Biotechnology Letters最新文献_第2页

Enhancement hispolon production from Phellinus linteus via epigenetic-modified culture to inhibit human breast cancer cells.

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-26 DOI: 10.1007/s10529-025-03561-z

Phongsakorn Chueaphromsri, Phongsakorn Kunhorm, Nipha Chaicharoenaudomrung, Parinya Noisa

{"title":"Enhancement hispolon production from Phellinus linteus via epigenetic-modified culture to inhibit human breast cancer cells.","authors":"Phongsakorn Chueaphromsri, Phongsakorn Kunhorm, Nipha Chaicharoenaudomrung, Parinya Noisa","doi":"10.1007/s10529-025-03561-z","DOIUrl":"https://doi.org/10.1007/s10529-025-03561-z","url":null,"abstract":"Phellinus linteus (PL) is a medicinal fungus known for producing hispolon, a bioactive compound with antioxidant, anti-inflammatory, and anticancer properties. However, the natural scarcity of PL and the unsuccessful cultivation of its fruiting bodies have led to the exploration of alternative methods for enhancing its bioactive compound production. In this study, static fermentation was employed, and Valproic acid (VPA), a histone deacetylase inhibitor (HDACi), was added to the culture medium to induce epigenetic modifications and enhance hispolon production. After 30 days of fermentation, the hispolon concentration was analyzed using high-performance liquid chromatography (HPLC), mycelial dry weight was measured, and the expression of hispolon synthesis-related enzymes was quantified using quantitative PCR (qPCR). Additionally, the anticancer potential of the fermented media was assessed in human breast adenocarcinoma HTB-26 cells using assays for cytotoxicity, reactive oxygen species (ROS) formation, apoptosis, antioxidant activity, and autophagy markers. The results revealed that the addition of 400 µM VPA increased hispolon production by 120% and mycelial dry weight by 41%, likely due to enhanced transcriptional accessibility. Furthermore, the PL fermentation media significantly inhibited HTB-26 cell growth through the induction of ROS formation, autophagy, and apoptosis. These findings suggest that VPA-enhanced static fermentation of PL offers a promising strategy for optimizing hispolon production and developing effective anticancer therapeutics.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 2","pages":"29"},"PeriodicalIF":2.0,"publicationDate":"2025-02-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143514655","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

Genome-wide screening of mitogen-activated protein kinase (MAPK) gene family and expression profile under heavy metal stress in Solanum lycopersicum.

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-19 DOI: 10.1007/s10529-025-03567-7

Hasan Can, Ilhan Dogan, Mehmet Emin Uras, Fatih Tabanli, Asli Hocaoglu-Ozyigit, Ibrahim Ilker Ozyigit

{"title":"Genome-wide screening of mitogen-activated protein kinase (MAPK) gene family and expression profile under heavy metal stress in Solanum lycopersicum.","authors":"Hasan Can, Ilhan Dogan, Mehmet Emin Uras, Fatih Tabanli, Asli Hocaoglu-Ozyigit, Ibrahim Ilker Ozyigit","doi":"10.1007/s10529-025-03567-7","DOIUrl":"https://doi.org/10.1007/s10529-025-03567-7","url":null,"abstract":"MAPKs are one of the essential signal transduction complexes which are responsible for the perception of abiotic stress and for the producing of related transcripts for responding to abiotic stress. For systematical analyzes of the mitogen-activated protein (MAP) kinase gene families and their expression profiles in Solanum lycopersicum L. exposed to diverse heavy metal stresses, 17 SlMAPK genes were studied in comparison with their 159 orthologs from various plant species. The result of phylogenetic analysis revealed that SlMAPKs were divided into four different subgroups (A, B, C, and D) based on their nucleic acid and protein sequence alignments. SlMAPKs including A, B and C group had lower molecular weights and more hydrophobic structures than D group SlMAPKs, while possible extra phosphorylation sites predicted in D-group SLMAPKs. 24 cis regulating elements such as Box 4, TATA-box, ABRE and CAAT-box were detected in their upstream parts of DNA sequences. Also, it was determined that SlMAPKs show interactions with important proteins such as Guanine nucleotide-binding protein beta subunit, heterotrimeric G-protein, protein phosphatase 2C and HY5. The results from our gene expression analyzes, significant increases were found in the expressions of the selected SLMAPK gene with applications of a range of increasing heavy metal concentrations (e.g., by the application of the 400 mM Ni + Pb exposure, a 16-fold increase in the expression of SlMAPK gene was noted). Overall, SlMAPK genes and proteins known were re-evaluated, and the SlMAPKs interactions with some other important proteins were observed. Also, some predictions about the extra phosphorylation sites of SlMAPKs having effects on their functions were done. In addition, the expression levels of SlMAPK genes were monitored under different levels of heavy metal stress exposures.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 2","pages":"27"},"PeriodicalIF":2.0,"publicationDate":"2025-02-19","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143447746","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

