RESUMEN
Magnaporthe oryzae is a rice blast pathogen that seriously threatens rice yield. Benzovindiflupyr is a succinate dehydrogenase inhibitor (SDHI) fungicide that effectively controls many crop diseases. Benzovindiflupyr has a strong inhibitory effect on M. oryzae; however, control of rice blast by benzovindiflupyr and risk of resistance to benzovindiflupyr are not well studied in this pathogen. In this study, six benzovindiflupyr-resistant strains were obtained by domestication induced in the laboratory. The MoSdhBH245D mutation was the cause of M. oryzae resistance to benzovindiflupyr, which was verified through succinate dehydrogenase (SDH) activity assays, molecular docking, and site-specific mutations. Survival fitness analysis showed no significant difference between the benzovindiflupyr-resistant and parent strains. Positive cross-resistance to benzovindiflupyr and other SDHIs and negative cross-resistance to azoxystrobin were observed. Therefore, the risk of benzovindiflupyr resistance in M. oryzae might be medium to high. It should be combined with other classes of fungicides (tebuconazole and azoxystrobin) to slow the development of resistance.
Asunto(s)
Farmacorresistencia Fúngica , Fungicidas Industriales , Mutación , Succinato Deshidrogenasa , Succinato Deshidrogenasa/genética , Succinato Deshidrogenasa/antagonistas & inhibidores , Fungicidas Industriales/farmacología , Farmacorresistencia Fúngica/genética , Enfermedades de las Plantas/microbiología , Magnaporthe/efectos de los fármacos , Magnaporthe/genética , Simulación del Acoplamiento Molecular , Oryza/microbiología , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Estrobilurinas/farmacología , AscomicetosRESUMEN
A CRISPR/Cas12a-coupled multiplexed strand displacement amplification (CMSDA) for the detection of miR155 has been developed. Non-specific amplification was avoided by designing a single-stranded DNA template with a hairpin structure. The detection target miR155 was used as a primer to initiate a multiple-strand displacement reaction to produce abundant ssDNA. ssDNA was recognized by the Cas12a/CrRNA binary complex, activating the trans-cleaving activity of Cas12a. The multiple-strand displacement reaction is more efficiently detected compared with a single-strand displacement reaction. The detection range is from 250 pM to 1 nM, and the limit of the detection is 6.5 pM. The proposed method showed a good applicability in complex serum environments, indicating that the method has a broad prospect for disease detection and clinical application. In addition, we designed a dual-cavity PCR tube, which realized one-tube detection of miRNA155 and avoided open-cap contamination.
Asunto(s)
Sistemas CRISPR-Cas , MicroARNs , MicroARNs/análisis , MicroARNs/sangre , MicroARNs/genética , Humanos , Sistemas CRISPR-Cas/genética , ADN de Cadena Simple/química , ADN de Cadena Simple/genética , Límite de Detección , Técnicas de Amplificación de Ácido Nucleico/métodos , Reacción en Cadena de la Polimerasa/métodos , Proteínas Bacterianas , Endodesoxirribonucleasas , Proteínas Asociadas a CRISPRRESUMEN
Accurate detection of cancer-associated mRNAs is beneficial to early diagnosis and potential treatment of cancer. Herein, for the first time, we developed a novel CRISPR/Cas12a-powered electrochemical/fluorescent (EC/FL) dual-mode controlled-release homogeneous biosensor for mRNA detection. A functionalized ssDNA P2-capped Fe3O4-NH2 loaded with methylene blue (P2@MB-Fe3O4-NH2) was synthesized as the signal probe, while survivin mRNA was chosen as the target RNA. In the presence of the target mRNA, the nicking endonuclease-mediated rolling circle amplification (NEM-RCA) was triggered to produce significant amounts of ssDNA, activating the collateral activity of Cas12a toward the surrounding single-stranded DNA. Thus, the ssDNA P1 completely complementary to ssDNA P2 was cleaved, resulting in that the ssDNA P2 bio-gate on Fe3O4-NH2 could not be opened due to electrostatic interactions. As a result, there was no or only a little MB in the supernatant after magnetic separation, and the measured EC/FL signal was exceedingly weak. On the contrary, the ssDNA P2 bio-gate was opened, enabling MB to be released into the supernatant, and generating an obvious EC/FL signal. Benefiting from the accuracy of EC/FL dual-mode cross-verification, high amplification efficiency, high specificity of NEM-RCA and CRISPR/Cas12a, and high loading of mesoporous Fe3O4-NH2 on signal molecules, the strategy shows aM-level sensitivity and single-base mismatch specificity. More importantly, the practical applicability of this dual-mode strategy was confirmed by mRNA quantification in complex serum environments and tumor cell lysates, providing a new way for developing a powerful disease diagnosis tool.
