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Aqueous zinc (Zn) chemistry features intrinsic safety, but suffers from severe irreversibility, as exemplified by low Coulombic efficiency, sustained water consumption and dendrite growth, which hampers practical applications of rechargeable Zn batteries. Herein, we report a highly reversible aqueous Zn battery in which the graphitic carbon nitride quantum dots additive serves as fast colloid ion carriers and assists the construction of a dynamic & self-repairing protective interphase. This real-time assembled interphase enables an ion-sieving effect and is found actively regenerate in each battery cycle, in effect endowing the system with single Zn2+ conduction and constant conformal integrality, executing timely adaption of Zn deposition, thus retaining sustainable long-term protective effect. In consequence, dendrite-free Zn plating/stripping at ~99.6% Coulombic efficiency for 200 cycles, steady charge-discharge for 1200 h, and impressive cyclability (61.2% retention for 500 cycles in a Zn | |MnO2 full battery, 73.2% retention for 500 cycles in a Zn | |V2O5 full battery and 93.5% retention for 3000 cycles in a Zn | |VOPO4 full battery) are achieved, which defines a general pathway to challenge Lithium in all low-cost, large-scale applications.
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Along with sexual maturity, the liver undergoes numerous metabolic processes to adapt the physiological changes associated with egg-laying in hens. However, mechanisms regulating the processes were unclear. In this study, comparative hepatic proteome and acetyl-proteome between pre- and peak-laying hens were performed. The results showed that the upregulated proteins were mainly related to lipid and protein biosynthesis, while the downregulated proteins were mainly involved in pyruvate metabolism and were capable of inhibiting gluconeogenesis and lactate synthesis in peak-laying hens compared with that in pre-laying hens. With unchanged expression level, the significant acetylated proteins were largely functioned on activation of polyunsaturated fatty acid oxidation in peroxisome, while the significant deacetylated proteins were principally used to elevate medium and short fatty acid oxidation in mitochondria and oxidative phosphorylation. Most of the proteins which involved in gluconeogenesis, lipid transport, and detoxification were influenced by both protein expression and acetylation. Taken overall, a novel mechanism wherein an alternate source of acetyl coenzyme A was produced by activation of FA oxidation and pyruvate metabolism to meet the increased energy demand and lipid synthesis in liver of laying hens was uncovered. This study provides new insights into molecular mechanism of adaptation to physiological changes in liver of laying hens.
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Hard carbon (HC) is a promising anode material for sodium-ion batteries, but the performance remains unsatisfactory and the sodiation mechanism in HC is one of the most debated topics. Here, from self-assembled cellulose nanocrystal sheets with crystallographic texture, unique HC nanosheets with vertically oriented (002) planes are fabricated and used as a model HC to investigate the sodiation mechanisms using synchrotron scanning transmission X-ray microscopy (STXM) coupled with analytical transmission electron microscopy (TEM). The model HC simplifies the 3D sodiation in a typical HC particle into a 2D sodiation, which facilitates the visualization of phase transformation at different states of charge. The results for the first time unveil that the sodiation in HC initiates heterogeneously, with multiple propagation fronts proceeding simultaneously, eventually merging into larger aggregates. The spatial correlation between the preferential adsorption and nucleation sites suggests that the heterogeneous nucleation is driven by the local Na-ion concentration, which is determined by defects or heteroatoms that have strong binding to Na ions. By identifying intercalation as the dominant sodium storage mechanism in the model HC, the findings highlight the importance of engineering the graphene layer orientation and the structural heterogeneity of edge sites to enhance the performances.
