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MicroRNAs (miRNAs) play vital roles in biological activities, but their in vivo imaging is still challenging due to the low abundance and the lack of efficient fluorescent tools. RNA aptamers with high affinity and low background emerge for bioimaging yet suffering from low brightness. We introduce a rational design based on target-mediated entropy-driven toehold exchange (EDTE) to induce the release of RNA aptamer and subsequently light up corresponding fluorophore, which achieves selective imaging of miRNAs with good stability in both living cells and tumor-bearing mouse. Through tailoring recognition unit of the EDTE probes, highly sensitive imaging of different miRNAs including miRNA-125b and miRNA-21 is achieved, confirming its universal bioimaging applications. In comparison with the reported "one-to-one" model, the EDTE strategy shows a remarkable 4.6-time improvement in signal/noise ratio for intracellular imaging of the same miRNA. Particularly, it realizes sensitive imaging of miRNA in vivo, providing a promising tool in investigating functions and interactions of disease-associated miRNAs.
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Aptâmeros de Nucleotídeos , Entropia , Corantes Fluorescentes , MicroRNAs , MicroRNAs/análise , MicroRNAs/metabolismo , Aptâmeros de Nucleotídeos/química , Animais , Corantes Fluorescentes/química , Camundongos , Humanos , Imagem Óptica , Camundongos NusRESUMO
OBJECTIVE: This systematic review and meta-analysis study aimed to evaluate the effectiveness of probiotics supplementation on glycaemic control in patients with type 2 diabetes mellitus (T2DM) based on the data from the randomised clinical trials (RCTs). METHODS: PubMed, Web of Sciences, Embase, and Cochrane Library were searched from the inception to October 2022, and RCTs about probiotics and T2DM were collected. The standardised mean difference (SMD) with 95% confidence interval (CI) was used to estimate the effects of probiotics supplementation on glycaemic control related parameters, e.g. fasting blood glucose (FBG), insulin, haemoglobin A1c (HbA1c), and homeostasis model of assessment of insulin resistance (HOMA-IR). RESULTS: Thirty RCTs including 1,827 T2MD patients were identified. Compared with the placebo group, the probiotics supplementation group had a significant decrease in the parameters of glycaemic control, including FBG (SMD = - 0.331, 95% CI - 0.424 to - 0.238, Peffect < 0.001), insulin (SMD = - 0.185, 95% CI - 0.313 to - 0.056, Peffect = 0.005), HbA1c (SMD = - 0.421, 95% CI - 0.584 to - 0.258, Peffect < 0.001), and HOMA-IR (SMD = - 0.224, 95% CI - 0.342 to - 0.105, Peffect < 0.001). Further subgroup analyses showed that the effect was larger in the subgroups of Caucasians, high baseline body mass index (BMI ≥ 30.0 kg/m2), Bifidobacterium and food-type probiotics (Psubgroup < 0.050). CONCLUSION: This study supported that probiotics supplementation had favourable effects on glycaemic control in T2DM patients. It may be a promising adjuvant therapy for patients with T2DM.
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Diabetes Mellitus Tipo 2 , Resistência à Insulina , Probióticos , Adulto , Humanos , Hemoglobinas Glicadas , Glicemia , Controle Glicêmico , Diabetes Mellitus Tipo 2/tratamento farmacológico , Probióticos/uso terapêutico , Probióticos/farmacologia , Insulina/uso terapêutico , Ensaios Clínicos Controlados Aleatórios como AssuntoRESUMO
The safety of CRISPR (clustered regularly interspaced short palindromic repeats)-based genome editing in the context of human gene therapy is largely unknown. CCR5 is a reasonable but not absolutely protective target for a cure of human immunodeficiency virus type 1 (HIV-1) infection, because CCR5-null blood cells are largely resistant to HIV-1 entry. We transplanted CRISPR-edited CCR5-ablated hematopoietic stem and progenitor cells (HSPCs) into a patient with HIV-1 infection and acute lymphoblastic leukemia. The acute lymphoblastic leukemia was in complete remission with full donor chimerism, and donor cells carrying the ablated CCR5 persisted for more than 19 months without gene editing-related adverse events. The percentage of CD4+ cells with CCR5 ablation increased by a small degree during a period of antiretroviral-therapy interruption. Although we achieved successful transplantation and long-term engraftment of CRISPR-edited HSPCs, the percentage of CCR5 disruption in lymphocytes was only approximately 5%, which indicates the need for further research into this approach. (Funded by the Beijing Municipal Science and Technology Commission and others; ClinicalTrials.gov number, NCT03164135.).
