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Mass spectrometry, coupled with innovative crosslinking techniques to decode protein conformations and interactions through uninterrupted signal connections, has undergone remarkable progress in recent years. It is crucial to develop selective crosslinking reagents that minimally disrupt protein structure and dynamics, providing insights into protein network regulation and biological functions. Compared to traditional crosslinkers, new bifunctional chemical crosslinkers exhibit high selectivity and specificity in connecting proximal amino acid residues, resulting in stable molecular crosslinked products. The conjugation with specific amino acid residues like lysine, cysteine, arginine and tyrosine expands the XL-MS toolbox, enabling more precise modeling of target substrates and leading to improved data quality and reliability. Another emerging crosslinking method utilizes unnatural amino acids (UAAs) derived from proximity-enabled reactivity with specific amino acids or sulfur-fluoride exchange (SuFEx) reactions with nucleophilic residues. These UAAs are genetically encoded into proteins for the formation of specific covalent bonds. This technique combines the benefits of genetic encoding for live cell compatibility with chemical crosslinking, providing a valuable method for capturing transient and weak protein-protein interactions (PPIs) for mapping PPI coordinates and improving the pharmacological properties of proteins. With continued advancements in technology and applications, crosslinking mass spectrometry is poised to play an increasingly significant role in guiding our understanding of protein dynamics and function in the future.
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Poria Cum Radix Pini is a rare medicinal fungus that contains several potential therapeutic ingredients. On this basis, a particle swarm mathematical model was used to optimize the extraction process of total triterpenes from P. Cum Radix Pini, and xanthine oxidase inhibitors were screened using affinity ultrafiltration mass spectrometry. Meanwhile, the accuracy of the ultrafiltration assay was verified by molecular docking experiments and molecular dynamics analysis, and the mechanism of action of the active compounds for the treatment of gout was analyzed by enzymatic reaction kinetics and network pharmacology. A high-speed countercurrent chromatography method combined with the consecutive injection and the economical two-phase solvent system preparation using functional activity coefficient of universal quasichemical model (UNIFAC) mathematical model was developed for increasing the yield of target compound. In addition, dehydropachymic acid and pachymic acid were used as competitive inhibitors, and 3-O-acetyl-16alpha-hydroxydehydrotrametenolic acid and dehydrotrametenolic acid were used as mixed inhibitors. Then, activity-oriented separation and purification were performed by high-speed countercurrent chromatography combined with semi-preparative high-performance liquid chromatography and the purity of the four compounds isolated was higher than 90%. It will help to provide more opportunities to discover and develop new potential therapeutic remedies from health care food resources.
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Gota , Poria , Poria/química , Xantina Oxidasa , Simulación del Acoplamiento Molecular , Cromatografía Líquida de Alta Presión/métodos , Inhibidores Enzimáticos/farmacología , Distribución en Contracorriente , Gota/tratamiento farmacológicoRESUMEN
INTRODUCTION: Studies show that Polyporus umbellatus has some pharmacological effects in enhancing immunity and against gout. OBJECTIVES: We aimed to establish new techniques for extraction, biological activity screening, and preparation of xanthine oxidase inhibitors (XODIs) from P. umbellatus. METHODS: First, the extraction of P. umbellatus was investigated using the back propagation (BP) neural network genetic algorithm mathematical regression model, and the extraction variables were optimised to maximise P. umbellatus yield. Second, XODIs were rapidly screened using ultrafiltration, and the change of XOD activity was tested by enzymatic reaction kinetics experiment to reflect the inhibitory effect of active compounds on XOD. Meanwhile, the potential anti-gout effects of the obtained active substances were verified using molecular docking, molecular dynamics simulations, and network pharmacology analysis. Finally, with activity screening as guide, a high-speed countercurrent chromatography (HSCCC) method combined with consecutive injection and two-phase solvent system preparation using the UNIFAC mathematical model was successfully developed for separation and purification of XODIs, and the XODIs were identified using MS and NMR. RESULTS: The results verified that polyporusterone A, polyporusterone B, ergosta-4,6,8(14),22-tetraen-3-one, and ergosta-7,22-dien-3-one of P. umbellatus exhibited high biological affinity towards XOD. Their structures have been further identified by NMR, indicating that the method is effective and applicable for rapid screening and identification of XODIs. CONCLUSION: This study provides new ideas for the search for natural XODIs active ingredients, and the study provide valuable support for the further development of functional foods with potential therapeutic benefits.