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Egg-laying performance is one of the most important economic traits in the poultry industry. Commercial layers can lay one egg almost every day during their peak-laying period. However, many Chinese indigenous chicken breeds show a relatively low egg-laying rate, even during their peak-laying period. To understand what makes the difference in egg production, we compared the hypothalamus transcriptome profiles of Lushi blue-shelled-egg chickens (LBS), a Chinese indigenous breed with low egg-laying rate and Rhode Island Red chickens (RIR), a commercial layer with relatively high egg-laying rate using RNA-seq. A total of 753 differentially expressed genes (DEGs) were obtained. Of these DEGs, 38 genes were enriched in 2 Gene Ontology (GO) terms, namely reproduction term and the reproductive process term, and 6 KEGG pathways, namely Wnt signaling pathway, Oocyte meiosis, GnRH signaling pathway, Thyroid hormone signaling pathway, Thyroid hormone synthesis and MAPK signaling pathway, which have been long known to be involved in egg production regulation. To further determine the core genes from the 38 DEGs, protein-protein interaction (PPI) network, co-expression network and transcriptional regulatory network analyses were carried out. After integrated analysis and experimental validation, 4 core genes including RAC1, MRE11A, MAP7 and SOX5 were identified as the potential core genes that are responsible for the laying-rate difference between the 2 breeds. These findings paved the way for future investigating the mechanism of egg-laying regulation and enriched the chicken reproductive regulation theory.
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Pollos , Transcriptoma , Animales , Pollos/genética , Femenino , Regulación de la Expresión Génica , Hipotálamo , OviposiciónRESUMEN
BACKGROUND: Estrogen plays an essential role in female development and reproductive function. In chickens, estrogen is critical for lipid metabolism in the liver. The regulatory molecular network of estrogen in chicken liver is poorly understood. To identify estrogen-responsive genes and estrogen functional sites on a genome-wide scale, we determined expression profiles of mRNAs, lncRNAs, and miRNAs in estrogen-treated ((17ß-estradiol)) and control chicken livers using RNA-Sequencing (RNA-Seq) and studied the estrogen receptor α binding sites by ChIP-Sequencing (ChIP-Seq). RESULTS: We identified a total of 990 estrogen-responsive genes, including 962 protein-coding genes, 11 miRNAs, and 17 lncRNAs. Functional enrichment analyses showed that the estrogen-responsive genes were highly enriched in lipid metabolism and biological processes. Integrated analysis of the data of RNA-Seq and ChIP-Seq, identified 191 genes directly targeted by estrogen, including 185 protein-coding genes, 4 miRNAs, and 2 lncRNAs. In vivo and in vitro experiments showed that estrogen decreased the mRNA expression of PPARGC1B, which had been reported to be linked with lipid metabolism, by directly increasing the expression of miR-144-3p. CONCLUSIONS: These results increase our understanding of the functional network of estrogen in chicken liver and also reveal aspects of the molecular mechanism of estrogen-related lipid metabolism.
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MicroARNs , ARN Largo no Codificante , Animales , Pollos/genética , Pollos/metabolismo , Estrógenos , Femenino , Metabolismo de los Lípidos/genética , Hígado/metabolismo , MicroARNs/genética , MicroARNs/metabolismo , ARN Largo no Codificante/metabolismoRESUMEN
Long chain acyl-CoA synthetases (ACSLs), which drive the conversion of long chain fatty acid into acyl-CoA, an ingredient of lipid synthesis, have been well-acknowledged to exert an indispensable role in many metabolic processes in mammals, especially lipid metabolism. However, in chicken, the evolutionary characteristics, expression profiles and regulatory mechanisms of ACSL gene family are rarely understood. Here, we analyzed the genomic synteny, gene structure, evolutionary event and functional domains of the ACSL gene family members using bioinformatics methods. The spatiotemporal expression profiles of ACSL gene family, and their regulatory mechanism were investigated via bioinformatics analysis incorporated with in vivo and in vitro estrogen-treated experiments. Our results indicated that ACSL2 gene was indeed evolutionarily lost in the genome of chicken. Chicken ACSLs shared an AMP-binding functional domain, as well as highly conversed ATP/AMP and FACS signature motifs, and were clustered into two clades, ACSL1/5/6 and ACSL3/4, based on high sequence similarity, similar gene features and conversed motifs. Chicken ACSLs showed differential tissue expression distributions, wherein the significantly decreased expression level of ACSL1 and the significantly increased expression level of ACSL5 were found, respectively, the expression levels of the other ACSL members remained unchanged in the liver of peak-laying hens versus pre-laying hens. Moreover, the transcription activity of ACSL1, ACSL3 and ACSL4 was silenced and ACSL6 was activated by estrogen, but no response to ACSL5. In conclusion, though having highly conversed functional domains, chicken ACSL gene family is organized into two separate groups, ACSL1/5/6 and ACSL3/4, and exhibits varying expression profiles and estrogen effects. These results not only pave the way for better understanding the specific functions of ACSL genes in avian lipid metabolism, but also provide a valuable evidence for gene family characteristics.