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Sistemas CRISPR-Cas , Edição de Genes/métodos , Infecções por HIV/terapia , HIV-1 , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas , Receptores CCR5/genética , Adulto , Antirretrovirais/uso terapêutico , Contagem de Células Sanguíneas , Contagem de Linfócito CD4 , Infecções por HIV/complicações , Infecções por HIV/tratamento farmacológico , HIV-1/genética , Humanos , Masculino , Leucemia-Linfoma Linfoblástico de Células Precursoras/complicações , Carga ViralRESUMO
The single batch normalization (BN) method is commonly used in the instance segmentation algorithms. The batch size is concerned with some drawbacks. A too small sample batch size leads to a sharp drop in accuracy, but a too large batch may result in the memory overflow of graphic processing units (GPU). These problems make BN not feasible to some instance segmentation tasks with inappropriate batch sizes. The self-adaptive normalization (SN) method, with an adaptive weight loss layer, shows good performance in instance segmentation algorithms, such as the YOLACT. However, the parameter averaging mechanism in the SN method is prone to problems in the weight learning and assignment process. In response to such a problem, the paper proposes to replace the single BN with an adaptive weight loss layer in SN models, based on which a weight learning method is developed. The proposed method increases the input feature expression ability of the subsequent layers. By building a Pytorch deep learning framework, the proposed method is validated in the MS-COCO data set and Autonomous Driving Cityscapes data set. The experimental results prove that the proposed method is effective in processing samples independent from the batch size. The stable accuracy for all kinds of target segmentation is achieved, and the overall loss value is significantly reduced at the same time. The convergence speed of the network is also improved.
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Particle production in ultrarelativistic heavy ion collisions depends on the details of the nucleon density distributions in the colliding nuclei. We demonstrate that the charged hadron multiplicity distributions in isobaric collisions at ultrarelativistic energies provide a novel approach to determine the poorly known neutron density distributions and thus the neutron skin thickness in finite nuclei, which can in turn put stringent constraints on the nuclear symmetry energy.
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Under the approximate chiral symmetry restoration, quark interactions with topological gluon fields in quantum chromodynamics can induce a chirality imbalance and parity violation in local domains. An electric charge separation (CS) could be generated along the direction of a strong magnetic field (B), a phenomenon called the chiral magnetic effect (CME). CS measurements by azimuthal correlators are contaminated by major backgrounds from elliptic flow anisotropy (v_{2}). Isobaric _{44}^{96}Ru+_{44}^{96}Ru and _{40}^{96}Zr+_{40}^{96}Zr collisions have been proposed to identify the CME (expected to differ between the two systems) out of the backgrounds (to be almost the same). We show, by using the density functional theory calculations of the proton and neutron distributions, that these expectations may not hold as originally anticipated because the two systems may have sizable differences in eccentricity and v_{2}.