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Polyporus , Xantina Oxidasa , Simulación del Acoplamiento Molecular , Polyporus/química , Inhibidores Enzimáticos/farmacologíaRESUMEN
INTRODUCTION: Sophora flavescens Aiton (Fabaceae), a ubiquitous plant species in Asia, contains a wide range of pharmacologically active compounds, such as flavonoids, with potential anti-Alzheimer's disease (anti-AD) effects. OBJECTIVES: The objective of the study is to develop a quaternity method for the screening, isolation, extraction optimization, and activity evaluation of acetylcholinesterase (AChE)-inhibiting compounds from S. flavescens to realize high-throughput screening of active substances in traditional Chinese medicine and to provide experimental data for the development of anti-AD drugs. METHODS: With AChE as the target molecule, affinity ultrafiltration and liquid chromatography-mass spectrometry were applied to screen for potential inhibitors of the enzyme in S. flavescens. Orthogonal array experiments combined with the multi-objective Non-Dominated Sorting Genetic Algorithm III was used for the first time to optimize the process for extracting the active substances. Enzyme inhibition kinetics and molecular docking studies were performed to verify the potential anti-AD effects of the active compounds. RESULTS: Five AChE-inhibiting compounds were identified: kushenol I, kurarinone, sophoraflavanone G, isokurarinone, and kushenol E. These were successfully separated at purities of 72.88%, 98.55%, 96.86%, 96.74%, and 95.84%, respectively, using the n-hexane/ethyl acetate/methanol/water (4.0/5.0/4.0/5.0, v/v/v/v), n-hexane/ethyl acetate/methanol/water (5.0/5.0/6.0/4.0, v/v/v/v), and n-hexane/ethyl acetate/methanol/water (4.9/5.1/5.7/4.3, v/v/v/v) mobile phase systems. Enzyme inhibition kinetics revealed that kushenol E had the best inhibitory effect. CONCLUSION: This study elucidates the mechanism of action of five active AChE inhibitors in S. flavescens and provides a theoretical basis for the screening and development of anti-AD and other therapeutic drugs.
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INTRODUCTION: Accurate screening and targeted preparative isolation of active substances from natural medicines have long been technical challenges in natural medicine research. OBJECTIVES: This study outlines a new approach for improving the efficiency of natural product preparation, focusing on the rapid and accurate screening of potential active ingredients in Ganoderma lucidum and efficient preparation of lipoxidase inhibitors, with the aim of providing new ideas for the treatment of Alzheimer's disease with G. lucidum. METHODS: The medicinal plant G. lucidum was selected through ultrafiltration coupled with liquid chromatography and mass spectrometry (UF-LC-MS) and computer-assisted screening for lipoxygenase (LOX) inhibitors. In addition, the inhibitory effect of the active compounds on LOX was studied using enzymatic reaction kinetics, and the underlying mechanism is discussed. Finally, based on the earlier activity screening guidelines, the identified ligands were isolated and purified through complex chromatography (high-speed countercurrent chromatography and semi-preparative high-performance liquid chromatography). RESULTS: Five active ingredients, ganoderic acids A, B, C2, D2, and F, were identified and isolated from G. lucidum. We improved the efficiency and purity of active compound preparation using virtual computer screening and enzyme inhibition assays combined with complex chromatography. CONCLUSION: The innovative methods of UF-LC-MS, computer-aided screening, and complex chromatography provide powerful tools for screening and separating LOX inhibitors from complex matrices and provide a favourable platform for the large-scale production of bioactive substances and nutrients.
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Antineoplásicos , Reishi , Inhibidores de la Lipooxigenasa/farmacología , Cromatografía Líquida de Alta Presión , Distribución en ContracorrienteRESUMEN
Baccatin III is a crucial precursor in the biosynthesis pathway of paclitaxel. Its main sources are extraction from Taxus or chemical synthesis using 10-deacetylbaccatin III (10-DAB) as substrate. However, these preparation approaches exhibit serious limitations, including the low content of baccatin III in Taxus and the complicated steps of chemical synthesis. Heterologous expression of 10-deacetylbaccatin III-10-O-acetyltransferase (TcDBAT) in microbial strains for biotransformation of 10-DAB is a promising alternative strategy for baccatin III production. Here, the promotion effects of glycerol supply and slightly acidic conditions with a low-temperature on the catalysis of recombinant TcDBAT strain were clarified using 10-DAB as substrate. Taxus needles is renewable and the content of 10-DAB is relatively high, it can be used as an effective source of the catalytic substrate 10-DAB. Baccatin III was synthesized by integrating the extraction of 10-DAB from renewable Taxus needles and in situ whole-cell catalysis in this study. 40 g/L needles were converted into 20.66 mg/L baccatin III by optimizing and establishing a whole-cell catalytic bioprocess. The method used in this study can shorten the production process of Taxus extraction for baccatin III synthesis and provide a reliable strategy for the efficient production of baccatin III by recombinant strains and the improvement of resource utilization rate of Taxus needles.
