Journal of bioscience and bioengineering最新文献

Letter to the editor on "Evaluation methods for decellularized tissues: A focus on human amniotic membrane".

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-04-04 DOI: 10.1016/j.jbiosc.2025.02.010

Mathilde Fénelon, Nicolas Lheureux, Florelle Gindraux

引用次数: 0

Deep-learning-assisted medium optimization improves hyaluronic acid production by Streptococcus zooepidemicus.

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-04-03 DOI: 10.1016/j.jbiosc.2025.03.001

Kazuki Watanabe, Yoshizumi Kawai, Tomoko Kagenishi, Tai-Ying Chiou, Masaaki Konishi

{"title":"Deep-learning-assisted medium optimization improves hyaluronic acid production by Streptococcus zooepidemicus.","authors":"Kazuki Watanabe, Yoshizumi Kawai, Tomoko Kagenishi, Tai-Ying Chiou, Masaaki Konishi","doi":"10.1016/j.jbiosc.2025.03.001","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.03.001","url":null,"abstract":"To improve the efficiency of hyaluronic acid production by Streptococcus zooepidemicus, the growth medium was optimized with a pipeline involving a deep learning (DL) algorithm. To train the DL model, the initial training dataset (OA01-18) was designed with the L18 orthogonal array, and hyaluronic acid (HA) was produced in small-scale cultures in deepwell plates. The range of HA production was 0.09-1.39 g/L under these conditions. In searching for the optimal medium composition, 54 candidate optimized media (OM01-54) were proposed by the system. According to the confirming culture experiment, the best production of HA (1.66 g/L) was achieved with OM30. During confirmation in a stirred-tank reactor, the volumetric production of HA in OA30 was larger than that in the control medium. In fed batch culture, HA accumulated to 5.13 and 9.96 g/Linitial volume after 10 and 30 h in culture, respectively. To avoid the suppression of HA production by the high viscosity of the medium conferred by HA, repeated batch culture with OM30 was performed by replacing 90 % of the broth volume approximately every 6 h. As a result, 21.4 g of HA was produced in 46 h, and productivity reached 0.465 g/Linitial volume/h.","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143795553","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

New natural products from Aspergillus candidus NRRL 5214 and their glycosylation by Streptomyces chromofuscus ATCC 49982.

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-04-03 DOI: 10.1016/j.jbiosc.2025.03.002

Jie Ren, Kyle Jackson, Caleb Don Barton, Hassan Sher, Yu Huang, Jixun Zhan

{"title":"New natural products from Aspergillus candidus NRRL 5214 and their glycosylation by Streptomyces chromofuscus ATCC 49982.","authors":"Jie Ren, Kyle Jackson, Caleb Don Barton, Hassan Sher, Yu Huang, Jixun Zhan","doi":"10.1016/j.jbiosc.2025.03.002","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.03.002","url":null,"abstract":"Fungi represent a rich source of bioactive natural products. In this study, we present the isolation and identification of two new diphenyl ethers, named aspergilluscandidus A (1) and aspergilluscandidus B (2), along with a known compound terphenyllin (3), from the fungal strain Aspergillus candidus NRRL 5214. The chemical structures of compounds 1-3 were characterized through extensive 1D and 2D NMR analysis. Compounds 1 and 3 were subsequently biotransformed into two new glycosides, namely aspergilluscandidus C (4) and terphenyllin-4″-O-β-d-glucuronide (5) by the actinomycete strain Streptomyces chromofuscus ATCC 49982. The cytotoxicity assay revealed that the glycosylated products 4 and 5 exhibited significantly improved activity against the glioblastoma 33 cell line compared to their respective substrates, decreasing the IC50 from 8.15 ± 1.09 μM (1) to 5.41 ± 0.30 μM (4) and from 88.29 ± 10.54 μM (3) to 31.25 ± 4.20 μM (5), respectively. Our study emphasizes A. candidus NRRL 5214 as a promising source of new natural products and presents an effective strategy for modifying both diphenyl ether and p-terphenyl compounds using S. chromofuscus ATCC 49982 to enhance their cytotoxicity activity.","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-04-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143795560","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

Polyvinyl alcohol, N-acetylcysteine, and methyl-β-cyclodextrin exhibit albumin functions in natural killer cell culture.