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Transplantation of hematopoietic stem cells (HSCs) with a naturally occurring CCR5 mutation confers a loss of detectable HIV-1 in the patient, making ablation of the CCR5 gene in HSCs an ideal therapy for an HIV-1 cure. Although CCR5 disruption has been attempted in CD4+ T cells and hematopoietic stem/progenitor cells (HSPCs), efficient gene editing with high specificity and long-term therapeutic potential remains a major challenge for clinical translation. Here, we established a CRISPR/Cas9 gene editing system in human CD34+ HSPCs and achieved efficient CCR5 ablation evaluated in long-term reconstituted NOD/Prkdcscid/IL-2Rγnull mice. The CCR5 disruption efficiency in our system remained robust in secondary transplanted repopulating hematopoietic cells. More importantly, an HIV-1 resistance effect was observed as indicated by significant reduction of virus titration and enrichment of human CD4+ T cells. Hence, we successfully established a CRISPR/Cas9 mediated CCR5 ablating system in long-term HSCs, which confers HIV-1 resistance in vivo. Our study provides evidence for translating CCR5 gene-edited HSC transplantation for an HIV cure to the clinic.
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Sistemas CRISPR-Cas , Marcação de Genes , Infecções por HIV/genética , Infecções por HIV/virologia , HIV-1/fisiologia , Células-Tronco Hematopoéticas/metabolismo , Receptores CCR5/genética , Animais , Antígenos CD34/metabolismo , Linhagem da Célula , Ensaio de Unidades Formadoras de Colônias , Modelos Animais de Doenças , Resistência à Doença , Mutação da Fase de Leitura , Edição de Genes , Transplante de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/citologia , Células-Tronco Hematopoéticas/virologia , Humanos , Imunofenotipagem , Camundongos , Camundongos Transgênicos , Fenótipo , RNA Guia de Cinetoplastídeos , Deleção de SequênciaRESUMO
OBJECTIVE: To analyze pathological characteristics of organs recovered during forensic autopsy submitted by legal medicine experts. METHODS: From Baoji city, 358 cases of forensic autopsy specimens from a series of routine exams were collected. And histopathological diagnoses were reviewed. RESULTS: Majority of the 358 cases were young men. The major causes of death were trauma, sudden death and poisoning. The cause of death was determined with histology in 250 cases. No typical histological changes were noted in 101 cases. The tissue autolysis and decomposition were present in 7 cases. The major pathological diagnosis was cardiovascular disease, followed by diseases in respiratory, nervous, and digestive systems. CONCLUSION: Forensic autopsy with its professional characteristics, is different from regular autopsy. When diagnosing cause of death by histopathological examination, pathologists should collaborate with legal medicine experts to know the details of the cases, circumstances surrounding the death, and specific forensic pathological characteristics.
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Autopsia , Causas de Morte , Medicina Legal , Patologia Legal , Ansiedade , Autólise , Doenças Cardiovasculares , China , Comportamento Cooperativo , Morte Súbita , Feminino , Humanos , Relações Interprofissionais , MasculinoRESUMO
Establishing ecological management zones based on the supply-demand relationship of ecosystem services (ESs) is essential for fostering sustainable development within social-ecological systems and improving human well-being. In this study, the spatial pattern between supply and demand in five ESs (grain production (GP), carbon sequestration (CS), soil conservation (SC), water conservation (WC), and habitat quality (HQ)) is analyzed using the ESs supply-demand ratio (ESDR) method, the spatial autocorrelation method, and the coupled coordination degree model. Zoning is performed according to the differences in their spatial combinations, and differential zoning management policies are proposed. The following results were obtained: (1) In terms of the ESDR, except for a slight increase in GP surplus from 2010 to 2020, there is a decline in the surplus of the other four ESs. (2) CS, WC, and HQ are dominated by cluster types LH and HL. GP and SC are dominated by cluster types HH and LL. The average value of the coupling coordination degree (CCD) of comprehensive ESs supply and demand show five types: moderate disharmony, slight disharmony, near disharmony, basic coordination, and slight coordination. (3) Based on the multiple spatial heterogeneity of ESs supply and demand, differentiated ecological management strategies are proposed at the grid scale. Overall, this study discover the spatial pattern of mismatch between the supply and demand of ecosystem services (ESs) in mountainous urban areas. This contribution enhances the discourse surrounding sustainable development theory and advances research on the coupling of social-ecological systems. Furthermore, it offers valuable insights for the formulation of sustainable ecological management policies tailored to mountainous urban settings.
