European Biophysics Journal最新文献_第8页

Physics of collective cell migration 集体细胞迁移的物理学。

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-09-14 DOI: 10.1007/s00249-023-01681-w

Ivana Pajic-Lijakovic, Milan Milivojevic

{"title":"Physics of collective cell migration","authors":"Ivana Pajic-Lijakovic, Milan Milivojevic","doi":"10.1007/s00249-023-01681-w","DOIUrl":"10.1007/s00249-023-01681-w","url":null,"abstract":"<div><p>Movement of cell clusters along extracellular matrices (ECM) during tissue development, wound healing, and early stage of cancer invasion involve various inter-connected migration modes such as: (1) cell movement within clusters, (2) cluster extension (wetting) and compression (de-wetting), and (3) directional cluster movement. It has become increasingly evident that dilational and volumetric viscoelasticity of cell clusters and their surrounding substrate significantly influence these migration modes through physical parameters such as: tissue and matrix surface tensions, interfacial tension between cells and substrate, gradients of surface and interfacial tensions, as well as, the accumulation of cell and matrix residual stresses. Inhomogeneous distribution of tissue surface tension along the cell–matrix biointerface can appear as a consequence of different contractility of various cluster regions. While the directional cell migration caused by the matrix stiffness gradient (i.e., durotaxis) has been widely elaborated, the structural changes of matrix surface caused by cell tractions which lead to the generation of the matrix surface tension gradient has not been considered yet. The main goal of this theoretical consideration is to clarify the roles of various physical parameters in collective cell migration based on the formulation of a biophysical model. This complex phenomenon is discussed with the help of model systems such as the movement of cell clusters on a collagen I gel matrix, simultaneously reviewing various experimental data with and without cells.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 8","pages":"625 - 640"},"PeriodicalIF":2.0,"publicationDate":"2023-09-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10231056","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

MIL-CELL: a tool for multi-scale simulation of yeast replication and prion transmission MIL-CELL:用于酵母复制和朊病毒传播的多尺度模拟工具。

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-09-05 DOI: 10.1007/s00249-023-01679-4

Damien Hall

{"title":"MIL-CELL: a tool for multi-scale simulation of yeast replication and prion transmission","authors":"Damien Hall","doi":"10.1007/s00249-023-01679-4","DOIUrl":"10.1007/s00249-023-01679-4","url":null,"abstract":"<div><p>The single-celled baker’s yeast, <i>Saccharomyces cerevisiae</i>, can sustain a number of amyloid-based prions, the three most prominent examples being [URE3], [PSI+], and [PIN+]. In the laboratory, haploid <i>S. cerevisiae</i> cells of a single mating type can acquire an amyloid prion in one of two ways (i) spontaneous nucleation of the prion within the yeast cell, and (ii) receipt via mother-to-daughter transmission during the cell division cycle. Similarly, prions can be lost due to (i) dissolution of the prion amyloid by its breakage into non-amyloid monomeric units, or (ii) preferential donation/retention of prions between the mother and daughter during cell division. Here we present a computational tool (<i>M</i>onitoring <i>I</i>nduction and <i>L</i>oss of prions in <i>Cell</i>s; MIL-CELL) for modelling these four general processes using a multiscale approach describing both spatial and kinetic aspects of the yeast life cycle and the amyloid-prion behavior. We describe the workings of the model, assumptions upon which it is based and some interesting simulation results pertaining to the wave-like spread of the epigenetic prion elements through the yeast population. MIL-CELL is provided as a stand-alone GUI executable program for free download with the paper. MIL-CELL is equipped with a relational database allowing all simulated properties to be searched, collated and graphed. Its ability to incorporate variation in heritable properties means MIL-CELL is also capable of simulating loss of the isogenic nature of a cell population over time. The capability to monitor both chronological and reproductive age also makes MIL-CELL potentially useful in studies of cell aging.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 8","pages":"673 - 704"},"PeriodicalIF":2.0,"publicationDate":"2023-09-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://link.springer.com/content/pdf/10.1007/s00249-023-01679-4.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10515694","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

N2 modified cap analogues as translation inhibitors and substrates for preparation of therapeutic mRNA N2修饰的帽类似物作为翻译抑制剂和用于制备治疗性信使核糖核酸的底物。

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-09-01 DOI: 10.1007/s00249-023-01676-7