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-03-24 DOI: 10.1016/j.jbiosc.2025.02.008

Ruisheng Xu, Guofeng Zhang, Huimin Huang, Yuanyuan Zhao, Wen-Song Tan, Haibo Cai

{"title":"Polyvinyl alcohol, N-acetylcysteine, and methyl-β-cyclodextrin exhibit albumin functions in natural killer cell culture.","authors":"Ruisheng Xu, Guofeng Zhang, Huimin Huang, Yuanyuan Zhao, Wen-Song Tan, Haibo Cai","doi":"10.1016/j.jbiosc.2025.02.008","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.008","url":null,"abstract":"Albumin is a crucial component of serum-free media, playing a significant role in ex vivo cell culture as a lipid carrier and antioxidant. However, purified albumin contains undefined substances, making it challenging to achieve clinical application standards for effector cell culture. This study used natural killer (NK)-92 cells as a model to investigate the effects of the albumin substitute replacing bovine serum albumin (BSA) on cell expansion and metabolism in an in-house-designed, chemically defined, serum-free medium. We selected polyvinyl alcohol (PVA), N-acetylcysteine (NAC), and methyl-β-cyclodextrin (M-β-CD) as an albumin substitute combination and optimized their concentrations by using response surface methodology. The optimized albumin substitute was named PVA-NAC-M-β-CD (PNM). After 8 days of culture, NK-92 cells cultured with the PNM exhibited phenotype and cytotoxic function comparable to cells cultured with different concentrations of BSA. The expansion fold was 89.22 ± 3.55, significantly higher than the 51.23 ± 6.57 observed in the 0.75 g/L BSA group (p < 0.05). Further verification of functions of PNM showed that intracellular fatty acid levels, cholesterol consumption rates, and the pSTAT5 level in the PNM group were significantly higher than those in the 0.75 g/L BSA group (p < 0.05). Reactive oxygen species levels remained controlled, and mitochondrial membrane potential was similar. These findings suggested that the PNM can effectively replace the functions of BSA as a fatty acid carrier, antioxidant, and, to some extent, a cholesterol carrier. This study provides insights for developing chemically defined media to prepare clinical-grade NK cells efficiently.","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-03-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143709905","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

Overproduction of secondary metabolites in Photorhabdus noenieputensis through rpoB mutations.

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-03-21 DOI: 10.1016/j.jbiosc.2025.02.004

Yu Imai

{"title":"Overproduction of secondary metabolites in Photorhabdus noenieputensis through rpoB mutations.","authors":"Yu Imai","doi":"10.1016/j.jbiosc.2025.02.004","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.004","url":null,"abstract":"Specific mutations of the rpoB gene, which encodes the β subunit of bacterial RNA polymerase, can enhance the production of secondary metabolites in bacteria such as actinomycetes. Entomopathogenic bacteria Photorhabdus and Xenorhabdus species produce a variety of secondary metabolites. Recently, these genera have attracted attention as a promising source for novel antibiotics. In this study, the effect of rpoB mutations on secondary metabolite production in Photorhabdus noenieputensis DSM 25462, a known producer of the antituberculosis antibiotic evybactin, was evaluated. Spontaneous rifampicin-resistant mutants, frequently carrying rpoB mutations, were generated by plating cells on agar medium containing four times the minimum inhibitory concentration (MIC) of rifampicin and evaluated their antibacterial production using Escherichia coli WO153 as a test strain. Among 190 spontaneous rifampicin-resistant mutants of P. noenieputensis, strain designated R191, which harbors the rpoB Q148K mutation (C442A), displayed higher antibacterial activity than that of the parental strain DSM 25462. The real-time quantitative RT-PCR analysis of 20 putative secondary metabolite biosynthetic gene clusters (BGCs) identified using antiSMASH revealed that seven of these BGCs were overexpressed in the strain R191. Furthermore, comparative high-pressure liquid chromatography (HPLC) analysis of the metabolite profile indicated that the strain R191 produced several compounds that were not detectable in the DSM 25462 culture. These findings suggest that the introduction of rpoB mutations into Photorhabdus strains is an effective strategy for enhancing secondary metabolite production and may lead to the discovery of novel antibiotics.","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143692225","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

Evaluation of antibody variants using a ribosome display and Brevibacillus choshinensis secretion system.

