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Lactate plays a crucial role in energy metabolism and greatly impacts protein activities, exerting diverse physiological and pathological effects. Therefore, convenient lactate assays for tracking spatiotemporal dynamics in living cells are desirable. In this paper, we engineered and optimized a red fluorescent protein sensor for l-lactate named FiLa-Red. This indicator exhibited a maximal fluorescence change of 730 % and an apparent dissociation constant (Kd) of approximately 460 µM. By utilizing FiLa-Red and other sensors, we monitored energy metabolism in a multiplex manner by simultaneously tracking lactate and NAD+/NADH abundance in the cytoplasm, nucleus, and mitochondria. The FiLa-Red sensor is expected to be a useful tool for performing metabolic analysis in vitro, in living cells and in vivo.
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Due to intrinsic defect of fire hazard security, improving the flame retardant capacity of paper is still insufficient in case of precombustion. Herein, we integrate the flame retardant with flame detection performance on the surface of paper, the restricts of graphene oxide (GO) are overcome between high thermoelectric performance and flexibility. A flexible lamellar thermoelectric composite (GO-LA-HP) is constructed through the co-assembly of GO, ascorbic acid (LA), and phenoxycycloposphazene (HP), lamellar GO-LA-HP avoid GO's toughness decline after modification, but also enhance fire sensitivity and flame resistance. The composite could significantly decrease the temperature rise stage (<150 °C), and trigger the fire-warning within 2 s. The hybrid coating composed of phytic acid/phosphoric acid (PyA/PA) and modified thermoelectric GO-LA-HP becomes excellent compatibility on the surface of the modified paper (GPP). GPP is able to extinguish itself immediately after leaving the fire in the vertical combustion, and display the excellent flame resistance. Furthermore, GPP's alarm signal could be timely generated as little as 3 s of contacting the flame, and the response time can exceed 600 s. According to the structure observation and analysis, the related synergistic fire detection reinforcing and flame-retardant mechanisms are also proposed and clarified.
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The translation elongation factor 1α (EF1α) protein is a highly conserved G protein that is crucial for protein translation in all eukaryotic organisms. EF1α quickly became insoluble at temperatures 42 °C treatment for 2h in vitro, but generally remained soluble in vivo even after being exposed to temperatures as high as 45 °C for an extended period, which suggests that protective mechanisms exist for keeping EF1α soluble in plant cells under heat stress. EF1α had fast in vivo insolubilization when exposed to 45 °C, resulting in about 40% of the protein aggregating after 9 h. Given its established role in protein translation, heat-induced aggregation is most likely to impact the function of the elongation factor. Overexpression of constitutive mutants in both GTP-bound and GDP-bound forms of EF1α resulted in significantly decreased heat tolerance. These findings provide evidence to support the critical role of EF1α, a thermosensitive protein, in the heat tolerance of plants.
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Factor 1 de Elongación Peptídica , Termotolerancia , Factor 1 de Elongación Peptídica/metabolismo , Factor 1 de Elongación Peptídica/genética , Termotolerancia/genética , Arabidopsis/genética , Arabidopsis/metabolismo , Calor , Agregado de Proteínas , Proteínas de Arabidopsis/metabolismo , Proteínas de Arabidopsis/genética , Respuesta al Choque Térmico/fisiologíaRESUMEN
CuInTe2 (CIT) is one of the typical ternary chalcogenides known for its characteristic mixed polyanionic/polycationic site defects, making it a subject of continuous interest in the field of thermoelectrics. In this work, we propose a chemical composition modulation strategy for CIT by alloying GeTe and then introducing a copper deficiency (denoted by VCu). This strategy aims to unpin its Fermi level (Fr) and shift Fr into the valence band (VB) while simultaneously enabling coupling between the optical and acoustic phonon, thereby providing an extra phonon scattering path at low frequencies. The simultaneous composition regulations not only enhance the carrier concentration (nH) to 1019-1020 cm-3 but also significantly reduce the lattice thermal conductivity (κL) to â¼0.48 W m-1 K-1, thus effectively realizing electro-acoustic coordination in the present material. As a consequence, the thermoelectric (TE) performance is remarkably improved with the highest TE figure of merit (ZT) of 1.51 at â¼838 K. This value ranks at a higher level among CIT-based materials, which showcases the great significance of chemical composition modulation.