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Pollos/genética , Coenzima A Ligasas/genética , Evolución Molecular , Metabolismo de los Lípidos/genética , Familia de Multigenes/genética , Acilcoenzima A/metabolismo , Animales , Células Cultivadas , Embrión de Pollo , Pollos/crecimiento & desarrollo , Pollos/metabolismo , Coenzima A Ligasas/metabolismo , Biología Computacional , Estrógenos/metabolismo , Ácidos Grasos/metabolismo , Femenino , Perfilación de la Expresión Génica , Regulación del Desarrollo de la Expresión Génica , Hepatocitos , Cultivo Primario de Células , Dominios Proteicos/genética , Análisis Espacio-Temporal , SinteníaRESUMEN
The emopamil binding protein (EBP) is an important enzyme participating in the final steps of cholesterol biosynthesis in mammals. A predictive gene EBP-like, which encodes the protein with a high identity to human EBP, was found in chicken genome. No regulatory mechanisms and biological functions of EBP-like have been characterized in chickens. In the present study, the coding sequence of EBP-like was cloned, the phylogenetic trees of EBP/EBP-like were constructed and the genomic synteny of EBP-like was analyzed. The regulatory mechanism of EBP-like were explored with in vivo and in vitro experiments. The biological functions of EBP-like in liver cholesterol biosynthetic were examined by using gain- or loss-of-function strategies. The results showed that chicken EBP-like gene was originated from a common ancestral with Japanese quail EBP gene, and was relatively conservative with EBP gene among different species. The EBP-like gene was highly expressed in liver, its expression level was significantly increased in peak-laying stage, and was upregulated by estrogen. Inhibition of the EBP-like mRNA expression could restrain the expressions of EBP-like downstream genes (SC5D, DHCR24, and DHCR7) in the cholesterol synthetic pathway, therefore downregulate the liver intracellular T-CHO level. In conclusion, as substitute of EBP gene in chickens, EBP-like plays a vital role in the process of chicken liver cholesterol synthesis. This research provides a basis for revealing the molecular regulatory mechanism of cholesterol synthesis in birds, contributes insights into the improvement of the growth and development, laying performance and egg quality in poultry.
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Techniques for scaling-up the direct-current (dc) triboelectricity generation in MoS2 multilayer-based Schottky nanocontacts are vital for exploiting the nanoscale phenomenon for real-world applications of energy harvesting and sensing. Here, we show that scaling-up the dc output can be realized by using various MoS2 multilayer-based heterojunctions including metal/semiconductor (MS), metal/insulator (tens of nanometers)/semiconductor (MIS), and semiconductor/insulator (a few nanometers)/semiconductor (SIS) moving structures. It is shown that the tribo-excited energetic charge carriers can overcome the interfacial potential barrier by different mechanisms, such as thermionic emission, defect conduction, and quantum tunneling in the case of MS, MIS, and SIS moving structures. By tailoring the interface structure, it is possible to trigger electrical conduction resulting in optimized power output. We also show that the band bending in the surface-charged region of MoS2 determines the direction of the dc power output. Our experimental results show that engineering the interface structure opens up new avenues for developing next-generation semiconductor-based mechanical energy conversion with high performance.