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-03-21 DOI: 10.1016/j.jbiosc.2025.02.009

Monami Kihara, Rio Okuda, Anri Okada, Teruyo Ojima-Kato, Hideo Nakano

{"title":"Evaluation of antibody variants using a ribosome display and Brevibacillus choshinensis secretion system.","authors":"Monami Kihara, Rio Okuda, Anri Okada, Teruyo Ojima-Kato, Hideo Nakano","doi":"10.1016/j.jbiosc.2025.02.009","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.009","url":null,"abstract":"In antibody engineering, the development of rapid and efficient strategies for improving affinity is highly necessary. In this study, we aimed to establish a method to efficiently enrich and analyze high-affinity antibody variants by combining protein synthesis using recombinant elements (PURE) ribosome display with next-generation sequencing (NGS) and Brevibacillus choshinensis secretion system using the NZ-1 antibody, which targets the PA tag peptide (GVAMPGAEDDVV) as a model antibody. From the mutated scFab library designed based on the structure, we performed a single-round of PURE ribosome display selection and analyzed the data by NGS to obtain high-affinity scFab candidates with high enrichment factor and high read counts. Subsequently, the most promising candidate was produced as a Fab in the B. choshinensis secretion system, and the purified Fab had an affinity (KD = 1.6 × 10-9 M) similar to the wild type. Overall, this study highlights the potential of the integrated PURE ribosome display with NGS analysis and the B. choshinensis secretion system for the rapid identification and analysis of high-affinity antibody variants.","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-03-21","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143692222","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

Application of silica monoliths for improved storage stability of metabolites in human plasma.

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-03-13 DOI: 10.1016/j.jbiosc.2025.02.007

Kazuhiro Kawamura, Eiichiro Fukusaki

{"title":"Application of silica monoliths for improved storage stability of metabolites in human plasma.","authors":"Kazuhiro Kawamura, Eiichiro Fukusaki","doi":"10.1016/j.jbiosc.2025.02.007","DOIUrl":"https://doi.org/10.1016/j.jbiosc.2025.02.007","url":null,"abstract":"In metabolomic studies, sample collection and analysis are typically performed at separate locations, necessitating the transport and storage of samples. However, sample transport and storage conditions are often constrained by the available facilities. Specifically, metabolite levels in biological and food samples can fluctuate due to the activity of endogenous enzymes, depending on the transport and storage conditions. Therefore, in this study, we aimed to achieve metabolite stabilization during storage by sampling human plasma on silica monoliths. Silica monoliths maintained the metabolite samples in a dry state, enabling their transport and storage at high temperatures. Plasmas stored at room temperature and refrigerated were measured using gas chromatography/mass spectrometry (GC/MS), and the fluctuations in metabolites between normal storage and storage on silica monoliths were compared. As a result, fluctuations in several metabolites such as glucose and amino acids were observed during normal storage. However, these were suppressed during storage on silica monoliths. Overall, our findings highlight the efficiency of silica monoliths for sample transport and storage at high temperatures.","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":" ","pages":""},"PeriodicalIF":2.3,"publicationDate":"2025-03-13","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143630501","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

Role of low-level alternating current and impedance for enhancing microalgae biomass and lipid production

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-03-10 DOI: 10.1016/j.jbiosc.2025.02.001

Carine Souza da Silva , Gustavo Miranda Pires Santos , Gabriele Rodrigues Conceição , Ian da Silva Andrade , Alana Nogueira Silva , Rodrigo Miranda Pires Santos , Paulo Fernando de Almeida , Fabio Alexandre Chinalia

{"title":"Role of low-level alternating current and impedance for enhancing microalgae biomass and lipid production","authors":"Carine Souza da Silva , Gustavo Miranda Pires Santos , Gabriele Rodrigues Conceição , Ian da Silva Andrade , Alana Nogueira Silva , Rodrigo Miranda Pires Santos , Paulo Fernando de Almeida , Fabio Alexandre Chinalia","doi":"10.1016/j.jbiosc.2025.02.001","DOIUrl":"10.1016/j.jbiosc.2025.02.001","url":null,"abstract":"<div><div>Microalgae hold significant potential for producing value-added bioproducts in pharmaceutical, cosmetic, food, and biofuel industries, with a global market value estimated at US$ 11.8 billion in 2023. Innovations in culturing systems, such as electric stimulation, aim to enhance growth performance, as it can improve cellular processes, including nutrient uptake and lipid accumulation. This study investigates the effect of low alternating electrical currents (μA) on the growth and lipid production of the halotolerant microalga Dunaliella salina across varying salt concentrations (3.5 % and 8.5 % Conway medium). Applying electric stimulation at 50, 750, and 990 μA for 30 min daily over 15 days resulted in significant enhancements, particularly at 3.5 % salinity, where lipid content increased by 144 %. The findings indicate that electrical stimulation notably reduced the lag phase and increased exponential growth rates, with superior growth coefficients correlating with higher medium impedance rather than direct current levels.</div></div>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":"139 5","pages":"Pages 369-376"},"PeriodicalIF":2.3,"publicationDate":"2025-03-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143604997","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

Development of an enzymatic cascade for semi de novo ATP production using thermophilic enzymes

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-03-09 DOI: 10.1016/j.jbiosc.2025.02.005