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Edad de Inicio , Vitíligo , Humanos , Vitíligo/epidemiología , Vitíligo/diagnóstico , Estudios Retrospectivos , Femenino , Niño , Masculino , Adolescente , PreescolarRESUMEN
BACKGROUND: Patients with spondyloarthritis (SpA)/HLA-B27-associated acute anterior uveitis (AAU) experience recurring acute flares, which pose significant visual and financial challenges. Despite established links between SpA and HLA-B27-associated AAU, the exact mechanism involved remains unclear, and further understanding is needed for effective prevention and treatment. METHODS: To investigate the acute pathogenesis of SpA/HLA-B27-associated AAU, Mendelian randomization (MR) and single-cell transcriptomic analyses were employed. The MR incorporated publicly available protein quantitative trait locus data from previous studies, along with genome-wide association study data from public databases. Causal relationships between plasma proteins and anterior uveitis were assessed using two-sample MR. Additionally, colocalization analysis was performed using Bayesian colocalization. Single-cell transcriptome analysis utilized the anterior uveitis dataset from the Gene Expression Omnibus (GEO) database. Dimensionality reduction, clustering, transcription factor analysis, pseudotime analysis, and cell communication analysis were subsequently conducted to explore the underlying mechanisms involved. RESULTS: Mendelian randomization analysis revealed that circulating levels of AIF1 and VARS were significantly associated with a reduced risk of developing SpA/HLA-B27-associated AAU, with AIF1 showing a robust correlation with anterior uveitis onset. Colocalization analysis supported these findings. Single-cell transcriptome analysis showed predominant AIF1 expression in myeloid cells, which was notably lower in the HLA-B27-positive group. Pseudotime analysis revealed dendritic cell terminal positions in differentiation branches, accompanied by gradual decreases in AIF1 expression. Based on cell communication analysis, CD141+CLEC9A+ classic dendritic cells (cDCs) and the APP pathway play crucial roles in cellular communication in the Spa/HLA-B27 group. CONCLUSIONS: AIF1 is essential for the pathogenesis of SpA/HLA-B27-associated AAU. Myeloid cell differentiation into DCs and decreased AIF1 levels are also pivotal in this process.
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Espondiloartritis , Uveítis Anterior , Humanos , Enfermedad Aguda , Teorema de Bayes , Perfilación de la Expresión Génica , Estudio de Asociación del Genoma Completo , Antígeno HLA-B27/genética , Antígeno HLA-B27/metabolismo , Espondiloartritis/genética , Espondiloartritis/metabolismo , Uveítis Anterior/genética , Uveítis Anterior/metabolismoRESUMEN
As a key glycolytic metabolite, lactate has a central role in diverse physiological and pathological processes. However, comprehensive multiscale analysis of lactate metabolic dynamics in vitro and in vivo has remained an unsolved problem until now owing to the lack of a high-performance tool. We recently developed a series of genetically encoded fluorescent sensors for lactate, named FiLa, which illuminate lactate metabolism in cells, subcellular organelles, animals, and human serum and urine. In this protocol, we first describe the FiLa sensor-based strategies for real-time subcellular bioenergetic flux analysis by profiling the lactate metabolic response to different nutritional and pharmacological conditions, which provides a systematic-level view of cellular metabolic function at the subcellular scale for the first time. We also report detailed procedures for imaging lactate dynamics in live mice through a cell microcapsule system or recombinant adeno-associated virus and for the rapid and simple assay of lactate in human body fluids. This comprehensive multiscale metabolic analysis strategy may also be applied to other metabolite biosensors using various analytic platforms, further expanding its usability. The protocol is suited for users with expertise in biochemistry, molecular biology and cell biology. Typically, the preparation of FiLa-expressing cells or mice takes 2 days to 4 weeks, and live-cell and in vivo imaging can be performed within 1-2 hours. For the FiLa-based assay of body fluids, the whole measuring procedure generally takes ~1 min for one sample in a manual assay or ~3 min for 96 samples in an automatic microplate assay.