O₂-microbubble of iron-porphyrin conjugated polyaspartamide for molecular ultrasound contrast effect.

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-19 DOI: 10.1007/s10529-025-03571-x

Yoon Na Cho, Jun Woo Lim, Seung Joo Oh, Sa Ra Han, Sungwoo Cho, Jimin Jeong, Byoung Hee Han, Jae Hyun Jeong

引用次数: 0

Evaluation of secretory signal peptides for heterologous protein secretion in Cyanobacterium aponinum PCC10605.

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-18 DOI: 10.1007/s10529-025-03569-5

Rajesh Nandru, Bhaskar Bhadra, Nilanjan Roy, Anshul Nigam, Penna Suprasanna

{"title":"Evaluation of secretory signal peptides for heterologous protein secretion in Cyanobacterium aponinum PCC10605.","authors":"Rajesh Nandru, Bhaskar Bhadra, Nilanjan Roy, Anshul Nigam, Penna Suprasanna","doi":"10.1007/s10529-025-03569-5","DOIUrl":"https://doi.org/10.1007/s10529-025-03569-5","url":null,"abstract":"Biomanufacturing of recombinant proteins in the microalgae has become an important field of research owing to sustainability, scalability, safety, and metabolic diversity of the microalgal system. Recovery of the recombinant protein from the host system needs to be devised and established, since the conventional downstream process for recombinant protein extraction is associated with high costs and resources. In a previous study, we have reported two putative signal peptides of C. aponinum using in silico approach. Herein, we evaluated the two secretory signal peptides for heterologous protein secretion in C. aponinum PCC10605. The green fluorescent protein was used as secretory protein and as a reporter. Signal peptides, thermitase and porin, fused with GFP were transformed in to C. aponinum for studying the expression and secretion. Following the antibiotic screening and GFP fluorescence analysis, transformants secreting GFP in the supernatant were validated by using western blotting. The results showed that fluorescence, as measured by FACS analysis and TECAN reader, varied among the two signal peptides and, higher fluorescence was recorded in the 'thermitase SP secreted GFP' supernatant. The thermitase signal peptide may offer as a new gateway for recombinant protein production and secretion in C. aponinum.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 2","pages":"25"},"PeriodicalIF":2.0,"publicationDate":"2025-02-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143439767","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

In-silico evaluation of potential plant-based tyrosinase inhibitors for cosmetic and pharmaceutical applications.

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-18 DOI: 10.1007/s10529-025-03570-y