Karol Kurpiejewski, Marzena Jankowska-Anyszka, Renata Grzela

{"title":"N2 modified cap analogues as translation inhibitors and substrates for preparation of therapeutic mRNA","authors":"Karol Kurpiejewski, Marzena Jankowska-Anyszka, Renata Grzela","doi":"10.1007/s00249-023-01676-7","DOIUrl":"10.1007/s00249-023-01676-7","url":null,"abstract":"<div><p>In recent years many scientists have begun to focus on the mRNA molecule’s emeregence as a new type of drug. Its fast-moving and successful career as a vaccine technology cannot be underestimated. mRNA provides new opportunities and allows for the rapid preparation of effective drugs at low cost. These extensive possibilities stem from a number of factors, but the small cap structure located at the 5′ end of the mRNA is one contributing factor. Cap protects mRNA and ensures efficient recruitment to the biosynthesis machinery. Furthermore, it allows for the easy introduction of various modifications that influence the activity of the entire mRNA. Among the many different cap analogues that have been reported, those modified at the N2 position of guanosine have been systematically developed. N2-modified caps in the form of nucleoside monophosphates or dinucleotides show favorable biological properties, as well as a high capacity to inhibit the translation process in the cell-free RRL system. Modified N2 dinucleotides are efficiently incorporated into the structure of the mRNA transcript, and in specific circumstances with the correct orientation, making them an interesting alternative for ARCA-type analogues. Moreover, mRNA transcripts containing cap structures modified within the exocyclic amino group show very high translational activity. Therefore, analogues modified at the N2 position may have future applications as therapeutics against various manifestations of cancer and as desirable tools in RNA engineering.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 6-7","pages":"511 - 519"},"PeriodicalIF":2.0,"publicationDate":"2023-09-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10132609","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Mammalian Nudt15 hydrolytic and binding activity on methylated guanosine mononucleotides 哺乳动物Nudt15对甲基化鸟苷单核苷酸的水解和结合活性。

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-08-29 DOI: 10.1007/s00249-023-01678-5

Maciej Lukaszewicz, Aleksandra Ferenc-Mrozek, Julia Kokosza, Anna Stefaniuk, Janusz Stepinski, Elzbieta Bojarska, Edward Darzynkiewicz

{"title":"Mammalian Nudt15 hydrolytic and binding activity on methylated guanosine mononucleotides","authors":"Maciej Lukaszewicz, Aleksandra Ferenc-Mrozek, Julia Kokosza, Anna Stefaniuk, Janusz Stepinski, Elzbieta Bojarska, Edward Darzynkiewicz","doi":"10.1007/s00249-023-01678-5","DOIUrl":"10.1007/s00249-023-01678-5","url":null,"abstract":"<div><p>The Nudt15 enzyme of the NUDIX protein family is the subject of extensive study due to its action on thiopurine drugs used in the treatment of cancer and inflammatory diseases. In addition to thiopurines, Nudt15 is enzymatically active in vitro on several nucleotide substrates. It has also been suggested that this enzyme may play a role in 5′RNA turnover by hydrolyzing m<sup>7</sup>GDP, a product of mRNA decapping. However, no detailed studies on this substrate with Nudt15 are available. Here, we analyzed the enzymatic activity of Nudt15 with m<sup>7</sup>GDP, its triphosphate form m<sup>7</sup>GTP, and the trimethylated counterparts (m<sub>3</sub><sup>2,2,7</sup>GDP and m<sub>3</sub><sup>2,2,7</sup>GTP). Kinetic data revealed a moderate activity of Nudt15 toward these methylated mononucleotides compared to the dGTP substrate. However m<sup>7</sup>GDP and m<sub>3</sub><sup>2,2,7</sup>GDP showed a distinct stabilization of Nudt15 upon ligand binding, in the same range as dGTP, and thus these two mononucleotides may be used as leading structures in the design of small molecule binders of Nudt15.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 6-7","pages":"487 - 495"},"PeriodicalIF":2.0,"publicationDate":"2023-08-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10109964","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Peptide nucleic acid conjugates and their antimicrobial applications—a mini-review 肽核酸偶联物及其抗菌应用——综述。

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-08-23 DOI: 10.1007/s00249-023-01673-w

Uladzislava Tsylents, Izabela Siekierska, Joanna Trylska

引用次数: 0

Structure and dynamics of pteridine reductase 1: the key phenomena relevant to enzyme function and drug design 蝶呤还原酶1的结构和动力学：与酶功能和药物设计相关的关键现象。

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-08-22 DOI: 10.1007/s00249-023-01677-6