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Técnicas Biosensibles , Ácido Láctico , Animales , Humanos , Ratones , Técnicas Biosensibles/métodos , Ácido Láctico/metabolismo , Ácido Láctico/análisisRESUMEN
Flameproof modification of paper can improve safety and application performance. However, traditional paper is prone to moisture absorption, resulting in significant reduction in flame retardant performance, even complete failure, greatly limiting the application environment. In order to achieve long-term flame retardant properties of paper, while avoiding the loss of physical properties caused by the introduction of flame retardants, in this work, a plant acid/phosphate and melamine formaldehyde coating (PyA/PA-MF) is prepared through electrostatic self-assembly for durable flame retardant performance of cellulose paper. Due to the electrostatic interaction, the paper surface become greatly rough with introduction of PyA/PA-MF, a uniform microsphere structure is formed on the surface of the paper cellulose, which effectively fix the phosphorus-containing groups. The oxygen index reaches 33 % and the carbon length was only 6.3 ± 0.2 cm, the pHRR and THR are decreased by 80 % and 73 %, respectively. After being immersed for 72 h, the oxygen index is still 31.4 % and carbon length is no more than 12 cm. mechanical property of modified paper is significant increased in the tensile strength (2.4 MPa) compared to the blank paper (1 MPa), as well as that the whiteness of the surface of the modified paper will not change. In summary, PyA/PA-MF endows paper long-term flame retardant performance while maintaining its basic performance.
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Retardadores de Llama , Electricidad Estática , Carbono , Celulosa , OxígenoRESUMEN
BACKGROUND: Canopy architecture is critical in determining the fruit-zone microclimate and, ultimately, in determining an orchard's success in terms of the quality and quantity of the fruit produced. However, few studies have addressed how the canopy environment leads to metabolomic and transcriptomic alterations in fruits. Designing strategies for improving the quality of pear nutritional components relies on uncovering the related regulatory mechanisms. RESULTS: We performed an in-depth investigation of the impact of canopy architecture from physiological, metabolomic and transcriptomic perspectives by comparing pear fruits grown in a traditional freestanding system (SP) or a flat-type trellis system (DP). Physiological studies revealed relatively greater fruit sizes, soluble solid contents and titratable acidities in pear fruits from DP systems with open canopies. Nontargeted metabolite profiling was used to characterize fruits at the initial ripening stage. Significant differences in fruit metabolites, including carbohydrates, nucleic acids, alkaloids, glycerophospholipids, sterol lipids, and prenol lipids, were observed between the two groups. Transcriptomic analysis indicated that a series of organic substance catabolic processes (e.g., the glycerol-3-phosphate catabolic process, pectin catabolic process and glucan catabolic process) were overrepresented in fruits of the DP system. Moreover, integrative analysis of the metabolome and transcriptome at the pathway level showed that DP pear fruits may respond to the canopy microenvironment by upregulating phenylpropanoid biosynthesis pathway genes such as PpPOD. Transient assays revealed that the contents of malic acid and citric acid were lower in the pear flesh of PpPOD RNAi plants, which was associated with regulating the expression of organic acid metabolism-related genes. CONCLUSIONS: Our results provide fundamental evidence that at the physiological and molecular levels, open-canopy architecture contributes to improving pear fruit quality and is correlated with increased levels of carbohydrates and lipid-like molecules. This study may lead to the development of rational culture practices for enhancing the nutritional traits of pear fruits.
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Pyrus , Frutas , Proteínas de Plantas/genética , Perfilación de la Expresión Génica , Carbohidratos , Lípidos , Regulación de la Expresión Génica de las PlantasRESUMEN
AIMS: This work sought to investigate the mechanism underlying the STING signaling pathway during myocardial infarction (MI), and explore the involvement and the role of SIRT6 in the process. MAIN METHODS: Mice underwent the surgery of permanent left anterior descending (LAD) artery constriction. Primary cardiomyocytes (CMs) and fibroblasts were subjected to hypoxia to mimic MI in vitro. STING expression was assessed in the infarct heart, and the effect of STING inhibition on cardiac fibrosis was explored. This study also evaluated the regulatory effect of STING by SIRT6 in macrophages. KEY FINDINGS: STING protein was increased in the infarct heart tissue, highlighting its involvement in the post-MI inflammatory response. Hypoxia-induced death of CMs and fibroblasts contributed to the upregulation of STING in macrophages, establishing the involvement of STING in the intercellular signaling during MI. Inhibition of STING resulted in a significant reduction of cardiac fibrosis at day 14 after MI. Additionally, this study identified SIRT6 as a key regulator of STING via influencing its acetylation and ubiquitination in macrophages, providing novel insights into the posttranscriptional modification and expression of STING at the acute phase after myocardial infarction. SIGNIFICANCE: This work shows the key role of SIRT6/STING signaling in the pathogenesis of cardiac injury after MI, suggesting that targeting this regulatory pathway could be a promising strategy to attenuate cardiac fibrosis after MI.