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Long noncoding RNAs (lncRNAs) play important roles in transcriptional and posttranscriptional regulation. However, the effects of lncRNAs on the meat quality of chicken hasn't been elucidated clearly yet. Gushi chickens are popular in China because of their superior meat quality, particularly the tender flesh, and unique flavor. Gushi chickens are popular in China because of their superior meat quality, delicate flesh, and unique flavor. We performed RNA-Seq analysis of breast muscle from Gushi chicken at two physiological stages, including juvenile (G20W) and laying (G55W). In total, 186 lncRNAs and 881 mRNAs were differentially expressed between G20W and G55W (fold change ≥ 2.0, P < 0.05). Among them, 131 lncRNAs presented upregulated and 55 were downregulated. We identified the cis and trans target genes of the differentially expressed lncRNAs, and constructed lncRNA-mRNA interaction networks. The results showed that differentially expressed mRNAs and lncRNAs were mainly involved in ECM-receptor interaction, glycerophospholipid metabolism, ubiquitin-mediated proteolysis, and the biosynthesis of amino acids. In summary, our study utilized RNA-seq analysis to predict the functions of lncRNA on chicken meat quality. Furthermore, comprehensive analysis identified lncRNAs and their target genes, which may contribute to a better understanding of the molecular mechanisms underlying in poultry meat quality and provide a theoretical basis for further research.
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Calidad de los Alimentos , Regulación de la Expresión Génica/fisiología , Proteínas de Aves de Corral , ARN Largo no Codificante , ARN Mensajero , Animales , Pollos/genética , Pollos/metabolismo , ARN Largo no Codificante/biosíntesis , ARN Largo no Codificante/genética , ARN Mensajero/biosíntesis , ARN Mensajero/genéticaRESUMEN
Although the generation of mechanical stress in the anode material is suggested as a possible reason for electrode degradation and fading of storage capacity in batteries, only limited knowledge of the electrode stress and its evolution is available at present. Here, we show real-time monitoring of the interfacial stress of a few-layer MoS2 system under the sodiation/desodiation process using microcantilever electrodes. During the first sodiation with a voltage plateau of 1.0 to 0.85 V, the MoS2 exhibits a compressive stress (2.1 Nm-1), which is substantially smaller than that measured (9.8 Nm-1) during subsequent plateaus at 0.85 to 0.4 V due to the differential volume expansion of the MoS2 film. The conversion reaction to Mo below 0.1 V generates an anomalous compressive stress of 43 Nm-1 with detrimental effects. These results also suggest the existence of a separate discharge stage between 0.6 and 0.1 V, where the generated stress is only approximately one-third of that observed below 0.1 V. This approach can be adapted to help resolve the localized stress in a wide range of electrode materials, to gain additional insights into mechanical effects of charge storage, and for long-lifetime battery design.
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Mechanical signaling involved in molecular interactions lies at the heart of materials science and biological systems, but the mechanisms involved are poorly understood. Here we use nanomechanical sensors and intact human cells to provide unique insights into the signaling pathways of connectivity networks, which deliver the ability to probe cells to produce biologically relevant, quantifiable and reproducible signals. We quantify the mechanical signals from malignant cancer cells, with 10 cells per ml in 1000-fold excess of non-neoplastic human epithelial cells. Moreover, we demonstrate that a direct link between cells and molecules creates a continuous connectivity which acts like a percolating network to propagate mechanical forces over both short and long length-scales. The findings provide mechanistic insights into how cancer cells interact with one another and with their microenvironments, enabling them to invade the surrounding tissues. Further, with this system it is possible to understand how cancer clusters are able to co-ordinate their migration through narrow blood capillaries.