Asunto(s)
Técnicas Biosensibles , Sistemas CRISPR-Cas , Sistemas CRISPR-Cas/genética , Preparaciones de Acción Retardada , ARN Mensajero/genética , ARN , Colorantes , ADN de Cadena Simple/genética , Endonucleasas , Inhibidores de Serina ProteinasaRESUMEN
An improved electrochemical sensor has been developed for sensitive detection of the p53 gene based on exponential amplification reaction (EXPAR) and CRISPR/Cas12a. Restriction endonuclease BstNI is introduced to specifically identify and cleave the p53 gene, generating primers to trigger the EXPAR cascade amplification. A large number of amplified products are then obtained to enable the lateral cleavage activity of CRISPR/Cas12a. For electrochemical detection, the amplified product activates Cas12a to digest the designed block probe, which allows the signal probe to be captured by the reduced graphene oxide-modified electrode (GCE/RGO), resulting in an enhanced electrochemical signal. Notably, the signal probe is labeled with large amounts of methylene blue (MB). Compared with traditional endpoint decoration, the special signal probe effectively amplifies the electrochemical signals by a factor of about 15. Experimental results show that the electrochemical sensor exhibits wide ranges from 500 aM to 10 pM and 10 pM to 1 nM, as well as a relatively low limit detection of 0.39 fM, which is about an order of magnitude lower than that of fluorescence detection. Moreover, the proposed sensor shows reliable application capability in real human serum, indicating that this work has great prospects for the construction of a CRISPR-based ultra-sensitive detection platform.
Asunto(s)
Sistemas CRISPR-Cas , Genes p53 , Humanos , Cartilla de ADN , Electrodos , FluorescenciaRESUMEN
Methylenetetrahydrofolate reductase (MTHFR) generates methyltetrahydrofolate for methylation reactions. Severe MTHFR deficiency results in homocystinuria and neurologic impairment. Mild MTHFR deficiency (677C > T polymorphism) increases risk for complex traits, including neuropsychiatric disorders. Although low dietary folate impacts brain development, recent concerns have focused on high folate intake following food fortification and increased vitamin use. Our goal was to determine whether high dietary folate during pregnancy affects brain development in murine offspring. Female mice were placed on control diet (CD) or folic acid-supplemented diet (FASD) throughout mating, pregnancy and lactation. Three-week-old male pups were evaluated for motor and cognitive function. Tissues from E17.5 embryos, pups and dams were collected for choline/methyl metabolite measurements, immunoblotting or gene expression of relevant enzymes. Brains were examined for morphology of hippocampus and cortex. Pups of FASD mothers displayed short-term memory impairment, decreased hippocampal size and decreased thickness of the dentate gyrus. MTHFR protein levels were reduced in FASD pup livers, with lower concentrations of phosphocholine and glycerophosphocholine in liver and hippocampus, respectively. FASD pup brains showed evidence of altered acetylcholine availability and Dnmt3a mRNA was reduced in cortex and hippocampus. E17.5 embryos and placentas from FASD dams were smaller. MTHFR protein and mRNA were reduced in embryonic liver, with lower concentrations of choline, betaine and phosphocholine. Embryonic brain displayed altered development of cortical layers. In summary, high folate intake during pregnancy leads to pseudo-MTHFR deficiency, disturbed choline/methyl metabolism, embryonic growth delay and memory impairment in offspring. These findings highlight the unintended negative consequences of supplemental folic acid.
Asunto(s)
Ácido Fólico/efectos adversos , Homocistinuria/genética , Memoria a Corto Plazo/efectos de los fármacos , Metilenotetrahidrofolato Reductasa (NADPH2)/deficiencia , Espasticidad Muscular/genética , Acetilcolina/genética , Acetilcolina/metabolismo , Animales , ADN (Citosina-5-)-Metiltransferasas/genética , ADN (Citosina-5-)-Metiltransferasas/metabolismo , ADN Metiltransferasa 3A , Dieta/efectos adversos , Femenino , Ácido Fólico/administración & dosificación , Homocistinuria/inducido químicamente , Homocistinuria/patología , Hígado/efectos de los fármacos , Hígado/metabolismo , Trastornos de la Memoria/inducido químicamente , Trastornos de la Memoria/fisiopatología , Metilación , Metilenotetrahidrofolato Reductasa (NADPH2)/genética , Ratones , Espasticidad Muscular/inducido químicamente , Espasticidad Muscular/patología , Embarazo , Trastornos Psicóticos/genética , Trastornos Psicóticos/patologíaRESUMEN
BACKGROUND: In recent decades, many workers from rural areas in China migrated to urban cities in search of a better livelihood. Due to the household registration policy and other financial barriers, more than 40 million of children were left behind in their rural home by their migrated parents in 2015. In this cross-sectional study, we aimed to investigate the influence of being left behind on these children's physical and mental health. METHODS: A self-administered questionnaire was completed by participants about their demographic background and health status. Chi-square Test was conducted to investigate the influence. RESULTS: A total of 1662 participants responded and completed all the questions in the questionnaire. Significant differences existed between left-behind children group and non-left-behind children group on several health issues such as not going to school due to sickness (p = 0.080), completeness of the vaccination scheme (p = 0.036) and feeling of loneliness (p = 0.039). However, regarding symptoms like fever, cough or respiratory difficulties, diarrhea and twitch, as well as mental health problems like unhappiness and insomnia, no significant difference was found. Gender difference was also demonstrated showing that girls were more vulnerable than boys to certain symptoms and emotional problems. CONCLUSION: This study indicated that both being left-behind and gender had an impact on the children's health. It is necessary to further reform the household registration system to improve rights of equal access to employment, education and health resources for workers and their children from rural areas in China.