Joanna Panecka-Hofman, Ina Poehner

{"title":"Structure and dynamics of pteridine reductase 1: the key phenomena relevant to enzyme function and drug design","authors":"Joanna Panecka-Hofman, Ina Poehner","doi":"10.1007/s00249-023-01677-6","DOIUrl":"10.1007/s00249-023-01677-6","url":null,"abstract":"<div><p>Pteridine reductase 1 (PTR1) is a folate and pterin pathway enzyme unique for pathogenic trypanosomatids. As a validated drug target, PTR1 has been the focus of recent research efforts aimed at finding more effective treatments against human parasitic diseases such as leishmaniasis or sleeping sickness. Previous PTR1-centered structural studies highlighted the enzyme characteristics, such as flexible regions around the active site, highly conserved structural waters, and species-specific differences in pocket properties and dynamics, which likely impacts the binding of natural substrates and inhibitors. Furthermore, several aspects of the PTR1 function, such as the substrate inhibition phenomenon and the level of ligand binding cooperativity in the enzyme homotetramer, likely related to the global enzyme dynamics, are poorly known at the molecular level. We postulate that future drug design efforts could greatly benefit from a better understanding of these phenomena through studying both the local and global PTR1 dynamics. This review highlights the key aspects of the PTR1 structure and dynamics relevant to structure-based drug design that could be effectively investigated by modeling approaches. Particular emphasis is given to the perspective of molecular dynamics, what has been accomplished in this area to date, and how modeling could impact the PTR1-targeted drug design in the future.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 6-7","pages":"521 - 532"},"PeriodicalIF":2.0,"publicationDate":"2023-08-22","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10407291","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Long-term memory in Staphylococcus aureus α-hemolysin ion channel kinetics 长期记忆对金黄色葡萄球菌α-溶血素离子通道动力学的影响。

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-08-05 DOI: 10.1007/s00249-023-01675-8

M. P. Silva, C. G. Rodrigues, D. C. Machado, R. A. Nogueira

{"title":"Long-term memory in Staphylococcus aureus α-hemolysin ion channel kinetics","authors":"M. P. Silva, C. G. Rodrigues, D. C. Machado, R. A. Nogueira","doi":"10.1007/s00249-023-01675-8","DOIUrl":"10.1007/s00249-023-01675-8","url":null,"abstract":"<div><p>The kinetics of an ion channel are classically understood as a random process. However, studies have shown that in complex ion channels, formed by multiple subunits, this process can be deterministic, presenting long-term memory. <i>Staphylococcus aureus</i> α-hemolysin (α-HL) is a toxin that acts as the major factor in <i>Staphylococcus aureus</i> virulence. α-HL is a water-soluble protein capable of forming ion channels into lipid bilayers, by insertion of an amphipathic β-barrel. Here, the α-HL was used as an experimental model to study memory in ion channel kinetics. We applied the approximate entropy (ApEn) approach to analyze randomness and the Detrended Fluctuation Analysis (DFA) to investigate the existence of long memory in α-HL channel kinetics. Single-channel currents were measured through experiments with α-HL channels incorporated in planar lipid bilayers. All experiments were carried out under the following conditions: 1 M NaCl solution, pH 4.5; transmembrane potential of + 40 mV and temperature 25 ± 1 °C. Single-channel currents were recorded in real-time in the memory of a microcomputer coupled to an A/D converter and a patch-clamp amplifier. The conductance value of the α-HL channels was 0.82 ± 0.0025 nS (<i>n</i> = 128). The DFA analysis showed that the kinetics of α-HL channels presents long-term memory (<span>({text{DFA}}_{{upalpha }})</span> = 0.63 ± 0.04). The ApEn outcomes showed low complexity to dwell times when open (ApEn<sub>o</sub> = 0.5514 ± 0.28) and closed (ApEn<sub>c</sub> = 0.1145 ± 0.08), corroborating the results of the DFA method.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 8","pages":"661 - 671"},"PeriodicalIF":2.0,"publicationDate":"2023-08-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9941465","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Proceedings of the 25th Analytical Ultracentrifugation Workshops and Symposium 第25届分析超离心研讨会论文集

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-08-01 DOI: 10.1007/s00249-023-01674-9

Borries Demeler, Robert Gilbert, Trushar R. Patel

{"title":"Proceedings of the 25th Analytical Ultracentrifugation Workshops and Symposium","authors":"Borries Demeler, Robert Gilbert, Trushar R. Patel","doi":"10.1007/s00249-023-01674-9","DOIUrl":"10.1007/s00249-023-01674-9","url":null,"abstract":"<div><p>The 25th International Analytical Ultracentrifugation (AUC) Workshops and Symposium (AUC2022) took place at the University of Lethbridge in Lethbridge, Canada, in July 2022. In total, 104 attendees (Attendance Profile: 104 attendees, 69 in-person, 35 remote. Brazil 1, Canada 24, China 1, Czech Republic 2, Finland 1, France 3, Germany 22, India 3, Italy 1, Japan 4, Spain 1, Switzerland 3, Taiwan 1, United Kingdom 5, United States 32) participated in the event and presented the latest advances in the field. While the primary focus of the conference was to showcase the applications of AUC in chemical, life sciences, and nanoparticle disciplines, several presentations also integrated complementary methods, such as isothermal titration calorimetry, microscale thermophoresis, light scattering (static and dynamic), small-angle X-ray scattering, X-ray crystallography, and cryo-electron microscopy. Additionally, the delegates gained valuable hands-on experience from 20 workshops covering a broad range of applications, experimental designs and systems, and the latest software innovations in solution biophysics. The AUC2022 special volume highlights the sustained innovation, utility and relevance of AUC and related solution biophysical methods across various disciplines, including biochemistry, structural biology, synthetic polymer chemistry, carbohydrate chemistry, protein and nucleic acid characterization, nano-science, and macromolecular interactions.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 4-5","pages":"195 - 201"},"PeriodicalIF":2.0,"publicationDate":"2023-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"4008302","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1