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Biotransformación , Taxoides , Taxus , Taxus/metabolismo , Taxus/química , Taxoides/metabolismo , Alcaloides/biosíntesis , Alcaloides/metabolismo , Alcaloides/química , Hojas de la Planta/metabolismo , Hojas de la Planta/química , Acetiltransferasas/metabolismo , Acetiltransferasas/genéticaRESUMEN
In this study, an efficient method that employs 5-lipoxygenase and acetylcholinesterase as biological target molecules in receptor-ligand affinity ultrafiltration-liquid chromatography was developed for the screening of enzyme inhibitors derived from the Astragalus membranaceus stems and leaves. The effects of the extraction time, number of extraction cycles, ethanol concentration, and liquid-solid ratio on the total yield of the target compounds were investigated using response surface methodology, and the bioactive components were isolated using a combination of semi-preparative high-performance liquid chromatography and high-speed countercurrent chromatography via a two-phase solvent system consisting of n-hexane-ethyl acetate-methanol-water (1:6:2:6, v/v/v/v). Subsequently, 10 naturally-occurring bioactive components in the Astragalus membranaceus stems and leaves, including wogonin, ononin, isoquercitrin, calycosin-7-glucoside, 3-hydroxy-9,10-dimethoxyptercarpan, hyperoside, 7,2'-dihydroxy-3',4'-dimethoxyisoflavan, baicalein, calycosin, and soyasaponin, were screened using affinity ultrafiltration to determine their potential effects against Alzheimer's disease. Consequently, all target compounds had purities higher than 95.0%, and the potential anti-Alzheimer's disease effect of the obtained bioactive compounds was verified using molecular docking analysis. Based on the results, the back-to-back screening of complex enzyme inhibitors and separation of the target bioactive compounds using complex chromatography could provide a new approach to the discovery and preparation of natural active ingredients.
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Acetilcolinesterasa , Astragalus propinquus , Astragalus propinquus/química , Inhibidores de la Colinesterasa , Araquidonato 5-Lipooxigenasa , Simulación del Acoplamiento Molecular , Cromatografía Líquida de Alta Presión/métodos , Distribución en Contracorriente/métodosRESUMEN
INTRODUCTION: Cicer arietinum L. is the choice of health food for people with diabetes, hypertension, and hyperlipidemia. As an essential source of high-nutrition legumes, it is also an important source of dietary isoflavones. OBJECTIVES: In order to improve the preparation efficiency of natural plants, a rapid biological activity screening and preparation of xanthine oxidase inhibitors from C. arietinum L. was established. METHODS: Xanthine oxidase (XOD) inhibitors were rapidly screened using ultrafiltration liquid chromatography-mass spectrometry (UF-LC-MS) based on receptor-ligand affinity. The change in XOD activity was evaluated by enzymatic reaction kinetics measurement. The potential bioactive compounds were verified through molecular docking. In addition, the biological activity of ligands screened was separated and purified by complex chromatography. The structures of the compounds were identified by nuclear magnetic resonance spectroscopy. RESULTS: Three active ingredients, namely daidzin, daidzein, calycosin with XOD binding affinities were identified and isolated from the raw plant materials via semi-preparative high-performance liquid chromatography (HPLC), 0-60 min, 5-50% B and countercurrent chromatography (CCC) (ethyl acetate/acetic acid/water [5:0.8:10, v/v/v]). CONCLUSION: This study will help to elucidate the mechanisms of action of natural plants of interest at the molecular level and could also provide more opportunities for the discovery and development of new nutritional value from other natural resources.