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Purpose: The aim of this study is to evaluate the efficacy and safety of Snap Needles (SN) in the management of Postoperative Hemorrhoidal Pain (POHP). Patients and Methods: A systematic search was conducted in various databases, including EMBASE, Web of Science, PubMed, WanFang database, China National Knowledge Infrastructure (CNKI), China Biomedical Literature Database (CBM), and China Science and Technology Journal Database (VIP), spanning from their inception to August 2023, to identify relevant randomized controlled trials (RCTs) on SN for POHP. The primary outcome measure was the Visual Analog Scale (VAS), while secondary outcomes encompassed the Total Effective Rate (TER), Wound Healing Time (WHT), Pain Relief Time (PRT), Pain Disappearance Time (PDT), and Adverse Events (AEs). The Cochrane Risk of Bias Tool was employed to assess the quality of individual studies. A meta-analysis was conducted using RevMan 5.4.1 software. Results: The meta-analysis included 11 RCTs involving 1188 POHP patients, with an overall assessment of study quality ranging from very low to moderate. The findings revealed that the SN group exhibited significant improvements in treatment outcomes when compared to the control group (CG). These improvements were reflected in reduced VAS scores (mean difference [MD] = -1.10, 95% confidence interval [CI]: -1.31, -0.89, P < 0.05), shorter WHT (MD = -2.55, 95% CI: -3.02, -2.09, P < 0.05), quicker PRT (MD = -7.99, 95% CI: -8.48, -7.49, P < 0.05), fewer AEs (risk ratio [RR] = 0.38, 95% CI: 0.22, 0.67, P < 0.05), improved TER (RR = 1.18, 95% CI: 1.09, 1.27, P < 0.05), and faster PDT (MD = 19.24, 95% CI: 14.17, 24.31, P < 0.05). Conclusion: The use of SN appears to yield favorable outcomes in the treatment of POHP, and is potentially an alternative therapy to western drug therapy.
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Thromboangiitis obliterans (TAO) is a non-atherosclerotic segmental inflammatory occlusive disease with a high recurrence rate, high disability rate, difficulty to cure, and poor prognosis. It has been clinically proven that Mailuoshutong pill (MLSTP) is an effective traditional Chinese medicine for treating TAO. As MLSTP contains hundreds of chemical components, the quality control of which is a challenge in the development of reliable quality evaluation metrics. This study aimed to evaluate the quality uniformity of MLSTP by establishing a multi-strategy platform. In the present study, the key targets and signaling pathways of MLSTP treating TAO were predicted by network pharmacology. It was further shown by in vivo validation experiments that MLSTP exerted therapeutic effects on TAO by modulating the PI3K-AKT signaling pathway, VEGF signaling pathway, and HIF-1 signaling pathway. In addition, UPLC fingerprints of MLSTP were established and screened for potential Q-markers of MLSTP in combination with network pharmacology results. Six components, including chlorogenic acid, liquiritin, paeoniflorin, calycosin-7-glucoside, berberine, and formononetin, were selected as potential quality markers (Q-markers) in MLSTP. Finally, the quantitative analysis of multi-components by single marker (QAMS) method was established to quantitatively analyze the six potential Q-markers, and the results were consistent with those obtained by the external standard method (ESM). Taken together, the multi-strategy platform established in this study would be conducive to the Q-markers screening and quality control of MLSTP, improving the quality standard of MLSTP and providing favorable assurance for the clinical management of TAO.