Jyoti Srivastava, Sukhendra Singh, Rupika Sinha

{"title":"In-silico evaluation of potential plant-based tyrosinase inhibitors for cosmetic and pharmaceutical applications.","authors":"Jyoti Srivastava, Sukhendra Singh, Rupika Sinha","doi":"10.1007/s10529-025-03570-y","DOIUrl":"https://doi.org/10.1007/s10529-025-03570-y","url":null,"abstract":"Tyrosinase is involved in a critical step of melanin synthesis; therefore, tyrosinase inhibitors are gaining more importance in the medicinal and cosmetic industry for the treatment of different pigmentary disorders. In the last decades, mushroom tyrosinase was used as a standard enzyme for the identification and advancement of most tyrosinase inhibitors. Due to differences in structure and substrate specificity between mushroom and human tyrosinase, there is a need for a more specific study with human tyrosinase. Additionally, the tyrosinase inhibitors which are currently in use have various side effects, therefore, safer inhibitors from natural sources are required. Different tyrosinase inhibitors from natural sources (aloesin, norartocarpetin, hesperetin, morin and taxifolin) were evaluated for an effective eco-friendly whitening agent using different bioinformatics tools. To check the efficacy and safety of the selected compounds ADME analysis was performed which showed that all the selected compounds fulfilled most of the parameters of general drug discovery. Docking of selected ligands was performed against the predicted structure of human tyrosinase; and the binding affinity (in kcal/mol) of kojic acid, aloesin, norartocarpetin, hesperetin, morin and taxifolin were obtained to be - 5.6, - 7.2, - 7.6, - 7.5, - 7.3 and - 7.2 respectively. Among all the selected ligands, norartocarpetin had the lowest binding affinity, i.e., - 7.6 kcal/mol, which showed that norartocarpetin could be used as a potent tyrosinase inhibitor. This bioactive compound is widely distributed in Moraceae plants and therefore, poses as a natural solution to various melanin-based dermatological issues and it can have a potential application in pharmaceuticals and cosmetic industries for the treatment of pigmentary disorders.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 2","pages":"26"},"PeriodicalIF":2.0,"publicationDate":"2025-02-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143439768","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

Mycoremediation of Sulfamethoxazole and metabolic pathway by Aspergillus tubingensis strain.

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-05 DOI: 10.1007/s10529-025-03568-6

Raveena Ann Alex, Jayanthi Abraham

引用次数: 0

Bacillus velezensis CE 100 controls anthracnose disease in walnut trees (Juglans regia L.) by inhibiting Colletotrichum gloeosporioides and eliciting induced systemic resistance. Velezensis CE 100 杆菌通过抑制球孢子菌（Colletotrichum gloeosporioides）和诱导系统抗性来控制核桃树（Juglans regia L.）的炭疽病。

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-05 DOI: 10.1007/s10529-025-03560-0

Vantha Choub, Sang-Jae Won, Jae-Hyun Moon, Su-In Choi, Henry B Ajuna, Young Sang Ahn

{"title":"Bacillus velezensis CE 100 controls anthracnose disease in walnut trees (Juglans regia L.) by inhibiting Colletotrichum gloeosporioides and eliciting induced systemic resistance.","authors":"Vantha Choub, Sang-Jae Won, Jae-Hyun Moon, Su-In Choi, Henry B Ajuna, Young Sang Ahn","doi":"10.1007/s10529-025-03560-0","DOIUrl":"10.1007/s10529-025-03560-0","url":null,"abstract":"Biological control of plant diseases is recognized as an effective and environmental friendly alternative to chemical fungicides. We demonstrated the dual biocontrol strategy of Bacillus velezensis CE 100 through the hydrolytic activity of chitinase and β-1,3-glucanase and the elicitation of induced systemic resistance (ISR) against Colletotrichum gloeosporioides that causes anthracnose disease in walnut trees. B. velezensis CE 100 produced a maximum of 62.1 units mL-1 (132.9 units mL-1) chitinase and 5.2 units mL-1 (9.4 units mL-1) β-1,3-glucanase enzymes in the broth culture (crude enzyme fraction), and inhibited spore germination and mycelial growth of C. gloeosporioides by 81.6% and 22.6%, respectively, at 100 µl mL-1 of crude enzyme fraction. The inoculation of B. velezensis CE 100 induced the production of pathogenesis-related (PR) chitinase in walnuts by 2.1-fold, and to a lesser extent PR β-1,3-glucanase, and reduced anthracnose disease severity by 3.0-fold compared to the control group. The bacterium produced a maximum of 11.4 µg mL-1 indole-3-acetic acid (IAA) and improved the chlorophyll content, shoot length, and root collar diameter of walnut trees compared to the fungicide treatment and control groups. B. velezensis CE 100 demonstrated the prospect of controlling walnut anthracnose by direct antagonism and ISR against C. gloeosporioides, while simultaneously enhancing walnut growth through IAA production.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 1","pages":"20"},"PeriodicalIF":2.0,"publicationDate":"2025-02-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143188030","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