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Cicer , Xantina Oxidasa , Humanos , Cicer/metabolismo , Simulación del Acoplamiento Molecular , Ligandos , Cromatografía Liquida/métodos , Inhibidores Enzimáticos/farmacología , Cromatografía Líquida de Alta Presión/métodosRESUMEN
INTRODUCTION: The spores of the medicinal fungus Ganoderma lucidum possess hepatoprotective properties. The main components, triterpenes, are particularly beneficial, making the screening and preparation of active triterpenes from Ganoderma lucidum significant. OBJECTIVES: We aimed to screen and verify cyclooxygenase-2 inhibitors from G. lucidum spores, establish a rapid online hyphenated technique for the preparation of active ingredients, and analyze the structures of the active ingredients. METHODS: Ultrafiltration LC combined with an enzyme inhibition assay and molecular docking was employed to screen and evaluate cyclooxygenase-2 ligands, which were prepared by pressurized liquid extraction coupled online with countercurrent chromatography and semi-preparative LC. The structures of the compounds were identified by nuclear magnetic resonance spectroscopy. RESULTS: Six cyclooxygenase-2 inhibitors, namely, ganoderic acids I, C2 , G, B, and A and ganoderenic acid A, were screened and evaluated. They were prepared using the online hyphenated technique, following which their structures were identified. CONCLUSION: This study provides opportunities for the discovery and development of new therapeutic drugs from other natural resources, as the present instrumental setup achieved efficient and systematic extraction and isolation of natural products compared with reference separation methods, thus exhibiting significant potential for industrial applications.
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Reishi , Triterpenos , Reishi/química , Inhibidores de la Ciclooxigenasa 2/farmacología , Inhibidores de la Ciclooxigenasa 2/análisis , Simulación del Acoplamiento Molecular , Esporas Fúngicas/química , Triterpenos/análisis , Distribución en ContracorrienteRESUMEN
Poria cocos (P. cocos) is a traditional Chinese medicinal product with the same origin as medicine and food. It has diuretic, anti-inflammatory and liver protection properties, and has been widely used in a Chinese medicine in the treatment of Alzheimer's disease (AD). This study was conducted to explore the activity screening, isolation of acetylcholinesterase inhibitors (AChEIs), and in vitro inhibiting effect of P. cocos. The aim was to develop a new extraction process optimization method based on the Matlab genetic algorithm combined with a traditional orthogonal experiment. Moreover, bio-affinity ultrafiltration combined with molecular docking was used to screen and evaluate the activity of the AChEIs, which were subsequently isolated and purified using high-speed counter-current chromatography (HSCCC) and semi-preparative high-performance liquid chromatography (semi-preparative HPLC). The change in acetylcholinesterase (AChE) activity was tested using an enzymatic reaction kinetics experiment to reflect the inhibitory effect of active compounds on AChE and explore its mechanism of action. Five potential AChEIs were screened via bio-affinity ultrafiltration. Molecular docking results showed that they had good binding affinity for the active site of AChE. Meanwhile, the five active compounds had reversible inhibitory effects on AChE: Polyporenic acid C and Tumulosic acid were non-competitive inhibitors; 3-Epidehydrotumulosic acid was a mixed inhibitor; and Pachymic acid and Dehydrotrametenolic acid were competitive inhibitors. This study provided a basis for the comprehensive utilization of P. cocos and drug development for the treatment of AD.
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Enfermedad de Alzheimer , Poria , Wolfiporia , Inhibidores de la Colinesterasa/farmacología , Inhibidores de la Colinesterasa/análisis , Acetilcolinesterasa , Simulación del Acoplamiento Molecular , Wolfiporia/química , Cromatografía Líquida de Alta Presión/métodos , Poria/químicaRESUMEN
We study continuous variable coherence of phase-dependent squeezed state based on an extended Hanbury Brown-Twiss scheme. High-order coherence is continuously varied by adjusting squeezing parameter r, displacement α, and squeezing phase θ. We also analyze effects of background noise γ and detection efficiency η on the measurements. As the squeezing phase shifts from 0 to π, the photon statistics of the squeezed state continuously change from the anti-bunching (g(n) < 1) to super-bunching (g(n) > n!) which shows a transition from particle nature to wave nature. The experiment feasibility is also examined. It provides a practical method to generate phase-dependent squeezed states with high-order continuous-variable coherence by tuning squeezing phase θ. The controllable coherence source can be applied to sensitivity improvement in gravitational wave detection and quantum imaging.