Tautomeric equilibrium and spectroscopic properties of 8-azaguanine revealed by quantum chemistry methods 用量子化学方法揭示了8-氮杂鸟嘌呤的互变异构平衡和光谱性质。

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-07-28 DOI: 10.1007/s00249-023-01672-x

Maciej Maciejczyk, Maciej Pyrka

{"title":"Tautomeric equilibrium and spectroscopic properties of 8-azaguanine revealed by quantum chemistry methods","authors":"Maciej Maciejczyk, Maciej Pyrka","doi":"10.1007/s00249-023-01672-x","DOIUrl":"10.1007/s00249-023-01672-x","url":null,"abstract":"<div><p>8-azaguanine is a triazolopyrimidine nucleobase analog possessing potent antibacterial and antitumor activities, and it has been implicated as a lead molecule in cancer and malaria therapy. Its intrinsic fluorescence properties can be utilized for monitoring its interactions with biological polymers like proteins or nucleic acids. In order to better understand these interactions, it is important to know the tautomeric equilibrium of this compound. In this work, the tautomeric equilibrium of all natural neutral and anionic compound forms (except highly improbable imino-enol tautomers) as well as their methyl derivatives and ribosides was revealed by quantum chemistry methods. It was shown that, as expected, tautomers protonated at positions 1 and 9 dominate neutral forms both in gas phase and in aqueous solution. 8-azaguanines methylated at any position of the triazole ring are protonated at position 1. The computed vertical absorption and emission energies are in very good agreement with the experimental data. They confirm the validity of the assumption that replacing the proton with the methyl group does not significantly change the positions of absorption and fluorescence peaks.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 6-7","pages":"545 - 557"},"PeriodicalIF":2.0,"publicationDate":"2023-07-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9885520","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 0

Suitability of double-stranded DNA as a molecular standard for the validation of analytical ultracentrifugation instruments 双链DNA作为分析超离心仪器验证的分子标准的适用性

IF 2 4区生物学

European Biophysics Journal Pub Date : 2023-07-27 DOI: 10.1007/s00249-023-01671-y

Maduni Ranasinghe, Jonathan M. Fogg, Daniel J. Catanese Jr., Lynn Zechiedrich, Borries Demeler

{"title":"Suitability of double-stranded DNA as a molecular standard for the validation of analytical ultracentrifugation instruments","authors":"Maduni Ranasinghe, Jonathan M. Fogg, Daniel J. Catanese Jr., Lynn Zechiedrich, Borries Demeler","doi":"10.1007/s00249-023-01671-y","DOIUrl":"10.1007/s00249-023-01671-y","url":null,"abstract":"<div><p>To address the current lack of validated molecular standards for analytical ultracentrifugation (AUC), we investigated the suitability of double-stranded DNA molecules. We compared the hydrodynamic properties of linear and circular DNA as a function of temperature. Negatively supercoiled, nicked, and linearized 333 and 339 bp minicircles were studied. We quantified the hydrodynamic properties of these DNAs at five different temperatures, ranging from 4 to 37 °C. To enhance the precision of our measurements, each sample was globally fitted over triplicates and five rotor speeds. The exceptional stability of DNA allowed each sample to be sedimented repeatedly over the course of several months without aggregation or degradation, and with excellent reproducibility. The sedimentation and diffusion coefficients of linearized and nicked minicircle DNA demonstrated a highly homogeneous sample, and increased with temperature, indicating a decrease in friction. The sedimentation of linearized DNA was the slowest; supercoiled DNA sedimented the fastest. With increasing temperature, the supercoiled samples shifted to slower sedimentation, but sedimented faster than nicked minicircles. These results suggest that negatively supercoiled DNA becomes less compact at higher temperatures. The supercoiled minicircles, as purified from bacteria, displayed heterogeneity. Therefore, supercoiled DNA isolated from bacteria is unsuitable as a molecular standard. Linear and nicked samples are well suited as a molecular standard for AUC and have exceptional colloidal stability in an AUC cell. Even after sixty experiments at different speeds and temperatures, measured over the course of 4 months, all topological states of DNA remained colloidal, and their concentrations remained essentially unchanged.</p></div>","PeriodicalId":548,"journal":{"name":"European Biophysics Journal","volume":"52 4-5","pages":"267 - 280"},"PeriodicalIF":2.0,"publicationDate":"2023-07-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"5043046","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

引用次数: 1