Asunto(s)
Salud Infantil/estadística & datos numéricos , Emigración e Inmigración/estadística & datos numéricos , Trastornos Mentales/psicología , Población Rural/estadística & datos numéricos , Adolescente , China , Estudios Transversales , Emigración e Inmigración/tendencias , Femenino , Humanos , Masculino , Relaciones Padres-Hijo , Padres/psicología , Factores Sexuales , Encuestas y CuestionariosRESUMEN
Background: Suboptimal folate intake, a risk factor for birth defects, is common even in areas with folate fortification. A polymorphism in methylenetetrahydrofolate dehydrogenase 1 (MTHFD1), R653Q (MTHFD1 c.1958 G > A), has also been associated with increased birth defect risk, likely through reduced purine synthesis. Objective: We aimed to determine if the interaction of MTHFD1 synthetase deficiency and low folate intake increases developmental abnormalities in a mouse model for MTHFD1 R653Q. Methods: Female Mthfd1S+/+ and Mthfd1S+/- mice were fed control or low-folate diets (2 and 0.3 mg folic acid/kg diet, respectively) before mating and during pregnancy. Embryos and placentas were examined for anomalies at embryonic day 10.5. Maternal 1-carbon metabolites were measured in plasma and liver. Results: Delays and defects doubled in litters of Mthfd1S+/- females fed low-folate diets compared to wild-type females fed either diet, or Mthfd1S+/- females fed control diets [P values (defects): diet 0.003, maternal genotype 0.012, diet × maternal genotype 0.014]. These adverse outcomes were associated with placental dysmorphology. Intrauterine growth restriction was increased by embryonic Mthfd1S+/- genotype, folate deficiency, and interaction of maternal Mthfd1S+/- genotype with folate deficiency (P values: embryonic genotype 0.045, diet 0.0081, diet × maternal genotype 0.0019). Despite a 50% increase in methylenetetrahydrofolate reductase expression in low-folate maternal liver (P diet = 0.0007), methyltetrahydrofolate concentration decreased 70% (P diet <0.0001) and homocysteine concentration doubled in plasma (P diet = 0.0001); S-adenosylmethionine decreased 40% and S-adenosylhomocysteine increased 20% in low-folate maternal liver (P diet = 0.002 and 0.0002, respectively). Conclusions: MTHFD1 synthetase-deficient mice are more sensitive to low folate intake than wild-type mice during pregnancy. Reduced purine synthesis due to synthetase deficiency and altered methylation potential due to low folate may increase pregnancy complications. Further studies and individualized intake recommendations may be required for women homozygous for the MTHFD1 R653Q variant.
