated esophageal carcinogenesis and prevention.
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Consumo de Bebidas Alcohólicas/efectos adversos , Aldehído Deshidrogenasa Mitocondrial/metabolismo , Benzamidas/administración & dosificación , Benzodioxoles/administración & dosificación , Carcinogénesis/efectos de los fármacos , Neoplasias Esofágicas/prevención & control , Carcinoma de Células Escamosas de Esófago/prevención & control , Acetaldehído/metabolismo , Acetaldehído/toxicidad , Aldehído Deshidrogenasa Mitocondrial/antagonistas & inhibidores , Aldehído Deshidrogenasa Mitocondrial/genética , Animales , Carcinogénesis/inducido químicamente , Carcinogénesis/genética , Cianamida/administración & dosificación , Aductos de ADN/efectos de los fármacos , Daño del ADN/efectos de los fármacos , Mucosa Esofágica/efectos de los fármacos , Mucosa Esofágica/patología , Neoplasias Esofágicas/etiología , Neoplasias Esofágicas/patología , Carcinoma de Células Escamosas de Esófago/etiología , Carcinoma de Células Escamosas de Esófago/patología , Etanol/metabolismo , Etanol/toxicidad , Técnicas de Sustitución del Gen , Humanos , Masculino , Ratones Transgénicos , Mutación , Neoplasias Experimentales/etiología , Neoplasias Experimentales/patología , Neoplasias Experimentales/prevención & control , Polimorfismo Genético , Factores de RiesgoRESUMEN
Esophageal cancer is prevalent in Cixian, China, but the etiology of this disease remains largely unknown. Therefore, we explored this by conducting a DNA adductome analysis. Both tumorous and nontumorous tissues were collected from patients who underwent surgical procedures at Cixian Cancer Hospital and the Fourth Hospital of Hebei Medical University, which is in a low-incidence area. N2-(3,4,5,6-Tetrahydro-2H-pyran-2-yl)deoxyguanosine (THP-dG) was the major adduct detected in samples from esophageal cancer patients in Cixian. The precursor of THP-dG, N-nitrosopiperidine (NPIP), exhibited a strong mutagenic activity under metabolic activation in the Ames test and a significant dose-dependent increase in mutation frequency during an in vivo mutagenicity test with guanine phosphoribosyltransferase (gpt) delta rats. The NPIP-induced mutation was dominated by A:T to C:G transversions, followed by G:C to A:T and A:T to G:C transitions, in the liver and esophagus of animal samples. A similar mutational pattern was observed in the mutational signature of esophageal cancer patients that demonstrated weak correlation with THP-dG levels. These findings suggested that NPIP exposure is partly involved in the development of esophageal cancer in Cixian residents.