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Lesiones Cardíacas , Infarto del Miocardio , Sirtuinas , Animales , Ratones , Modelos Animales de Enfermedad , Fibrosis , Lesiones Cardíacas/metabolismo , Hipoxia/metabolismo , Macrófagos/metabolismo , Ratones Endogámicos C57BL , Infarto del Miocardio/metabolismo , Miocardio/metabolismo , Miocitos Cardíacos/metabolismo , Transducción de Señal , Sirtuinas/metabolismoRESUMEN
Traditional methods of coherent diffraction imaging using random masks result in an insufficient difference between the diffraction patterns, making it challenging to form a strong amplitude constraint, causing significant speckle noise in the measurement results. Hence, this study proposes an optimized mask design method combining random and Fresnel masks. Increasing the difference between diffraction intensity patterns enhances the amplitude constraint, suppresses the speckle noise effectively, and improves the phase recovery accuracy. The numerical distribution of the modulation masks is optimized by adjusting the combination ratio of the two mask modes. The simulation and physical experiments show that the reconstruction results of PSNR and SSIM using the proposed method are higher than those using random masks, and the speckle noises are effectively reduced.
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Autophagy and glycolysis are highly conserved biological processes involved in both physiological and pathological cellular programs, but the interplay between these processes is poorly understood. Here, we show that the glycolytic enzyme lactate dehydrogenase A (LDHA) is activated upon UNC-51-like kinase 1 (ULK1) activation under nutrient deprivation. Specifically, ULK1 directly interacts with LDHA, phosphorylates serine-196 when nutrients are scarce and promotes lactate production. Lactate connects autophagy and glycolysis through Vps34 lactylation (at lysine-356 and lysine-781), which is mediated by the acyltransferase KAT5/TIP60. Vps34 lactylation enhances the association of Vps34 with Beclin1, Atg14L, and UVRAG, and then increases Vps34 lipid kinase activity. Vps34 lactylation promotes autophagic flux and endolysosomal trafficking. Vps34 lactylation in skeletal muscle during intense exercise maintains muscle cell homeostasis and correlates with cancer progress by inducing cell autophagy. Together, our findings describe autophagy regulation mechanism and then integrate cell autophagy and glycolysis.
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Fosfatidilinositol 3-Quinasas Clase III , Lisina , Homólogo de la Proteína 1 Relacionada con la Autofagia/genética , Homólogo de la Proteína 1 Relacionada con la Autofagia/metabolismo , Proteínas Relacionadas con la Autofagia/metabolismo , Fosfatidilinositol 3-Quinasas Clase III/genética , Fosfatidilinositol 3-Quinasas Clase III/metabolismo , LípidosRESUMEN
The present study aims to investigate the role of AKT2 in the pathogenesis of hepatic and cardiac lipotoxicity induced by lipid overload-induced obesity and identify its downstream targets. WT and Akt2 KO mice were fed either normal diet, or high-fat diet (HFD) to induce obesity model in vivo. Human hepatic cell line (L02 cells) and neonatal rat cardiomyocytes (NRCMs) were used as in vitro models. We observed that during HFD-induced obesity, Akt2 loss-of-function mitigated lipid accumulation and oxidative stress in the liver and heart tissue. Mechanistically, down-regulation of Akt2 promotes SIRT6 expression in L02 cells and NRCMs, the latter deacetylates SOD2, which promotes SOD2 activity and therefore alleviates oxidative stress-induced injury of hepatocytes and cardiomyocytes. Furthermore, we also proved that AKT2 inhibitor protects hepatocytes and cardiomyocytes from HFD-induced oxidative stress. Therefore, our work prove that AKT2 plays an important role in the regulation of obesity-induced lipid metabolic disorder in the liver and heart. Our study also indicates AKT2 inhibitor as a potential therapy for obesity-induced hepatic and cardiac injury.