Asunto(s)
Anomalías Congénitas/etiología , Deficiencia de Ácido Fólico/complicaciones , Ácido Fólico/administración & dosificación , Formiato-Tetrahidrofolato Ligasa/deficiencia , Genotipo , Meteniltetrahidrofolato Ciclohidrolasa/deficiencia , Metilenotetrahidrofolato Deshidrogenasa (NADP)/deficiencia , Enzimas Multifuncionales/deficiencia , Polimorfismo Genético , Complicaciones del Embarazo/etiología , Animales , Metilación de ADN , Dieta , Modelos Animales de Enfermedad , Femenino , Desarrollo Fetal , Retardo del Crecimiento Fetal/etiología , Ácido Fólico/sangre , Deficiencia de Ácido Fólico/sangre , Deficiencia de Ácido Fólico/genética , Deficiencia de Ácido Fólico/metabolismo , Formiato-Tetrahidrofolato Ligasa/genética , Formiato-Tetrahidrofolato Ligasa/metabolismo , Ligasas , Hígado/metabolismo , Meteniltetrahidrofolato Ciclohidrolasa/genética , Meteniltetrahidrofolato Ciclohidrolasa/metabolismo , Metilenotetrahidrofolato Deshidrogenasa (NADP)/genética , Metilenotetrahidrofolato Deshidrogenasa (NADP)/metabolismo , Metilenotetrahidrofolato Reductasa (NADPH2)/metabolismo , Ratones , Enzimas Multifuncionales/genética , Enzimas Multifuncionales/metabolismo , Placenta , Embarazo , Complicaciones del Embarazo/sangre , Complicaciones del Embarazo/genética , Complicaciones del Embarazo/metabolismo , Preñez , S-Adenosilhomocisteína/metabolismo , S-Adenosilmetionina/metabolismo , Tetrahidrofolatos/sangreRESUMEN
Cellulose nanofibrils (CNFs) are becoming increasingly ubiquitous in diverse technologies requiring sustainable nanoscale species to form or modify films. The objective of the present study is to investigate the swelling behavior and accompanying free volume of self-standing TEMPO-oxidized (TO) CNF films in the presence of water vapor. For this purpose, we have performed time-resolved swelling experiments on films, prepared according to different experimental protocols, at 90% relative humidity (RH) and ambient temperature. Corresponding free-volume characteristics are elucidated by positron annihilation lifetime spectroscopy (PALS) conducted at ambient temperature and several RH levels. Increasing the drying temperature of the films (from ambient to 50 °C) is observed to promote an increase in film density, which serves to reduce bulk swelling. These elevated drying temperatures likewise cause the free-volume pore size measured by PALS to decrease, while the corresponding total free-volume fraction remains nearly constant. Similarly, dispersion of TO-CNF into aqueous suspensions by ultrasonication prior to film formation increases both the total free-volume fraction and pore size but reduces the size of individual nanofibrils with little net change in bulk swelling. The swelling and concurrent free-volume measurements reported here generally reveal an increase in the free volume of TO-CNF films with increasing RH.
Asunto(s)
Celulosa/química , Óxidos N-Cíclicos/química , Membranas Artificiales , Nanofibras/química , Oxidación-ReducciónRESUMEN
Block ionomers can, in the same fashion as their neutral block copolymer analogs, microphase-order into various nanoscale morphologies. The added benefit of a copolymer possessing a charged species is that the resultant block ionomer becomes amphiphilic and capable of imbibing polar liquids, including water. This characteristic facilitates incorporation of metallic species into the soft nanostructure for a wide range of target applications. In this study, the nonpolar and polar constituents of solvent-templated midblock-sulfonated block ionomers (SBIs) are first selectively metallated for complementary morphological analysis. Next, four different salts, with cationic charges ranging from +1 to +3, are introduced into three hydrated SBIs varying in their degree of sulfonation (DOS), and cation uptake is measured as a function of immersion time. These results indicate that uptake generally increases with increasing salt concentration, cationic charge, and specimen DOS. Swelling and nanoindentation measurements conducted at ambient temperature demonstrate that water uptake decreases, while the surface modulus increases, with increasing cationic charge. Chemical spectra acquired from energy-dispersive X-ray spectroscopy (EDS) confirm the presence of each of the ion-exchanged species, and corresponding EDS chemical maps reveal that the spatial distribution of these species is relatively uniform throughout the block ionomer films.
Asunto(s)
Compuestos Organometálicos/química , Ácidos Sulfónicos/química , Estructura Molecular , Tamaño de la Partícula , Propiedades de SuperficieRESUMEN
The common R653Q variant (â¼20% homozygosity in Caucasians) in the synthetase domain of the folate-metabolizing enzyme MTHFD1 reduces purine synthesis. Although this variant does not appear to affect risk for colorectal cancer, we questioned whether it would affect growth of colorectal tumors. We induced tumor formation in a mouse model for MTHFD1-synthetase deficiency (Mthfd1S+/- ) using combined administration of azoxymethane (AOM) and dextran sodium sulfate (DSS) in male and female wild-type and Mthfd1S+/- mice. Tumor size was significantly smaller in MthfdS+/- mice, particularly in males. A reduction in the proliferation of MthfdS+/- mouse embryonic fibroblast cell lines, compared with wild-type lines, was also observed. Tumor number was not influenced by genotype. The amount of inflammation observed within tumors from male Mthfd1S+/- mice was lower than that in wild-type mice. Gene expression analysis in tumor adjacent normal (pre-neoplastic) tissue identified several genes involved in proliferation (Fosb, Fos, Ptk6, Esr2, Atf3) and inflammation (Atf3, Saa1, TNF-α) that were downregulated in MthfdS+/- males. In females, MthfdS+/- genotype was not associated with these gene expression changes, or with differences in tumor inflammation. These findings suggest that the mechanisms directing tumor growth differ significantly between males and females. We suggest that restriction of purine synthesis, reduced expression of genes involved in proliferation, and/or reduced inflammation lead to slower tumor growth in MTHFD1-synthetase deficiency. These findings may have implications for CRC tumor growth and prognosis in individuals with the R653Q variant. © 2016 Wiley Periodicals, Inc.