Enhancement of reduction of biodesulfurization end products by Paenibacillus strains. 增强Paenibacillus菌株对生物硫化最终产物的还原作用。

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-05 DOI: 10.1007/s10529-025-03563-x

Taner Sar, Murat Ozturk, Benjamin C Stark, Meltem Yesilcimen Akbas

{"title":"Enhancement of reduction of biodesulfurization end products by Paenibacillus strains.","authors":"Taner Sar, Murat Ozturk, Benjamin C Stark, Meltem Yesilcimen Akbas","doi":"10.1007/s10529-025-03563-x","DOIUrl":"10.1007/s10529-025-03563-x","url":null,"abstract":"The work reported here aimed to enhance the reduction of the downstream intermediates 2-hydroxybiphenyl (2-HBP) and 2-(2'hydroxyphenyl ethan-1-al) produced by biodesulfurization of DBT (dibenzothiophene) and BT (benzothiophene) using Paenibacillus strains (32O-W and 32O-Y). Salicylaldehyde was used as a surrogate for 2-(2'hydroxybiphenyl ethan-1-al), as the two compounds are structurally very similar while the latter is not commercially available. Five strategies were tested using growth in media containing either 2-HBP or salicylaldehyde: use of single strain cultures, co-culturing, genetic engineering to express Vitreoscilla hemoglobin (VHb), cell immobilization, and nanoparticle coating of cells. Cell growth and reduction of 2-HBP and salicylaldehyde were measured during 96 h of culturing. Regarding 2-HBP reduction, 32O-Y was generally better than 32O-W (about 50% for free cells, and as much as 16% for immobilized cells, and 24% for coated cells); co-culturing did not provide any consistent advantage, while VHb expression increased utilization only for 32O-W (by about 50%). Immobilization and coating resulted in large improvements for both strains (as much as 3700%). Free, immobilized, and coated cells of 32O-Y all removed salicylaldehyde, while only immobilized 32O-W cells were able to do so. For 32O-Y, co-culturing and coating, but not VHb expression, resulted in improvements in salicylaldehyde reduction (of up to 31%). Thus, alginate immobilization or nanoparticle coating of bacterial cells may be useful approaches for enhancing the reduction of DBT or BT biodesulfurization end products, and thus the overall biodesulfurization process for petroleum and petroleum products.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 1","pages":"21"},"PeriodicalIF":2.0,"publicationDate":"2025-02-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143188037","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

Comparative study of nanohydroxyapatite-emdogain effects on apical papilla stem cell survival and differentiation.

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-05 DOI: 10.1007/s10529-024-03557-1

Elham Khoshbin, Hamed Karkehabadi, Razieh Salehi, Abbas Farmany, Rezvan Najafi, Roshanak Abbasi