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Dieta Alta en Grasa , Sirtuinas , Humanos , Animales , Ratones , Ratas , Dieta Alta en Grasa/efectos adversos , Hígado/metabolismo , Estrés Oxidativo , Obesidad/metabolismo , Miocitos Cardíacos/metabolismo , Sirtuinas/metabolismo , Lípidos , Ratones Endogámicos C57BL , Ratones Noqueados , Proteínas Proto-Oncogénicas c-akt/metabolismoRESUMEN
Rewiring of redox metabolism has a profound impact on tumor development, but how the cellular heterogeneity of redox balance affects leukemogenesis remains unknown. To precisely characterize the dynamic change in redox metabolism in vivo, we developed a bright genetically encoded biosensor for H2O2 (named HyPerion) and tracked the redox state of leukemic cells in situ in a transgenic sensor mouse. A H2O2-low (HyPerion-low) subset of acute myeloid leukemia (AML) cells was enriched with leukemia-initiating cells, which were endowed with high colony-forming ability, potent drug resistance, endosteal rather than vascular localization, and short survival. Significantly high expression of malic enzymes, including ME1/3, accounted for nicotinamide adenine dinucleotide phosphate (NADPH) production and the subsequent low abundance of H2O2. Deletion of malic enzymes decreased the population size of leukemia-initiating cells and impaired their leukemogenic capacity and drug resistance. In summary, by establishing an in vivo redox monitoring tool at single-cell resolution, this work reveals a critical role of redox metabolism in leukemogenesis and a potential therapeutic target.
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Peróxido de Hidrógeno , Leucemia Mieloide Aguda , Ratones , Animales , Leucemia Mieloide Aguda/tratamiento farmacológico , Leucemia Mieloide Aguda/genética , Leucemia Mieloide Aguda/metabolismo , Oxidación-Reducción , Ratones Transgénicos , Resistencia a Antineoplásicos/genéticaRESUMEN
BACKGROUND: Anesthesiologists need to appreciate the impact of preoperative anxiety in children. The present study aimed to explore whether interactive multimedia-based home-initiated interventions could effectively relieve preoperative anxiety in pediatric patients. METHODS: In this prospective study, we compared preoperative anxiety between two groups of children aged 4-9 years. Children in the control group received a question-and-answer (Q&A) introduction, and children in the intervention group received multimedia-based home-initiated preoperative education using comic booklets, videos, and coloring game books. Differences in anxiety between the two groups were evaluated by the modified Yale Preoperative Anxiety Scale-Short Form (mYPAS-SF) at four time points: in the ophthalmology outpatient clinic before intervention as the baseline (T0); in the preoperative waiting area (T1); at the time of separating from their parents and moving to the operating room (T2); and at the time of anesthesia induction (T3). Parental anxiety was assessed by the Self-rating Anxiety Scale (SAS) and Visual Analog Scale (VAS) at T0 and T2. Other related information was collected by questionnaire. RESULTS: Eighty-four children who underwent pediatric strabismus in our center between November 2020 and July 2021 were included in this study. An intention-to-treat (ITT) analysis was performed on data from 78 enrolled children. Children in the intervention group exhibited lower m-YPAS-SF scores at T1, T2, and T3 than those in the control group (all p < 0.001). By using a mixed-effect model with repeated measurement (MMRM) after adjusting the m-YPAS score at T0 as a covariate, the interventional effect in terms of themYPAS-SF score was also significant over time (p < 0.001). The percentage of children with perfect induction compliance (ICC = 0) in the intervention group was significantly higher than that in the control group [18.4% vs. 7.5%], and poor induction compliance (ICC>4) was lower (2.6% vs. 17.5%, p = 0.048). The mean parental VAS score at T2 in the intervention group was significantly lower than that in the control group (p = 0.021). CONCLUSIONS: Interactive multimedia-based home-initiated intervention could reduce preoperative anxiety in children and improve the quality of anesthesia induction based on ICC scores, which may in turn impose a positive impact on parental anxiety.