Asunto(s)
Aminohidrolasas/deficiencia , Neoplasias Colorrectales/patología , Formiato-Tetrahidrofolato Ligasa/deficiencia , Meteniltetrahidrofolato Ciclohidrolasa/deficiencia , Metilenotetrahidrofolato Deshidrogenasa (NADP)/deficiencia , Metilenotetrahidrofolato Deshidrogenasa (NADP)/genética , Antígenos de Histocompatibilidad Menor/genética , Complejos Multienzimáticos/deficiencia , Enzimas Multifuncionales/deficiencia , Polimorfismo de Nucleótido Simple , Animales , Azoximetano/efectos adversos , Proliferación Celular , Células Cultivadas , Neoplasias Colorrectales/inducido químicamente , Neoplasias Colorrectales/genética , Sulfato de Dextran/efectos adversos , Femenino , Regulación Neoplásica de la Expresión Génica , Humanos , Masculino , RatonesRESUMEN
BACKGROUND: A single nucleotide polymorphism (SNP) in the synthetase domain of the trifunctional folate-dependent enzyme MTHFD1 (c.1958G>A, R653Q) has been linked to adverse pregnancy outcomes, neural tube defects, and possibly congenital heart defects. Maternal folate deficiency may also modify the risk associated with these disorders. We recently established a mouse model with a mild deficiency of 10-formyltetrahydrofolate synthetase activity in MTHFD1 (Mthfd1S(+/-) mice) to investigate disorders associated with SNPs in this gene. The effect of synthetase deficiency on embryonic heart development has not yet been examined. METHODS: Female Mthfd1S(+/+) and (+/-) mice were placed on control and folate-deficient diets for 6 weeks before mating to Mthfd1S(+/-) males. Embryos and placentae were collected at embryonic day 14.5. Embryos were evaluated for congenital heart defects by histological examination. RESULTS: Embryonic Mthfd1S(+/-) genotype was associated with an increased incidence of heart defects, primarily ventricular septal defects. Other markers of embryonic development (crown-rump length, embryonic weight, embryonic delay, placental weight, and thickness of the ventricular myocardium) were not affected by embryonic genotype. Maternal genotype and diet did not have a significant effect on these outcomes. CONCLUSION: Deficiency of the MTHFD1 10-formyltetrahydrofolate synthetase activity in embryos is associated with increased incidence of congenital heart defects.
Asunto(s)
Aminohidrolasas/genética , Modelos Animales de Enfermedad , Formiato-Tetrahidrofolato Ligasa/genética , Cardiopatías Congénitas/genética , Metilenotetrahidrofolato Deshidrogenasa (NADP)/genética , Complejos Multienzimáticos/genética , Animales , Femenino , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones NoqueadosRESUMEN
Hyperhomocysteinaemia can contribute to cognitive impairment and brain atrophy. MTRR (methionine synthase reductase) activates methionine synthase, which catalyses homocysteine remethylation to methionine. Severe MTRR deficiency results in homocystinuria with cognitive and motor impairments. An MTRR polymorphism may influence homocysteine levels and reproductive outcomes. The goal of the present study was to determine whether mild hyperhomocysteinaemia affects neurological function in a mouse model with Mtrr deficiency. Mtrr+/+, Mtrr+/gt and Mtrrgt/gt mice (3 months old) were assessed for short-term memory, brain volumes and hippocampal morphology. We also measured DNA methylation, apoptosis, neurogenesis, choline metabolites and expression of ChAT (choline acetyltransferase) and AChE (acetylcholinesterase) in the hippocampus. Mtrrgt/gt mice exhibited short-term memory impairment on two tasks. They had global DNA hypomethylation and decreased choline, betaine and acetylcholine levels. Expression of ChAT and AChE was increased and decreased respectively. At 3 weeks of age, they showed increased neurogenesis. In the cerebellum, mutant mice had DNA hypomethylation, decreased choline and increased expression of ChAT. Our work demonstrates that mild hyperhomocysteinaemia is associated with memory impairment. We propose a mechanism whereby a deficiency in methionine synthesis leads to hypomethylation and compensatory disturbances in choline metabolism in the hippocampus. This disturbance affects the levels of acetylcholine, a critical neurotransmitter in learning and memory.
