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MetA is a ''thermal fuse'' that inhibits growth and protects Escherichia coli at elevated temperatures. MetA是一种“热保险丝”,在高温下抑制生长并保护大肠杆菌。
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2025-01-18 DOI: 10.1016/j.celrep.2025.115257
Severin J Schink, Zara Gough, Elena Biselli, Mariel Garcia Huiman, Yu-Fang Chang, Markus Basan, Ulrich Gerland
{"title":"MetA is a ''thermal fuse'' that inhibits growth and protects Escherichia coli at elevated temperatures.","authors":"Severin J Schink, Zara Gough, Elena Biselli, Mariel Garcia Huiman, Yu-Fang Chang, Markus Basan, Ulrich Gerland","doi":"10.1016/j.celrep.2025.115257","DOIUrl":"10.1016/j.celrep.2025.115257","url":null,"abstract":"","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":"44 1","pages":"115257"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143000782","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
CPSF1 inhibition promotes widespread use of intergenic polyadenylation sites and impairs glycolysis in prostate cancer cells.
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2025-01-22 DOI: 10.1016/j.celrep.2024.115211
Kiel T Tietz, Braedan M McCluskey, Conor R Miller, Yingming Li, Sarah A Munro, Scott M Dehm
{"title":"CPSF1 inhibition promotes widespread use of intergenic polyadenylation sites and impairs glycolysis in prostate cancer cells.","authors":"Kiel T Tietz, Braedan M McCluskey, Conor R Miller, Yingming Li, Sarah A Munro, Scott M Dehm","doi":"10.1016/j.celrep.2024.115211","DOIUrl":"10.1016/j.celrep.2024.115211","url":null,"abstract":"<p><p>Localized prostate cancer can be cured by radiation or surgery, but advanced prostate cancer continues to be a clinical challenge. Altered alternative polyadenylation occurs in numerous cancers and can downregulate tumor-suppressor genes and upregulate oncogenes. We found that the cleavage and polyadenylation specificity factor (CPSF) complex factor CPSF1 is upregulated in patients with advanced prostate cancer, with high CPSF1 expression correlating with worse progression-free survival. Knockdown of CPSF1 selectively inhibited the growth of prostate cancer cells and reduced glycolytic output. Evaluating the changes in global poly(A) site usage in prostate cancer cells following CPSF1 knockdown revealed widespread usage of intergenic poly(A) sites distal to annotated 3' UTRs, which lengthened 3' UTRs and decreased levels of thousands of mRNAs, including key glycolysis genes. These findings uncover a role for CPSF1 in the suppression of intergenic poly(A) sites in prostate cancer and nominate CPSF1 as a therapeutic target in advanced prostate cancer.</p>","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":"44 1","pages":"115211"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143028046","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
A nonsecretory antimicrobial peptide mediates inflammatory organ damage in Drosophila renal tubules. 一种非分泌性抗菌肽介导果蝇肾小管的炎症性器官损伤。
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2024-12-23 DOI: 10.1016/j.celrep.2024.115082
Ayano Oi, Natsuki Shinoda, Shun Nagashima, Masayuki Miura, Fumiaki Obata
{"title":"A nonsecretory antimicrobial peptide mediates inflammatory organ damage in Drosophila renal tubules.","authors":"Ayano Oi, Natsuki Shinoda, Shun Nagashima, Masayuki Miura, Fumiaki Obata","doi":"10.1016/j.celrep.2024.115082","DOIUrl":"10.1016/j.celrep.2024.115082","url":null,"abstract":"<p><p>An excessive immune response damages organs, yet its molecular mechanism is incompletely understood. Here, we screened a factor mediating organ damage upon genetic activation of the innate immune pathway using Drosophila renal tubules. We found that an antimicrobial peptide, Attacin-D (AttD), causes organ damage upon immune deficiency (Imd) pathway activation in the Malpighian tubules. Loss of AttD function suppresses most of the pathological phenotypes induced by Imd activation, such as cell death, bloating of the whole animal, and mortality, without compromising the immune activation. AttD is required for the immune-induced damage specifically in the Malpighian tubules and not the midgut. Unlike other antimicrobial peptides, AttD lacks a signal peptide and stays inside tubular cells, potentially damaging the tubular cells via aggregation and oligomerization. Suppression of AttD almost completely attenuates the pathology caused by a gut-tumor-induced immune activation. Our study elucidates the mechanistic effector of immune-induced organ damage.