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Medicamentos de Ervas Chinesas , Medicamentos de Ervas Chinesas/análise , Fosfatidilinositol 3-Quinases/metabolismo , Medicina Tradicional Chinesa , Transdução de Sinais , Controle de QualidadeRESUMO
DNA methyltransferase is significant in cellular activities and gene expression, and its aberrant expression is closely linked to various cancers during initiation and progression. Currently, there is a great demand for reliable and label-free techniques for DNA methyltransferase evaluation in tumor diagnosis and cancer therapy. Herein, a low-background fluorescent RNA aptamer-based sensing approach for label-free quantification of cytosine-guanine (CpG) dinucleotides methyltransferase (M.SssI) is reported. The fluorogenic light-up RNA aptamers-based strategy exhibits high selectivity via restriction endonuclease, padlock-based recognition, and RNA transcription. By combining rolling circle amplification (RCA), and RNA transcription with fluorescence response of RNA aptamers of Spinach-dye compound, the proposed platform exhibited efficiently ultrahigh sensitivity toward M.SssI. Eventually, the detection can be achieved in a linear range of 0.02-100 U mL-1 with a detection limit of 1.6 × 10-3 U mL-1. Owing to these superior features, the method is further applied in serum samples spiked M.SssI, which delivers a recovery ranging from 92.0 to 107.0% and a relative standard deviation <7.0%, providing a promising and practical tool for determining M.SssI in complex biological matrices.
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Aptâmeros de Nucleotídeos , Aptâmeros de Nucleotídeos/genética , Metilases de Modificação do DNA , Técnicas de Amplificação de Ácido Nucleico/métodos , DNA/metabolismo , RNARESUMO
(1) Background: Exercise is effective in promoting and maintaining bone mass. The aim of this study was to detect the exercise-induced metabolic changes in bone tissue of zebrafish. (2) Methods: Thirty-eight zebrafish (Danio rerio, six months old) were analyzed. The exercise group (n = 19) received 8 weeks of counter-current swimming training. The control group (n = 19) was not subjected to exercise. Mineralization was quantified, and alkaline phosphatase (Alp) and anti-tartrate acid phosphatase (Trap) activities were estimated (n = 12). The metabolomics (n = 12) and transcriptomics (n = 14) data of bone tissue were used for the integration analyses. (3) Results: The results showed that the exercise training improved the bone mineralization of zebrafish, e.g., the exercise group (5.74 × 104 ± 7.63 × 103) had a higher mean optical density than the control group (5.26 × 104 ± 8.56 × 103, p = 0.046) for the caudal vertebrae. The amount of mineralized matrix in scales of the exercised zebrafish was also higher (0.156 ± 0.012 vs. 0.102 ± 0.003, p = 0.005). Both histological staining and biochemical analysis revealed increased Alp activity (0.81 ± 0.26 vs. 0.76 ± 0.01, p = 0.002) and decreased Trap activity (1.34 ± 0.01 vs. 1.36 ± 0.01, p = 0.005) in the exercise group. A total of 103 different metabolites (DMs, VIP ≥ 1, fold change (FC) ≥ 1.20 or ≤0.83, p < 0.050) were identified. Alanine, aspartate and glutamate metabolism, ß-alanine metabolism, pyrimidine metabolism, and pantothenate and CoA biosynthesis were the significantly enriched metabolic pathways (p < 0.050). A total of 35 genes (q ≤ 0.050 (BH), |Log2FC| ≥ 0.5) were coenriched with the 103 DMs in the four identified pathways. Protein-protein interaction network analysis of the 35 genes showed that entpd3, entpd1, and cmpk2 were the core genes. (4) Conclusions: The results of this study suggest that alanine, aspartate and glutamate metabolism, ß-alanine metabolism, pyrimidine metabolism, and pantothenate and CoA biosynthesis contributed to exercise-induced improvements in bone mass.