Asunto(s)
Cerebelo/metabolismo , Ferredoxina-NADP Reductasa/genética , Hipocampo/metabolismo , Homocistinuria/metabolismo , Hiperhomocisteinemia/metabolismo , Memoria a Corto Plazo , Acetilcolinesterasa/genética , Acetilcolinesterasa/metabolismo , Animales , Apoptosis , Betaína/metabolismo , Cerebelo/patología , Colina/metabolismo , Colina O-Acetiltransferasa/genética , Colina O-Acetiltransferasa/metabolismo , Metilación de ADN , Modelos Animales de Enfermedad , Ferredoxina-NADP Reductasa/deficiencia , Expresión Génica , Hipocampo/patología , Homocisteína/metabolismo , Homocistinuria/genética , Homocistinuria/patología , Hiperhomocisteinemia/genética , Hiperhomocisteinemia/patología , Masculino , Metionina/metabolismo , Ratones , Ratones Noqueados , Transmisión SinápticaRESUMEN
Colorectal cancer risk is increased when dietary folate intake is low, with or without a deficiency in methylenetetrahydrofolate reductase (MTHFR). We have observed that intestinal tumors are induced in mice fed low-folate diets, and that tumor incidence is increased when these mice also have MTHFR deficiency. This study was undertaken to identify differentially expressed proteins in conditions favoring initial steps of murine carcinogenesis in normal preneoplastic intestine. We compared the proteome of BALB/c normal intestine in Mthfr(+/+) mice fed control diets for 1 year (low susceptibility to tumorigenesis) with the proteome of Mthfr(+/-) animals fed low folate diets (higher tumor susceptibility). Our data suggest that the NuRD complex, KRAS-related proteins, the protein synthetic machinery, and fatty acid-related metabolic proteins are upregulated in the early stages of tumorigenesis. These proteins may serve as biomarkers or targets for colorectal cancer diagnosis or therapy.
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Carcinogénesis/metabolismo , Ácido Fólico/metabolismo , Homocistinuria/complicaciones , Neoplasias Intestinales/etiología , Neoplasias Intestinales/metabolismo , Intestinos/patología , Metilenotetrahidrofolato Reductasa (NADPH2)/deficiencia , Espasticidad Muscular/complicaciones , Proteoma/metabolismo , Animales , Carcinogénesis/patología , Dieta , Modelos Animales de Enfermedad , Femenino , Mucosa Intestinal/metabolismo , Neoplasias Intestinales/patología , Metilenotetrahidrofolato Reductasa (NADPH2)/metabolismo , Ratones , Ratones Endogámicos BALB C , Proteómica , Trastornos Psicóticos/complicacionesRESUMEN
Intracellular glucose detection is crucial due to its pivotal role in metabolism and various physiological processes. Precise glucose monitoring holds significance in diabetes management, metabolic studies, and biotechnological applications. In this study, we developed an innovative and expedient cell-permeable nanoreactor for intracellular glucose based on surface-enhanced Raman scattering (SERS). The nanoreactor was designed with gold nanoparticles (AuNPs), which were engineered with glucose oxide (GOx) and a H2O2-responsive Raman reporter 2-mercaptohydroquinone (2-MHQ). The interaction between 2-MHQ and H2O2 generated by glucose and GOx could simultaneously induce the appearance in the peak at 985 cm-1. Our results showed excellent performance in detecting glucose within the concentration range from 0.1 µM to 10 mM, with a low detection limitation of 14.72 nM. In addition, the glucose distribution in single HeLa cells was evaluated by real time SERS mapping. By combining noble metal particles and natural oxidases, the nanoreactor possesses both Raman activity and enzymatic functionality, thus enables sensitive glucose detection and facilitates imaging at a single cell level, which offers an insightful monitoring of cellular processes.
Asunto(s)
Glucosa , Oro , Nanopartículas del Metal , Espectrometría Raman , Espectrometría Raman/métodos , Humanos , Células HeLa , Oro/química , Nanopartículas del Metal/química , Glucosa/análisis , Glucosa/metabolismo , Peróxido de Hidrógeno/análisis , Peróxido de Hidrógeno/química , Glucosa Oxidasa/química , Glucosa Oxidasa/metabolismoRESUMEN
Hypoxia is a hallmark of cancer development. However, the molecular mechanisms by which hypoxia promotes tumor metastasis are not fully understood. In this study, we demonstrate that hypoxia promotes breast cancer metastasis through suppression of ΔNp63α in a HIF1α-independent manner. We show that hypoxia-activated XBP1s forms a stable repressor protein complex with HDAC2 and EZH2 to suppress ΔNp63α transcription. Notably, H3K27ac is predominantly occupied on the ΔNp63 promoter under normoxia, while H3K27me3 on the promoter under hypoxia. We show that XBP1s binds to the ΔNp63 promoter to recruit HDAC2 and EZH2 in facilitating the switch of H3K27ac to H3K27me3. Pharmacological inhibition or the knockdown of either HDAC2 or EZH2 leads to increased H3K27ac, accompanied by the reduced H3K27me3 and restoration of ΔNp63α expression suppressed by hypoxia, resulting in inhibition of cell migration. Furthermore, the pharmacological inhibition of IRE1α, but not HIF1α, upregulates ΔNp63α expression in vitro and inhibits tumor metastasis in vivo. Clinical analyses reveal that reduced p63 expression is correlated with the elevated expression of XBP1, HDAC2, or EZH2, and is associated with poor overall survival in human breast cancer patients. Together, these results indicate that hypoxia-activated XBP1s modulates the epigenetic program in suppression of ΔNp63α to promote breast cancer metastasis independent of HIF1α and provides a molecular basis for targeting the XBP1s/HDAC2/EZH2-ΔNp63α axis as a putative strategy in the treatment of breast cancer metastasis.