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We focus on the generation of mechanical squeezing by using periodically amplitude-modulated laser to drive an active-passive-coupled double-cavity optomechanical system, where the coupled gain cavity and loss cavity can form into a parity-time (P T)-symmetry system. The numerical analysis of the system stability shows that the system is more likely to be stable in the unbroken-P T-symmetry regime than in the broken-P T-symmetry regime. The mechanical squeezing in the active-passive system exhibits stronger robustness against the thermal noise than that in the passive-passive system, and the so-called 3 dB limit can be broken in the resolved-sideband regime. Furthermore, it is also found that the mechanical squeezing obtained in the unbroken-P T-symmetry region is stronger than that in the broken-P T-symmetry region. This work may be meaningful for the quantum state engineering in the gain-loss quantum system that contributes to the study of P T-symmetric physics in the quantum regime.
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INTRODUCTION: This study aims to investigate the feasibility of a noninvasive handheld electroporation pulses delivery device (EPDD) for electroporation-based treatment (EBT) of skin superficial lesions through numerical analysis and animal study. METHODS: Finite element analysis was performed to investigate the performance of the EPDD. The electric field, temperature, EI and TI were calculated under pulse voltages of 600, 800, and 1000 V. A mouse subcutaneous tumor model was established to evaluate the performance of the EPDD through histopathology and survival analyses. RESULTS: The electrical field strength increased from 151 (600 V) to 252 V/cm (1000 V) in the skin and from 1302 (600 V) to 2171 V/cm (1000 V) in the tumor. The volume of EI grew and reached a plateau at the 165th pulse, whereas the maximum volume of EI increased with higher voltage. The growth tendency of TI differed between groups, and it was higher in the high-voltage group (HVG) than in the low-voltage group. Histopathological analysis showed that the depth and range of the ablation area could be controlled by adjusting pulse voltage. Survival analysis showed that the survival of the HVG was better than that of the low-voltage and the control group (p < 0.01). CONCLUSIONS: The results demonstrate that the EPDD is feasible, safe, and effective for skin EBT. The volume of EP tissue injury can be controlled by adjusting the pulse voltage, pulse number, and other parameters. The proposed noninvasive handheld EPDD can be a potential therapeutic tool for EBT of superficial skin lesions in the future.
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Electroporación , Piel , Animales , Modelos Animales de Enfermedad , Electroporación/métodos , RatonesRESUMEN
Adolescence is a significant period for the formation of relationship networks and the development of internalizing problems. With a sample of Chinese adolescents (N = 3,834, 52.01% girls, Mage = 16.68 at Wave 1), the present study aimed to identify the configuration of adolescents' relationship qualities from four important domains (i.e., relationship quality with mother, father, peers, and teachers) and how distinct profiles were associated with the development of internalizing problems (indicated by depressive and anxiety symptoms) across high school years. Latent profile analysis identified a five-profile configuration with four convergent profiles (i.e., relationship qualities with others were generally good or bad) and one "Father estrangement" profile (i.e., the relationship quality with others were relatively good but that with father was particularly poor). Further conditional latent growth curve analysis indicated the "Father estrangement" profile was especially vulnerable to an increase in the internalizing problems as compared with other relationship profiles. This study contributes to understanding the characteristics of interpersonal relationship qualities and their influences on adolescent internalizing problems in a non-Western context. Results were further discussed from a culturally specific perspective.
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A simple and efficient method was developed for the rapid screening and identification of ligands for monoamine oxidase B. A new ionic-liquid-based ultrasound-assisted extraction method for medicinal herbs was also developed and validated. In addition, the hyphenated technique of countercurrent chromatography and semipreparative-LC was developed and applied to the isolation of the chemical constituents for Pueraria thomsonii Benth. Three potent monoamine oxidase B inhibitors, namely, daidzein-4',7-diglucoside (42.2 mg), puerarin 6''-O-xyloside (88.3 mg), and 3'-hydroxypuerarin (48.5 mg) with purities of 98.2, 96.3, and 97.1%, respectively, were obtained from 500 g of P. thomsonii raw material using semi-preparative high-performance liquid chromatography, whereas 3'-methoxypuerarin (76.2 mg), daidzein-8-C-apiosyl (1â6) glucoside (84.2 mg), and tectorigenin (75.1 mg) with purities of 98.5, 96.4, and 96.8%, respectively, were obtained from 500 g raw material via countercurrent chromatography using a two-phase solvent system comprising n-hexane-ethyl acetate-methanol-water at a volume ratio of 1.85:1.00:0.86:3.69 (v/v/v/v). Then, the anti-Alzheimer activity of the phytochemicals was assessed using a PC12 cell model. Treatment with tectorigenin, daidzein-4',7-diglucoside, puerarin 6''-O-xyloside, 3'-hydroxypuerarin, 3'-methoxypuerarin, and daidzein-8-C-apiosyl (1â6) glucoside (100 µg/mL), resulted in cell viabilities of 69.00, 65.81, 59.69, 57.90, 55.61, and 54.59%, respectively (p < 0.001). The protocol was proved to be very accurate and efficient.