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The growing need to prevent pathogen outbreaks is irrefutable in the case of the food industry. Early detection in products, especially beverages, contaminated with bacterial strains is vital to avoid infected foods from reaching the consumer. If E. coli is pesent in such foods, it can cause infections. It can also be an indicator of the existence of other harmful coliforms. In this study, we have investigated the detection of Escherichia coli ( E. coli) in orange juice using a portable nanofiber-light addressable potentiometric sensor (NF-LAPS). We have chosen electrospun pH-sensitive poly(vinyl alcohol)/poly(acrylic acid) (PVA/PAA) hydrogel NFs as the sensitive layer. The successful detection of E. coli was reported with the NF-LAPS in less than 1 h. The limit of detection (LOD) measured in the sensor is found to be102 CFU/mL. We have confirmed the selectivity of the biosensor toward E. coli by examining the response of the NF-LAPS against Salmonella typhimurium ( S. typhi), also commonly found in orange juice. Despite the complex nature of orange juice, the response of the biosensor is in no way affected while orange juice is tested as is.
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Escherichia coli/aislamiento & purificación , Jugos de Frutas y Vegetales/microbiología , Nanofibras/química , PotenciometríaRESUMEN
The direct conversion of mechanical energy into electricity by nanomaterial-based devices offers potential for green energy harvesting 1-3 . A conventional triboelectric nanogenerator converts frictional energy into electricity by producing alternating current (a.c.) triboelectricity. However, this approach is limited by low current density and the need for rectification 2 . Here, we show that continuous direct-current (d.c.) with a maximum density of 106 A m-2 can be directly generated by a sliding Schottky nanocontact without the application of an external voltage. We demonstrate this by sliding a conductive-atomic force microscope tip on a thin film of molybdenum disulfide (MoS2). Finite element simulation reveals that the anomalously high current density can be attributed to the non-equilibrium carrier transport phenomenon enhanced by the strong local electrical field (105-106 V m-2) at the conductive nanoscale tip 4 . We hypothesize that the charge transport may be induced by electronic excitation under friction, and the nanoscale current-voltage spectra analysis indicates that the rectifying Schottky barrier at the tip-sample interface plays a critical role in efficient d.c. energy harvesting. This concept is scalable when combined with microfabricated or contact surface modified electrodes, which makes it promising for efficient d.c. triboelectricity generation.
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Spontaneous self-assemblies of biomolecules can generate geometrical patterns. Our findings provide an insight into the mechanism of self-assembled ring pattern generation by human serum albumin (HSA). The self-assembly is a process guided by kinetic and thermodynamic parameters. The generated protein ring patterns display a behavior which is geometrically related to a n-simplex model and is explained through thermodynamics and chemical kinetics.
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Albúmina Sérica Humana/química , Algoritmos , Humanos , Cinética , Microscopía de Fuerza Atómica , Albúmina Sérica Humana/ultraestructura , TermodinámicaRESUMEN
Apolipoprotein B (ApoB) is a major protein component of plasma lipoproteins. It is involved in many important biological processes such as lipid transportation, enzyme activity regulation, and receptor recognition. Extensive studies have shown that the expression of ApoB is regulated at multiple levels. However, the regulation of ApoB expression by microRNAs (miRNAs) still remains unknown. In the present study, identified are miRNAs that are predicted to interact with ApoB in chicken. The predicted relationship between the identified miRNAs and ApoB was verified through dual luciferase reporter assay in chicken DF1 cells, and the effect of miRNAs on ApoB expression was analyzed in chicken embryo hepatocytes stimulated by 17ß-estradiol. The results show that miR-101-2-5p was predicted to interact with ApoB. Dual luciferase reporter assay together with the miR-101-2-5p mimics study demostrate that ApoB is the target of miR-101-2-5p, which suppresses the expression of ApoB through binding with the 3'UTR of ApoB. Our experiments suggest that miR-101-2-5p might be involved in lipid metabolism through binding to the 3'UTR of ApoB in the liver of egg-laying chickens.