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Neoplasias de la Mama , Proteína Potenciadora del Homólogo Zeste 2 , Epigénesis Genética , Histona Desacetilasa 2 , Subunidad alfa del Factor 1 Inducible por Hipoxia , Proteínas Supresoras de Tumor , Proteína 1 de Unión a la X-Box , Humanos , Neoplasias de la Mama/patología , Neoplasias de la Mama/genética , Neoplasias de la Mama/metabolismo , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Proteína Potenciadora del Homólogo Zeste 2/genética , Proteína 1 de Unión a la X-Box/metabolismo , Proteína 1 de Unión a la X-Box/genética , Histona Desacetilasa 2/metabolismo , Histona Desacetilasa 2/genética , Femenino , Subunidad alfa del Factor 1 Inducible por Hipoxia/metabolismo , Subunidad alfa del Factor 1 Inducible por Hipoxia/genética , Proteínas Supresoras de Tumor/metabolismo , Proteínas Supresoras de Tumor/genética , Animales , Línea Celular Tumoral , Metástasis de la Neoplasia , Ratones , Regulación Neoplásica de la Expresión Génica , Factores de Transcripción/metabolismo , Factores de Transcripción/genética , Hipoxia de la Célula/genéticaRESUMEN
Fast, sensitive, and portable detection of genetic modification contributes to agricultural security and food safety. Here, we developed RPA-CRISPR/Cas12a-G-quadruplex colorimetric assays that can combine with intelligent recognition by deep learning algorithms to achieve sensitive, rapid, and portable detection of the CaMV35S promoter. When the crRNA-Cas12a complex recognizes the RPA amplification product, Cas12 cleaves the G-quadruplex, causing the G4-Hemin complex to lose its peroxide mimetic enzyme function and be unable to catalyze the conversion of ABTS2- to ABTS, allowing CaMV35S concentration to be determined based on ABTS absorbance. By utilizing the RPA-CRISPR/Cas12a-G4 assay, we achieved a CaMV35S limit of detection down to 10 aM and a 0.01 % genetic modification sample in 45 min. Deep learning algorithms are designed for highly accurate classification of color results. Yolov5 objective finding and Resnet classification algorithms have been trained to identify trace (0.01 %) CaMV35S more accurately than naked eye colorimetry. We also coupled deep learning algorithms with a smartphone app to achieve portable and rapid photo identification. Overall, our findings enable low cost ($0.43), high accuracy, and intelligent detection of the CaMV35S promoter.
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Sistemas CRISPR-Cas , Colorimetría , Aprendizaje Profundo , G-Cuádruplex , Colorimetría/métodos , Sistemas CRISPR-Cas/genética , Regiones Promotoras Genéticas , Proteínas Asociadas a CRISPR/genética , Proteínas Asociadas a CRISPR/metabolismo , Límite de Detección , Proteínas Bacterianas/genética , EndodesoxirribonucleasasRESUMEN
BACKGROUND: The incidence of neural tube defects has diminished considerably since the implementation of food fortification with folic acid (FA). However, the impact of excess FA intake, particularly during pregnancy, requires investigation. In a recent study, we reported that a diet supplemented with 20-fold higher FA than the recommended intake for rodents had adverse effects on embryonic mouse development at embryonic days (E)10.5 and 14.5. In this report, we examined developmental outcomes in E14.5 embryos after administering a diet supplemented with 10-fold higher FA than recommended to pregnant mice with and without a mild deficiency of methylenetetrahydrofolate reductase (MTHFR). METHODS: Pregnant mice with or without a deficiency in MTHFR were fed a control diet (recommended FA intake of 2 mg/kg diet for rodents) or an FA-supplemented diet (FASD; 10-fold higher than the recommended intake [20 mg/kg diet]). At E14.5, mice were examined for embryonic loss and growth retardation, and hearts were assessed for defects and for ventricular wall thickness. RESULTS: Maternal FA supplementation was associated with embryonic loss, embryonic delays, a higher incidence of ventricular septal defects, and thinner left and right ventricular walls, compared to mothers fed control diet. CONCLUSIONS: Our work suggests that even moderately high levels of FA supplementation may adversely affect fetal mouse development. Additional studies are warranted to evaluate the impact of high folate intake in pregnant women. Birth Defects Research (Part A), 2013. © 2012 Wiley Periodicals, Inc.