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Ansiedad , Padres , Cuidados Preoperatorios , Humanos , Niño , Ansiedad/prevención & control , Cuidados Preoperatorios/métodos , Anestesiólogos , Estudios Prospectivos , Estrabismo/cirugía , Padres/educación , Educación del Paciente como AsuntoRESUMEN
OBJECTIVES: This study aims to investigate the usefulness of diffusion-weighted imaging (DWI) and arterial spin labeling (ASL) for predicting final infarct volume in patients with acute atherothrombotic subtype cerebral infarction (AT-type stroke). METHODS: The data of 77 patients with AT-type stroke were retrospectively analyzed. ASL and DWI values of minimum apparent diffusion coefficient (min ADC), mean ADC (mean ADC), minimum cerebral blood flow (min CBF), and mean CBF (mean CBF) of the infarction lesions were measured. Changes in cerebral infarction volume (ΔVolume) were determined by DWI reexamination on the 7th day after onset. Correlations of ADC and CBF with Δ Volume were analyzed. Receiver operating characteristic (ROC) curve analysis was used to determine the usefulness of ADC and CBF values for predicting final infarct volume. RESULTS: There was a significant difference in the distribution of the ΔVolume in AT-type stroke (P<0.0001). The ADC and min CBF values were negatively correlated with the infarct ΔVolume (P<0.05); mean CBF and ΔCBF were not correlated with ΔVolume. When min ADC was ≤0.303 × 10-3 mm2/s, min CBF 1.5 ≤2.415 mL/100 g/min, or min CBF2.5 ≤4.25 mL/100 g/min, ΔVolume was likely to be large. The ROC curve showed the highest predictive value for min ADC and min CBF. CONCLUSION: Distinctive patterns of quantitative ADC and CBF can be used as a simple and rapid method for predicting change in infarction volume in AT-type stroke.
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Isquemia Encefálica , Accidente Cerebrovascular , Humanos , Estudios Retrospectivos , Marcadores de Spin , Accidente Cerebrovascular/diagnóstico por imagen , Isquemia Encefálica/diagnóstico , Enfermedad Aguda , Infarto Cerebral/diagnóstico por imagen , InfartoRESUMEN
BACKGROUND: Depression is a typical outcome of the repair of posttraumatic stress disorder (PTSD). Based on network pharmacology and neuropharmacology experiments, this study aimed to explore how gastrodin (GAS) reverses depressive symptoms in traumatically stressed rats. METHODS: GAS-related targets were predicted by SwissTargetPrediction; depression-related targets were collected from GeneCards and therapeutic target database (TTD); protein-protein interaction (PPI) network was constructed with its action mechanism being predicted by gene ontology (GO) analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment. The animal model of PTSD was replicated by single prolonged stress (SPS). The antidepressant effect of GAS was investigated by the forced swim test (FST) and tail suspension test (TST). The levels of tyrosine hydroxylase (TH) and corticotropin-releasing factor type I receptor (CRF1) in locus ceruleus (LC) and the expression of corticotropin-releasing factor (CRF) in the paraventricular nucleus of the hypothalamus (PVN) and central amygdala (CeA) were measured by immunofluorescence. RESULTS: GAS significantly shortened the tail suspension and swimming immobility in SPS rats in TST and FST experiments (p < 0.05 or p < 0.01). The network analysis showed that the critical antidepressant targets of GAS were 86 targets such as GAPDH, CASP3 MMP9, HRAS, DPP4, and TH, which were significantly enriched in the pathways such as pathways neuroactive ligandreceptor interaction. High doses of GAS could significantly reduce the level of TH and CRF in CEA in the brain of rats with depressive symptoms (p < 0.01) and, at the same time, lower the expression of CRF in PVN (p < 0.05). CONCLUSION: The effect of GAS on depressive symptoms in SPS rats may be closely related to its reduction of CRF expression in PVN and CeA and inhibition of neuron (NE) synthesis in LC.