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Transcriptoma , Peixe-Zebra , Animais , Ácido Aspártico , Metabolômica , Alanina , beta-Alanina , Pirimidinas , GlutamatosRESUMO
BACKGROUND: Mailuo Shutong Pills (MLST) have displayed pharmacological activity against thromboangiitis obliterans (TAO). However, the active ingredients and therapeutic mechanism of MLST against TAO remained to be further clarified. PURPOSE: The aim of this study was to explore the active components of MLST and their synergistic mechanism against TAO by integrating pharmacokinetics (PK) and pharmacometabolomics (PM). METHODS: TAO model rats were established by sodium laurate solution. Firstly, the efficacy of MLST was evaluated by gangrene score, blood flow velocity, and hematoxylin-eosin (H&E) staining. Secondly, PK research was conducted on bioavailable components to characterize their dynamic behaviors under TAO. Thirdly, multiple plasma and urine metabolic biomarkers for sodium laurate-induced TAO rats were found by untargeted metabolomics, and then variations in TAO-altered metabolites following MLST treatment were analyzed utilizing multivariate and bioinformatic analysis. Additionally, metabolic pathway analysis was performed using MetaboAnalyst. Finally, the dynamic link between absorbed MLST-compounds and TAO-associated endogenous metabolites was established by correlation analysis. RESULTS: MLST significantly alleviated gangrene symptoms by improving the infiltration of inflammatory cells and blood supply in TAO rats. Significant differences in metabolic profiles were found in 17 differential metabolites in plasma and 24 in urine between Sham and TAO rats. The 10 bioavailable MLST-compounds, such as chlorogenic acid and paeoniflorin, showed positive or negative correlations with various TAO-altered metabolites related to glutamate metabolism, histidine metabolism, arachidonic acid metabolism and so on. CONCLUSION: This study originally investigated the dynamic interaction between MLST and the biosystem, providing unique insight for disclosing the active components of MLST and their synergistic mechanisms against TAO, which also shed light on new therapeutic targets for TAO and treatment.
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Medicina Tradicional do Leste Asiático , Tromboangiite Obliterante , Ratos , Animais , Tromboangiite Obliterante/tratamento farmacológico , Tromboangiite Obliterante/induzido quimicamente , Gangrena , Tipagem de Sequências MultilocusRESUMO
(1) Background: Optimal bone mass accumulation during adolescence is crucial for maximising peak bone mass during adulthood. Dietary antioxidant vitamins may contribute to bone mass accumulation. This 2.5-year-long longitudinal study aimed to evaluate the relationships between dietary vitamin A, C, and E intakes and the annual changes in bone parameters among Chinese adolescents. (2) Method: Subjects aged 10-18 years (n = 1418) were recruited from a secondary school in Jiangmen, China. Dietary vitamin A, C, and E intakes were assessed using 24 h dietary records over 3 consecutive days. The Sahara Clinical Bone Sonometer was used to measure the broadband ultrasound attenuation (BUA) and the speed of sound (SOS). Their annual changes were then calculated (i.e., BUA%/year, SOS%/year). The associations were detected after adjusting for the baseline bone phenotype; age; sex; weight; height; pubertal stage; physical activity; and dietary intakes of vitamin D, calcium and energy. (3) Results: A curvilinear relationship was found between the dietary intake of vitamin C and BUA%/year (p = 0.026); further analyses in the subgroups revealed that this relationship was observed in male adolescents (p = 0.012). A positive association was observed only in boys with a dietary vitamin C intake of ≥159.01 mg/day (ß = 0.395, p = 0.036). Moreover, a linear positive association was shown between the dietary intake of vitamin E and BUA%/year in female adolescents (ß = 0.082, p = 0.033). (4) Conclusion: Our findings indicated that dietary vitamin C intake has a threshold effect on bone mass gain in male adolescents and that dietary vitamin E intake could be a positive predictor of bone mass gain in female adolescents.