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Líquidos Iónicos , Pueraria , Cromatografía Líquida de Alta Presión/métodos , Distribución en Contracorriente/métodos , Monoaminooxidasa , Pueraria/químicaRESUMEN
The extraction of Scutellaria baicalensis Georgi was investigated using the response surface methodology-genetic algorithm mathematical regression model, and the extraction variables were optimized to maximize the flavonoid yield. Furthermore, a simple and efficient ultrafiltration-liquid chromatography-mass spectrometry and molecular docking methods were developed for the rapid screening and identification of acetylcholinesterase inhibitors present in Scutellaria baicalensis Georgi. Subsequently, four major chemical constituents, namely baicalein, norwogonin, wogonin, and oroxylin A, were identified as potent acetylcholinesterase inhibitors. This novel approach, involving the use of ultrafiltration-liquid chromatography-mass spectrometry and molecular docking methods combined with stepwise flow rate counter-current chromatography and semi-preparative high-performance liquid chromatography, could potentially provide a powerful tool for the screening and extraction of acetylcholinesterase inhibitors from complex matrices and be a useful platform for the production of bioactive and nutraceutical ingredients.
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Inhibidores de la Colinesterasa , Scutellaria baicalensis , Acetilcolinesterasa , Cromatografía Líquida de Alta Presión/métodos , Flavonoides , Simulación del Acoplamiento Molecular , Scutellaria baicalensis/químicaRESUMEN
Black bean, in which isoflavones are the main active constituent, also contains saponins and monoterpenes. Soybean isoflavone is a secondary metabolite that is formed during the growth of soybean; it exhibits antioxidant and cardiovascular activities and traces estrogen-like effects. In this study, black bean isoflavones were extracted with n-butanol, and ultrafiltration-liquid chromatography-mass spectrometry was used to screen their activity. Subsequently, the inhibitors were isolated and purified using semipreparative liquid chromatography and stepwise flow rate countercurrent chromatography. Thereafter, five active compounds were identified using mass spectrometry and nuclear magnetic resonance experiments. Finally, the inhibition types of the xanthine oxidase inhibitors were determined using enzymatic kinetic studies. The IC50 values of daidzin, glycitein-7-O-glucoside, genistin, daidzein, and genistein were determined to be 35.08, 56.22, 30.76, 68.79, and 95.37 µg/mL, respectively. Daidzin, genistin, and daidzein exhibited reversible inhibition, whereas glycitein-7-O-glucoside and genistein presented irreversible inhibition. This novel approach, which was based on ultrafiltration-liquid chromatography-mass spectrometry and stepwise flow rate countercurrent chromatography, is a powerful method for screening and isolating xanthine oxidase inhibitors from complex matrices. The study of enzyme inhibition types is helpful for understanding the underlying inhibition mechanism. Therefore, a beneficial platform was developed for the large-scale production of bioactive and nutraceutical ingredients.
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Distribución en Contracorriente , Isoflavonas , Xantina Oxidasa , Cromatografía Líquida de Alta Presión/métodos , Cromatografía Liquida , Isoflavonas/química , Cinética , Phaseolus/química , Proteínas de Plantas/química , Xantina Oxidasa/antagonistas & inhibidoresRESUMEN
Alzheimer's disease (AD) is a common dementia disease in the elderly. To get a better understanding of the pathophysiology, we performed a proteomic analysis of the urine exosomes (U-exo) in AD model mice (J20). The polymer precipitation method was used to isolate U-exo from the urine of 3-month-old J20 and wild-type (WT) mice. Neuron-derived exosome (N-exo) was isolated from U-exo by immunoprecipitation. iTRAQ-based MALDI TOF MS/MS was used for proteomic analysis. The results showed that compared to WT, the levels of 61 and 92 proteins were increased in the J20 U-exo and N-exo, respectively. Gene ontology enrichment analysis demonstrated that the sphingolipid catabolic process, ceramide catabolic process, membrane lipid catabolic process, Aß clearance, and Aß metabolic process were highly enriched in U-exo and N-exo. Among these, Asah1 was shown to be the key protein in lipid metabolism, and clusterin, ApoE, neprilysin, and ACE were related to Aß metabolism and clearance. Furthermore, protein-protein interaction analysis identified four protein complexes where clusterin and ApoE participated as partner proteins. Thus, J20 U-exo and N-exo contain proteins related to lipid- and Aß-metabolism in the early stages of AD, providing a new insight into the underlying pathological mechanism of early AD.