{"title":"Comparative study of nanohydroxyapatite-emdogain effects on apical papilla stem cell survival and differentiation.","authors":"Elham Khoshbin, Hamed Karkehabadi, Razieh Salehi, Abbas Farmany, Rezvan Najafi, Roshanak Abbasi","doi":"10.1007/s10529-024-03557-1","DOIUrl":"10.1007/s10529-024-03557-1","url":null,"abstract":"Background: The study was designed to explore the enhanced impact of nano-hydroxyapatite and emdogain on the survival and osteogenic/odontogenic differentiation of human stem cells isolated from the apical papilla (hSCAPs).Materials and methods: In this in vitro trial, hSCAPS obtained from intact impacted immature third molars were confirmed to have characteristic cell surface markers, then exposed to nanohydroxyapatite, emdogain, and nanohydroxyapatite coated with emdogain for durations of 1-3 days. The survival of apical papilla stem cells was measured using a methyl thiazolyl tetrazolium assay. The quantitative reverse transcription polymerase chain reaction, alkaline phosphatase activity (ALP) and Alizarin red staining were used to evaluate osteogenic-odontogenic differentiation. Analysis of the data was done using one-way ANOVA, t-test, and Mann-Whitney test (α = 0.05).Results: At 1-3 days, emdogain exhibited no significant impact on the survival of human stem cells from the apical papilla. In contrast, nanohydroxyapatite (α > 0.05) and nanohydroxyapatite coated with emdogain demonstrated a notable reduction in cell survival compared to the control group (α < 0.05). The expression of dentin sialophosphoprotein, dentin matrix protein 1, and bone sialoprotein genes demonstrated a notable increase in the group treated with nanohydroxyapatite coated with emdogain compared to the other groups (α < 0.05), and furthermore, this group exhibited more pronounced mineralized nodules than the other experimental groups.Conclusion: In contrast to nanohydroxyapatite, Emdogain did not demonstrate a detrimental effect on the survival of hSCAPs. Nanohydroxyapatite, emdogain, and nanohydroxyapatite coated with emdogain increased osteogenic/odontogenic differentiation of hSCAPs.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 1","pages":"24"},"PeriodicalIF":2.0,"publicationDate":"2025-02-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143188032","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

Determining the function of ripening associated genes and biochemical changes during tomato (Solanum lycopersicum L.) fruit maturation.

IF 2 4区生物学

Biotechnology Letters Pub Date : 2025-02-05 DOI: 10.1007/s10529-025-03565-9

Darshan Dorairaj, Shivangi Sharma, Kiran Suresh Mawale, Bijesh Puthusseri, Giridhar Parvatam, Nandini Prasad Shetty

{"title":"Determining the function of ripening associated genes and biochemical changes during tomato (Solanum lycopersicum L.) fruit maturation.","authors":"Darshan Dorairaj, Shivangi Sharma, Kiran Suresh Mawale, Bijesh Puthusseri, Giridhar Parvatam, Nandini Prasad Shetty","doi":"10.1007/s10529-025-03565-9","DOIUrl":"10.1007/s10529-025-03565-9","url":null,"abstract":"This article examines biochemical alterations and gene expression changes during tomato fruit physiology. The chroma index increases from mature green (41.27) to red ripe (48.36) stages, and the texture softens from mature green (43.56 N) to red ripe (24.75 N). Reducing sugar and total carotenoid levels rise at the red ripe stage. Free radical content was elevated in the early stages (7 nM) of ripening and declined at the later stages (4 nM). The specific activity of α-mannosidase and β-N-acetyl hexosaminidase was high at the breaker (0.077 & 0.075 U/mg, respectively) stages, while polygalacturonase activity was high at red ripe (1.173 U/mg) stage. qPCR experiments revealed that the α-mannosidase was upregulated during the breaker (1.2 fold) stages of tomato ripening, the β-N-acetyl Hexosaminidase was upregulated throughout the breaker (2 fold), and pink (1.2 fold) stages of tomato ripening, and the β-xylosidase was upregulated significantly during the breaker stage (3.9 fold) of tomato ripening. The current findings revealed that the α-Mannosidase (0.77), β-N-acetylhexosaminidase (0.99), xylosidase (0.85), ethylene-responsive factors (0.86), aminocylco propane carboxylic oxidase (0.90), and pectin methylesterase (0.83), were significantly associated with textural softening. Polygalacturonase (0.75) positively correlated to reducing sugar formation, aminocylco propane carboxylic synthase 4 (0.96) expression correlates with chroma changes during tomato fruit ripening. These correlations illustrate the complex interplay between gene expression and the physical and biochemical changes occurring during tomato fruit ripening.","PeriodicalId":8929,"journal":{"name":"Biotechnology Letters","volume":"47 1","pages":"22"},"PeriodicalIF":2.0,"publicationDate":"2025-02-05","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143188034","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