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Antioxidantes , Calcâneo , Animais , Ácido Ascórbico , Densidade Óssea , Calcâneo/diagnóstico por imagem , Cálcio , Ingestão de Alimentos , Feminino , Estudos Longitudinais , Masculino , Ultrassonografia , Vitamina A , Vitamina D , Vitamina E , VitaminasRESUMO
BACKGROUND: Mailuoshutong pill (MLSTP) is a traditional Chinese medicine (TCM) for the treatment of Thromboangiitis obliterans (TAO, Buerger's disease) which is a segmental non-atherosclerotic inflammatory occlusive disorder. However, the mechanism and quality standards of MLSTP have not been sufficiently studied. PURPOSE: This work aims to investigate the potential mechanisms and quality markers (Q-markers) of MLSTP treating TAO based on the chinmedomics strategy. METHODS: The therapeutical effect of MLSTP on TAO rats was evaluated by changes in body weight and clinical score, regional blood flow velocity and perfused blood vessel distribution, hematoxylin-eosin (H&E) staining, serum metabolic profile. Moreover, both endogenous metabolites and exogenous components were simultaneously detected in serum based on ultra-high performance liquid chromatography coupled with a Q Exactive hybrid quadrupole-orbitrap high resolution mass spectrometry (UHPLC-Q-Orbitrap HRMS), and multivariate analysis was applied to identify the biomarkers, as well as the dynamic changes of metabolites were observed to explore the mechanism of action of MLSTP. In addition, the pharmacodynamic material basis were identified by correlation analysis between biomarkers and absorbed constituents. Finally, the Q-markers of MLSTP were determined according to the screening principles of Q-marker and validated the measurability. RESULTS: MLSTP treatment alleviated disease severity of TAO, reduced inflammatory infiltration, and ameliorated vascular function. 26 potential biomarkers associated with glutamate metabolism, linoleic acid metabolism, arachidonic acid metabolism and so on were identified. Besides, 27 prototypical components were identified in serum, 16 of which were highly correlated with efficacy and could serve as the pharmacodynamic material basis of MLSTP against TAO. In addition, 7 compounds, namely, sweroside, chlorogenic acid, calycosin-7-glucoside, formononetin, paeoniflorin, liquiritigenin and 3-butylidenephthalide, were considered as potential Q-markers of MLSTP. Ultimately, the measurability of the seven Q-markers was validated by rapid identifcation and quantifcation. CONCLUSION: This study successfully clarified the therapeutic effect and Q-markers of MLSTP by chinmedomics strategy, which is of great significance for the establishment of quality standards. Furthermore, it provides a certain reference for the screening of Q-markers in TCM prescriptions.
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Biomarcadores , Medicamentos de Ervas Chinesas , Tromboangiite Obliterante , Animais , Biomarcadores/análise , Cromatografia Líquida de Alta Pressão/métodos , Medicamentos de Ervas Chinesas/química , Medicina Tradicional Chinesa , Metabolômica , Ratos , Tromboangiite Obliterante/tratamento farmacológicoRESUMO
Cholesterol and tocopherols, which are important quality indicators in milk powder, are essential nutrients for the human body. Current pretreatment methods for the detection of cholesterol and four isomers of vitamin E (α-tocopherol, ß-tocopherol, γ-tocopherol, and δ-tocopherol) are based on national food safety standards, which are complicated, time-consuming, and unsuited for simultaneous measurements. Thus, developing a simple, fast, and simultaneous detection method for cholesterol and the four kinds of tocopherols is of practical significance. In this study, gas chromatography-tandem mass spectrometry (GC-MS/MS) was used to establish qualitative and quantitative methods for the determination of cholesterol and the above mentioned four isomers of vitamin E. The sample was digested with lipase and then saponified rapidly using a potassium carbonate-ethanol system. The optimal pretreatment method was established by optimizing the enzymolysis time, saponification temperature, type and volume of the extraction solvent, and extraction time. Then, cholesterol and the four tocopherols in milk power were simultaneously determined. The results revealed a good linear relationship for cholesterol and the tocopherols in the range of 0.5-50.0 mg/L and 0.25-25.0 mg/L, respectively. The correlation coefficients (r2) were greater than 0.99; the recoveries were 76.6%-93.1%; and the relative standard deviations were 0.9%-3.3%. The limits of quantification for cholesterol and the tocopherols were 10.0 µg/100 g and 5.0 µg/100 g, respectively. The recoveries of the added standards did not fully reflect the ability of the method to decompose and extract the actual sample, especially given that the five compounds considered in this study were fat-soluble. Thus, the added standard recovery could not verify the enzymatic hydrolysis effect. In order to investigate the effectiveness of this method for actual milk powder samples, the amounts of cholesterol and the four tocopherols in infant milk powder were determined according to the national standard methods (GB 5009.82-2016, GB 5009.128-2016) and the proposed method. For each method, six sets of measurements were carried out in parallel. The cholesterol content measured by this method was slightly lower than that measured by the national standard method, while the amounts of the four tocopherols were slightly higher. There was no significant difference (p> 0.05) between the national standard method and our method on the amounts of cholesterol and the four tocopherols in milk powder. Twenty kinds of infant formula milk powder and four kinds of low-fat milk powder were randomly selected from the market, and the amounts of cholesterol and the four tocopherols were analyzed. The results showed that the amounts of cholesterol and the four tocopherols in the infant formula milk powder were higher than those in the low-fat milk powder. This method is simple, fast, sensitive, and accurate, thus meeting the detection requirements for cholesterol and tocopherols in milk powder. The findings of the study would provide a theoretical foundation for the rapid estimation of milk powder quality.