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Apolipoproteínas B/genética , Pollos/genética , Regulación de la Expresión Génica , Hígado/metabolismo , MicroARNs/metabolismo , Animales , Células Cultivadas , FemeninoRESUMEN
Poultry meat quality is associated with breed, age, tissue and other factors. Many previous studies have focused on distinct breeds; however, little is known regarding the epigenetic regulatory mechanisms in different age stages, such as DNA methylation. Here, we compared the global DNA methylation profiles between juvenile (20 weeks old) and later laying-period (55 weeks old) hens and identified candidate genes related to the development and meat quality of breast muscle using whole-genome bisulfite sequencing. The results showed that the later laying-period hens, which had a higher intramuscular fat (IMF) deposition capacity and water holding capacity (WHC) and less tenderness, exhibited higher global DNA methylation levels than the juvenile hens. A total of 2,714 differentially methylated regions were identified in the present study, which corresponded to 378 differentially methylated genes, mainly affecting muscle development, lipid metabolism, and the ageing process. Hypermethylation of the promoters of the genes ABCA1, COL6A1 and GSTT1L and the resulting transcriptional down-regulation in the later laying-period hens may be the reason for the significant difference in the meat quality between the juvenile and later laying-period hens. These findings contribute to a better understanding of epigenetic regulation in the skeletal muscle development and meat quality of chicken.
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Pollos , Metilación de ADN , Epigénesis Genética , Calidad de los Alimentos , Carne , Envejecimiento , Animales , Metabolismo de los Lípidos , Músculo Esquelético/crecimiento & desarrollo , Secuenciación Completa del Genoma/métodosRESUMEN
A conjugated polymer interface consisting of an oxaborole containing polymer and a glycopolymer was used for achieving very high selectivity in dopamine (DA) detection. The optimum binding affinity between the polymers promotes the selectivity to DA through a displacement mechanism while remaining unaffected by other structurally related analogs and saccharide derivatives. Real-time detection of DA with very high selectivity and sensitivity has been demonstrated by immobilizing the polymer conjugates on surface plasmon resonance (SPR) and microcantilever (MCL) sensor platforms. Using the conjugated polymer sensing layer, the SPR biosensor was capable of detecting DA in the concentration range of 1 × 10-9 to 1 × 10-4 mol L-1, whereas the MCL sensor showed a limit of detection (LOD) of 5 × 10-11 mol L-1. We find that the sensing mechanism is based on DA-induced reversible swelling of the conjugated polymer layer and this allows regeneration and reuse of the sensor multiple times. Also, we conclude that SPR is a suitable sensor platform for DA in-line detection at clinical level considering the detection time and stability, whereas MCL can achieve a much lower LOD.
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Dopamina/química , Técnicas Biosensibles , Límite de Detección , Polímeros , Resonancia por Plasmón de SuperficieRESUMEN
The elongation of very long chain fatty acids protein 6 (ELOVL6) encodes a fatty acid elongase that is responsible for the final step in endogenous saturated fatty acid synthesis and involves in de novo lipogenesis. Though the regulatory mechanism of ELOVL6 expression has been studied extensively, little is known about the role of miRNA in regulating ELOVL6 gene expression in chicken until now. To investigate the regulatory mechanism of miRNA on the expression of ELOVL6 gene, bioinformatics analysis was employed to predict the potential miRNAs that binding with the 3'untranslated region (3'UTR) of ELOVL6. The putative miRNA was further screened by comparative analysis with previous miRNA-seq results. Gga-miR-22-3p, which could bind with the 3'UTR of ELOVL6 and showed negative expression correlation with ELOVL6 gene in chicken liver, was obtained. Tissue expression profiles showed that gga-miR-22-3p and ELOVL6 are extensively expressed in many tissues, and ELOVL6 with high expression level in kidney and liver tissues, and gga-miR-22-3p with high expression in lung and heart. Dual-luciferase reporter assays results indicated that the expression of luciferase reporter gene linked with part sequence of the 3'UTR of chicken ELOVL6 gene was down-regulated by the overexpression of gga-miR-22-3p in the DF1 cells, and the down-regulation behavior was abolished when the gga-miR-22-3p binding site in 3'UTR of ELOVL6 was mutated (P>0.05). Furthermore, the ELOVL6 expression in chicken hepatocytes was down-regulated when miR-22-3p was over-expressed. Therefore, we concluded that miR-22-3p might involve in controlling the hepatic lipid composition through affecting the expression of ELOVL6 gene, and could serve as a regulator of lipid metabolism in the liver of egg-laying hen.