Takuma Suzuki , Suryatin Alim Gladwin , Kentaro Miyazaki , Hiroya Tomita , Kohsuke Honda

{"title":"Development of an enzymatic cascade for semi de novo ATP production using thermophilic enzymes","authors":"Takuma Suzuki , Suryatin Alim Gladwin , Kentaro Miyazaki , Hiroya Tomita , Kohsuke Honda","doi":"10.1016/j.jbiosc.2025.02.005","DOIUrl":"10.1016/j.jbiosc.2025.02.005","url":null,"abstract":"<div><div>Industrial production of ATP has mostly relied on extraction from living cells. Although microbial and enzymatic ATP production have also been developed, the former suffers from complexity in product separation, while the latter requires expensive substrates, making their practical use difficult. To tackle these problems, we newly developed an enzymatic cascade for ATP production, which does not use expensive substrates, by assembling 16 thermophilic enzymes prepared through a heat-purification from the crude extract of recombinant Escherichia coli. This cascade consists of two modules: an ATP regeneration module based on a non-oxidative glycolysis and an ADP supply module. The ATP regeneration module can provide the energy required for phosphorylation of AMP and ADP to ATP while simultaneously supplying ribose-5-phosphate, a building block of adenosine phosphates, from inexpensive starch and inorganic phosphate. Ribose-5-phosphate is then adenylated with exogenously supplied adenine in the ADP supply module and further phosphorylated to ATP. This ATP production cascade is not accompanied by CO2 emission and is expected to be a novel ATP manufacturing platform with less environmental impact. In the present study, ATP production with 100 % molar conversion yield was achieved from 1 mM adenine. However, increasing the initial adenine concentration resulted in lower yields. Enzyme characterization and docking simulations revealed that this decline was due to non-competitive inhibition of certain enzymes by ATP, which could potentially be mitigated through protein engineering.</div></div>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":"139 5","pages":"Pages 354-361"},"PeriodicalIF":2.3,"publicationDate":"2025-03-09","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143597122","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

Multi-gene metabolic engineering of Pichia pastoris to synthesize ectoine

IF 2.3 4区生物学

Journal of bioscience and bioengineering Pub Date : 2025-03-07 DOI: 10.1016/j.jbiosc.2025.02.006

Shuai Zhang , Bingjie Cheng , Qing Liao , Xuewu Huang , Mengjiao Mi , Ming Huang , Yue Wu , Shuyan Wu , Xiaoyuang Wang , Xiaoqing Hu

{"title":"Multi-gene metabolic engineering of Pichia pastoris to synthesize ectoine","authors":"Shuai Zhang , Bingjie Cheng , Qing Liao , Xuewu Huang , Mengjiao Mi , Ming Huang , Yue Wu , Shuyan Wu , Xiaoyuang Wang , Xiaoqing Hu","doi":"10.1016/j.jbiosc.2025.02.006","DOIUrl":"10.1016/j.jbiosc.2025.02.006","url":null,"abstract":"<div><div>As a promising osmolyte, ectoine has been widely applied in cosmetics, food, and pharmaceutical industries in recent years, therefore its biomanufacturer has attracted increasing interest. Ectoine-producing isolates were previously screened from halophilic microorganisms. After ectoine synthetase was identified, genetic engineering of Escherichia coli, Corynebacterium glutamicum, and Hansenula polymorpha were employed to produce ectoine. However, Pichia pastoris, another successful host capable of high-density cell culture, had not yet been exploited as an ectoine-synthesizing host. In this study, therefore, P. pastoris was employed for the first time to produce ectoine through multi-gene metabolic engineering. Firstly, Chromohalobacter salexigens HZS/E, a halophilic isolate producing 46.96 mg/mL ectoine, was identified, while ectABC encoding ectoine synthetase was cloned. Later, ectABC was introduced into P. pastoris GS115 under the control of two different promoters. The results showed that PGAP-based HZS02 accumulated 8.03 g/L, 12.62 % higher than 7.13 g/L produced by PAOX-based HZS01. Finally, to enhance the supply of the precursor l-aspartate-β-semialdehyde, three genes (aspC, aK, and asD) were individually and collectively overexpressed. The highest ectoine yield was achieved at 10.88 g/L by GS115/pGAPZ A-ectABC-aspC-aK-asD. This study demonstrated that P. pastoris was a highly effective host for ectoine biosynthesis.</div></div>","PeriodicalId":15199,"journal":{"name":"Journal of bioscience and bioengineering","volume":"139 5","pages":"Pages 347-353"},"PeriodicalIF":2.3,"publicationDate":"2025-03-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143585665","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