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Colesterol/análise , Leite/química , Vitamina E/análise , Animais , Cromatografia Líquida de Alta Pressão , Cromatografia Gasosa-Espectrometria de Massas , Fórmulas Infantis/análise , Pós , Espectrometria de Massas em TandemRESUMO
BACKGROUND: Metastatic breast cancer (MBC) remains an incurable disease worldwide. Tumor gene mutations have evolved and led to drug resistance in the treatment course of MBC. However, data on the mutation profiles and druggable genomic alterations of MBC remain limited, particularly among Chinese patients. Our study aimed to depict the mutation profiles and identify druggable mutations in circulating tumor DNA (ctDNA) in Chinese MBC patients. METHODS: Targeted deep sequencing of a 1021-gene panel was performed on 17 blood samples and 5 available tissue samples from 17 Chinese MBC patients. RESULTS: We identified 60 somatic mutations in 17 blood samples (sensitivity 100%). Somatic mutations were identified in the blood samples of all patients, and 41.18% (7/17) of patients harbored at least one druggable mutation. A high ctDNA level in plasma is associated with shorter progression-free survival. CONCLUSION: Targeted deep sequencing of cell free DNA is a highly sensitive, noninvasive method to depict tumor mutation profiles, identify druggable mutations in MBC, and predict patient outcome. Our study shed light on the utility of ctDNA as noninvasive "liquid biopsy" in the management of MBC.
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Povo Asiático/genética , Neoplasias da Mama/genética , DNA Tumoral Circulante/genética , Análise Mutacional de DNA/métodos , Adulto , Ácidos Nucleicos Livres/genética , China , Feminino , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Humanos , Biópsia Líquida , Pessoa de Meia-Idade , Metástase Neoplásica , Sensibilidade e Especificidade , Análise de Sequência de DNA/métodosRESUMO
One major strategy to generate genetically modified mouse models is gene targeting in mouse embryonic stem (ES) cells, which is used to produce gene-targeted mice for wide applications in biomedicine. However, a major bottleneck in this approach is that the robustness of germline transmission of gene-targeted ES cells can be significantly reduced by their genetic and epigenetic instability after long-term culturing, which impairs the efficiency and robustness of mouse model generation. Recently, we have established a new type of pluripotent cells termed extended pluripotent stem (EPS) cells, which have superior developmental potency and robust germline competence compared to conventional mouse ES cells. In this study, we demonstrate that mouse EPS cells well maintain developmental potency and genetic stability after long-term passage. Based on gene targeting in mouse EPS cells, we established a new approach to directly and rapidly generate gene-targeted mouse models through tetraploid complementation, which could be accomplished in approximately 2 months. Importantly, using this approach, we successfully constructed mouse models in which the human interleukin 3 (IL3) or interleukin 6 (IL6) gene was knocked into its corresponding locus in the mouse genome. Our study demonstrates the feasibility of using mouse EPS cells to rapidly generate mouse models by gene targeting, which have great application potential in biomedical research.