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Acetiltransferasas/genética , Pollos/genética , Hígado/metabolismo , MicroARNs/genética , Regiones no Traducidas 3' , Acetiltransferasas/metabolismo , Animales , Pollos/fisiología , Elongasas de Ácidos Grasos , Femenino , Riñón/metabolismo , Lipólisis/genética , Pulmón/metabolismo , Miocardio/metabolismo , OviparidadRESUMEN
Ligand-directed targeting and capturing of cancer cells is a new approach for detecting circulating tumor cells (CTCs). Ligands such as antibodies have been successfully used for capturing cancer cells and an antibody based system (CellSearch(®)) is currently used clinically to enumerate CTCs. Here we report the use of a peptide moiety in conjunction with a microcantilever array system to selectively detect CTCs resulting from cancer, specifically breast cancer. A sensing microcantilever, functionalized with a breast cancer specific peptide 18-4 (WxEAAYQrFL), showed significant deflection on cancer cell (MCF7 and MDA-MB-231) binding compared to when exposed to noncancerous (MCF10A and HUVEC) cells. The peptide-functionalized microcantilever allowed efficient capture and detection of cancer cells in MCF7 spiked human blood samples emulating CTCs in human blood. A detection limit of 50-100 cancer cells mL(-1) from blood samples was achieved with a capture yield of 80% from spiked whole blood samples. The results emphasize the potential of peptide 18-4 as a novel peptide for capturing and detecting cancer cells in conjunction with nanomechanical cantilever platform. The reported peptide-based cantilever platform represents a new analytical approach that can lead to an alternative to the various detection platforms and can be leveraged to further study CTCs.
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Neoplasias de la Mama/patología , Oligopéptidos/química , Secuencia de Aminoácidos , Recuento de Células/instrumentación , Recuento de Células/métodos , Separación Celular , Femenino , Células Endoteliales de la Vena Umbilical Humana/fisiología , Humanos , Límite de Detección , Células MCF-7 , Células Neoplásicas Circulantes/patología , Resonancia por Plasmón de SuperficieRESUMEN
We employed a direct peptide-bacteria binding assay to screen peptide fragments for high and specific binding to Listeria monocytogenes. Peptides were screened from a peptide array library synthesized on cellulose membrane. Twenty four peptide fragments (each a 14-mer) were derived from three potent anti-listerial peptides, Leucocin A, Pediocin PA1, and Curvacin A, that belong to class IIa bacteriocins. Fragment Leu10 (GEAFSAGVHRLANG), derived from the C-terminal region of Leucocin A, displayed the highest binding among all of the library fragments toward several pathogenic Gram-positive bacteria, including L. monocytogenes, Enterococcus faecalis, and Staphylococcus aureus. The specific binding of Leu10 to L. monocytogenes was further validated using microcantilever (MCL) experiments. Microcantilevers coated with gold were functionalized with peptides by chemical conjugation using a cysteamine linker to yield a peptide density of â¼4.8×10(-3) µmol/cm2 for different peptide fragments. Leu10 (14-mer) functionalized MCL was able to detect Listeria with same sensitivity as that of Leucocin A (37-mer) functionalized MCL, validating the use of short peptide fragments in bacterial detection platforms. Fragment Leu10 folded into a helical conformation in solution, like that of native Leucocin A, suggesting that both Leu10 and Leucocin A may employ a similar mechanism for binding target bacteria. The results show that peptide-conjugated microcantilevers can function as highly sensitive platforms for Listeria detection and hold potential to be developed as biosensors for pathogenic bacteria.