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Hormona Liberadora de Corticotropina , Depresión , Ratas , Animales , Hormona Liberadora de Corticotropina/genética , Hormona Liberadora de Corticotropina/metabolismo , Depresión/tratamiento farmacológico , Depresión/metabolismo , Farmacología en Red , Receptores de Hormona Liberadora de Corticotropina/genética , Receptores de Hormona Liberadora de Corticotropina/metabolismo , Antidepresivos/farmacología , Antidepresivos/uso terapéuticoRESUMEN
Despite its central importance in cellular metabolism, many details remain to be determined regarding subcellular lactate metabolism and its regulation in physiology and disease, as there is sensitive spatiotemporal resolution of lactate distribution, and dynamics remains a technical challenge. Here, we develop and characterize an ultrasensitive, highly responsive, ratiometric lactate sensor, named FiLa, enabling the monitoring of subtle lactate fluctuations in living cells and animals. Utilizing FiLa, we demonstrate that lactate is highly enriched in mammalian mitochondria and compile an atlas of subcellular lactate metabolism that reveals lactate as a key hub sensing various metabolic activities. In addition, FiLa sensors also enable direct imaging of elevated lactate levels in diabetic mice and facilitate the establishment of a simple, rapid, and sensitive lactate assay for point-of-care clinical screening. Thus, FiLa sensors provide powerful, broadly applicable tools for defining the spatiotemporal landscape of lactate metabolism in health and disease.
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Diabetes Mellitus Experimental , Animales , Ratones , Diabetes Mellitus Experimental/metabolismo , Mitocondrias/metabolismo , Ácido Láctico/metabolismo , MamíferosRESUMEN
The sirtuin 6 (SIRT6) participates in regulating glucose and lipid homeostasis. However, the function of SIRT6 in the process of cardiac pathogenesis caused by obesity-associated lipotoxicity remains to be unveiled. This study was designed to elucidate the role of SIRT6 in the pathogenesis of cardiac injury due to nutrition overload-induced obesity and explore the downstream signaling pathways affecting oxidative stress in the heart. In this study, we used Sirt6 cardiac-specific knockout murine models treated with a high-fat diet (HFD) feeding to explore the function and mechanism of SIRT6 in the heart tissue during HFD-induced obesity. We also took advantage of neonatal cardiomyocytes to study the role and downstream molecules of SIRT6 during HFD-induced injury in vitro, in which intracellular oxidative stress and mitochondrial content were assessed. We observed that during HFD-induced obesity, Sirt6 loss-of-function aggravated cardiac injury including left ventricular hypertrophy and lipid accumulation. Our results evidenced that upon increased fatty acid uptake, SIRT6 positively regulated the expression of endonuclease G (ENDOG), which is a mitochondrial-resident molecule that plays an important role in mitochondrial biogenesis and redox homeostasis. Our results also showed that SIRT6 positively regulated superoxide dismutase 2 (SOD2) expression post-transcriptionally via ENDOG. Our study gives a new sight into SIRT6 beneficial role in mitochondrial biogenesis of cardiomyocytes. Our data also show that SIRT6 is required to reduce intracellular oxidative stress in the heart triggered by high-fat diet-induced obesity, involving the control of ENDOG/SOD2.
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Estrés Oxidativo , Sirtuinas , Ratones , Animales , Estrés Oxidativo/fisiología , Sirtuinas/metabolismo , Obesidad/etiología , Obesidad/metabolismo , LípidosRESUMEN
Metformin is accepted as a first-line drug for the therapy of Type 2 diabetes (T2D), while its mechanism is still controversial. In the present study, by taking advantage of mouse model of high-fat-diet (HFD)-induced obesity and primary mouse hepatocytes (PMHCs) as well as human hepatocyte L02 cell line, we aimed to investigate the involvement of SIRTs during the application of metformin for the therapy of T2D. Our data evidenced that during HFD-induced obesity, there was elevation of nucleus protein acetylation. Analysis of liver tissue showed that among all SIRT members, SIRT6 expression was significantly down-regulated during HFD feeding, which was sustained to regular level with metformin administration. Our result also showed that SIRT6 suppressed intracellular oxidative stress upon FAs stimulation in PMHCs and L02 cells. Mechanistically, SIRT6, but not SIRT1 promoted PGC-1α expression. We further prove that ENDOG is downstream of PGC-1α. In addition, we evidenced that ENDOG protects hepatocytes from lipid-induced oxidative stress, and down-regulation of Endog blunted the protective role of metformin in defending against FAs-induced oxidative stress. Our study established a novel mechanism of metformin in counteracting lipid-induced hepatic injury via activating SIRT6/PGC-1α/ENDOG signaling, thus providing novel targets of metformin in the therapy of T2D.