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Enfermedad de Alzheimer , Exosomas , Ratones , Animales , Ratones Transgénicos , Péptidos beta-Amiloides/metabolismo , Clusterina/metabolismo , Exosomas/metabolismo , Espectrometría de Masas en Tándem , Proteómica , Enfermedad de Alzheimer/metabolismo , Apolipoproteínas E/metabolismo , Modelos Animales de Enfermedad , Precursor de Proteína beta-Amiloide/metabolismoRESUMEN
BACKGROUND: Tertiary lymphoid structures (TLSs), organizationally resemble lymph nodes, are frequently present in breast cancer (BCa). It is usually, but not always, associated with a positive prognosis or immunotherapy response in cancer patients. This meta-analysis was performed to assess the prognostic and clinical impact of TLSs in BCa. METHODS: We conducted a systematic search in PubMed, Embase, Cochrane Library, Web of Science, China National Knowledge Infrastructure, and WanFang Database to obtain eligible research data up to May 30, 2021. This meta-analysis is focusing on the studies evaluated the prognostic value of TLSs and the associated clinicopathologic indicators, related gene expression and survival. STATA software 16.0 software was used to assess the prognostic significance and clinical impact of TLSs. RESULTS: Nine studies involved with 2281 cases were incorporated in this meta-analysis, in which four of them evaluated the prognostic value of TLSs. There are 6 studies assessed the relationship of TLSs and 4 studies investigated the clinicopathologic parameters as well as the key gene expression, respectively. The results showed the presence of TLSs were predicting a better OS (HR = 0.61, 95% CI: 0.51-0.73, p < 0.001) and DFS (HR = 0.40, 95% CI: 0.17-0.93, p < 0.001) of BCa patients. It also revealed that the presence of TLSs was significantly correlated with tumor differentiation (p < 0.001), pTNM stage (p < 0.001), lymph node metastasis (p < 0.001), and TILs density (p < 0.001) of BCa, and the expression of Her2 (p < 0.001), ER (p < 0.001), PR (p < 0.001) and Ki67 (p = 0.009) of the tumor cell. CONCLUSION: Our results indicated that high levels of TLSs could predict a favorable prognosis for BCa. Moreover, the TLSs were significantly correlated with the clinicopathological indicators and the critical gene expression of BCa, indicating its potential clinical impact on BCa patients.
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BACKGROUND: Irreversible electroporation (IRE) is an emerging tissue ablation technique with widespread potential, especially for cancer treatment. Although the safety and efficacy of IRE for gastric tissue ablation have been demonstrated, there is a gap of knowledge regarding the effect of electroporation pulse (EP) on the physiology and histopathology of the stomach. This study applied EP to the stomach of healthy rats and investigated the digestive function, serum marker levels, and gastric tissue structure of EP-treated rats. METHODS: Ninety male rats were divided into nine groups and examined up to 28 days post-treatment. A single burst of electroporation pulse (500 V, 99 pluses, 1 Hz, 100 µs) was delivered to the stomachs of rats using a tweezer-style round electrode. Gastric emptying, small intestinal transit, and gastric secretion were measured to evaluate the digestive function. Serum marker levels were determined using ELISA. Haematoxylin-eosin, Masson trichrome, and immunofluorescence were performed for histopathological analysis. RESULTS: No significant effect on gastric emptying or secretion was found post-EP, whereas the small intestinal transit decreased at 4 h and rapidly recovered to normal on 1-day post-EP. Further, serum TNF-α and IL-1ß levels temporarily changed during the acute phase but returned to baseline within 28 days. Moreover, histopathological analysis revealed that cell death occurred immediately post-EP in the ablation area, whereas the gastric wall scaffold in the ablation region remained intact post-EP. CONCLUSIONS: This study demonstrates the safety and efficacy of EP on the physiology and histopathology of the stomach and lays a foundation for more comprehensive applications of this technique.