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Embrión de Mamíferos/efectos de los fármacos , Desarrollo Embrionario/efectos de los fármacos , Ácido Fólico/toxicidad , Complejo Vitamínico B/toxicidad , Animales , Relación Dosis-Respuesta a Droga , Pérdida del Embrión/inducido químicamente , Femenino , Corazón/efectos de los fármacos , Corazón/embriología , Defectos del Tabique Interventricular/inducido químicamente , Ventrículos Cardíacos/efectos de los fármacos , Ventrículos Cardíacos/embriología , Masculino , Metilenotetrahidrofolato Reductasa (NADPH2)/sangre , Metilenotetrahidrofolato Reductasa (NADPH2)/deficiencia , Ratones , Ratones Endogámicos BALB C , EmbarazoRESUMEN
[Purpose/Significance] Under the influence of various factors such as the pandemic, oil prices, and economic conditions, the global automotive industry has entered a period of downturn. Therefore, how to activate market potential and increase consumers' willingness to purchase cars has become an important research topic. Unlike many other products, test drives play a significant role in the car-buying process. [Method/Procedure] This study employs a questionnaire survey to explore how consumer perceptions of product aesthetics, space quality, and service quality during their initial visit to an automobile 4S store influence their test drive intention through two dimensions of brand image: symbolic and experiential. A structural equation model is used to establish a test drive intention impact model incorporating these dimensions. [Results/Conclusions] The study found that brand image, both symbolic and experiential, plays a significant mediating role in enhancing potential consumers' test drive intentions. Space image had the most significant impact on brand image. Although product aesthetics did not directly affect brand image experientially, they remained an important factor in enhancing brand image symbolically. [Contribution/Value] The results of this study can provide insights for automotive brand managers, automobile 4S store designers, and others aiming to promote the sustainable development of automotive consumption.
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The selective separation of metal species from various sources is highly desirable in applications such as hydrometallurgy, water treatment, and energy production but also challenging. Monovalent cation exchange membranes (CEMs) show a great potential to selectively separate one metal ion over others of the same or different valences from various effluents in electrodialysis. Selectivity among metal cations is influenced by both the inherent properties of membranes and the design and operating conditions of the electrodialysis process. The research progress and recent advances in membrane development and the implication of the electrodialysis systems on counter-ion selectivity are extensively reviewed in this work, focusing on both structure-property relationships of CEM materials and influences of process conditions and mass transport characteristics of target ions. Key membrane properties, such as charge density, water uptake, and polymer morphology, and strategies for enhancing ion selectivity are discussed. The implications of the boundary layer at the membrane surface are elucidated, where differences in the mass transport of ions at interfaces can be exploited to manipulate the transport ratio of competing counter-ions. Based on the progress, possible future R&D directions are also proposed.
RESUMEN
Palladium is a vital commodity in the industry. To guarantee a stable supply in the future, it is imperative to adopt more effective recycling practices. In this proof-of-concept study, we explore the potential of electrodialysis to enhance the palladium concentration in a residual solution of palladium recycling, thus promoting higher recovery rates. Experiments were conducted using an industrial hydrochloric acid solution containing around 1000 mg/L of palladium, with a pH below 1. Two sets of membranes, Selemion AMVN/CMVN and Fujifilm Type 12 AEM/CEM, were tested at two current levels. The Fujifilm membranes, which are designed for low permeability of water, show promising results, recovering around 40% of palladium within a two-hour timeframe. The Selemion membranes were inefficient due to excessive water transport. All membranes accumulated palladium in their structures. Anion-exchange membranes showed higher palladium accumulation at lower currents, while cation-exchange membranes exhibited increased palladium accumulation at higher currents. Owing to the low concentration of palladium and the presence of abundant competing ions, the current efficiency remained below 2%. Our findings indicate a strong potential for augmenting the palladium stage in industrial draw solutions through electrodialysis, emphasizing the importance of membrane properties and process parameters to ensure a viable process. Beyond the prominent criteria of high permselectivity and low resistance, minimizing the permeability of water within IEMs remains a key challenge to mitigating the efficiency loss associated with uncontrolled mixing of the electrolyte solution.