</p>","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":" ","pages":"115082"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142884977","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
Anti-herpetic tau preserves neurons via the cGAS-STING-TBK1 pathway in Alzheimer's disease. 抗疱疹tau通过cGAS-STING-TBK1通路在阿尔茨海默病中保存神经元。
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2025-01-02 DOI: 10.1016/j.celrep.2024.115109
Vanesa R Hyde, Chaoming Zhou, Juan R Fernandez, Krishnashis Chatterjee, Pururav Ramakrishna, Amanda Lin, Gregory W Fisher, Orhan Tunç Çeliker, Jill Caldwell, Omer Bender, Peter Joseph Sauer, Jose Lugo-Martinez, Daniel Z Bar, Leonardo D'Aiuto, Or A Shemesh
{"title":"Anti-herpetic tau preserves neurons via the cGAS-STING-TBK1 pathway in Alzheimer's disease.","authors":"Vanesa R Hyde, Chaoming Zhou, Juan R Fernandez, Krishnashis Chatterjee, Pururav Ramakrishna, Amanda Lin, Gregory W Fisher, Orhan Tunç Çeliker, Jill Caldwell, Omer Bender, Peter Joseph Sauer, Jose Lugo-Martinez, Daniel Z Bar, Leonardo D'Aiuto, Or A Shemesh","doi":"10.1016/j.celrep.2024.115109","DOIUrl":"10.1016/j.celrep.2024.115109","url":null,"abstract":"<p><p>Alzheimer's disease (AD) diagnosis relies on the presence of extracellular β-amyloid (Aβ) and intracellular hyperphosphorylated tau (p-tau). Emerging evidence suggests a potential link between AD pathologies and infectious agents, with herpes simplex virus 1 (HSV-1) being a leading candidate. Our investigation, using metagenomics, mass spectrometry, western blotting, and decrowding expansion pathology, detects HSV-1-associated proteins in human brain samples. Expression of the herpesvirus protein ICP27 increases with AD severity and strongly colocalizes with p-tau but not with Aβ. Modeling in human brain organoids shows that HSV-1 infection elevates tau phosphorylation. Notably, p-tau reduces ICP27 expression and markedly decreases post-infection neuronal death from 64% to 7%. This modeling prompts investigation into the cGAS-STING-TBK1 pathway products, nuclear factor (NF)-κB and IRF-3, which colocalizes with ICP27 and p-tau in AD. Furthermore, experimental activation of STING enhances tau phosphorylation, while TBK1 inhibition prevents it. Together, these findings suggest that tau phosphorylation acts as an innate immune response in AD, driven by cGAS-STING.</p>","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":" ","pages":"115109"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142926818","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
The glial UDP-glycosyltransferase Ugt35b regulates longevity by maintaining lipid homeostasis in Drosophila. 胶质udp -糖基转移酶Ugt35b通过维持果蝇脂质稳态来调节寿命。
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2024-12-24 DOI: 10.1016/j.celrep.2024.115099
Lihong Sheng, Jianpeng Gao, Qingyuan Wei, Ye Gong, Zhi-Xiang Xu
{"title":"The glial UDP-glycosyltransferase Ugt35b regulates longevity by maintaining lipid homeostasis in Drosophila.","authors":"Lihong Sheng, Jianpeng Gao, Qingyuan Wei, Ye Gong, Zhi-Xiang Xu","doi":"10.1016/j.celrep.2024.115099","DOIUrl":"10.1016/j.celrep.2024.115099","url":null,"abstract":"<p><p>Lipid droplets (LDs) are dynamic organelles essential for lipid storage and organismal survival. Studies have highlighted the importance of glial function in brain LD formation during aging; however, the genes and mechanisms involved remain elusive. Here, we found that Ugt35b, a member of the uridine diphosphate (UDP)-glycosyltransferases that catalyze the transfer of glycosyl groups to acceptors, is highly expressed in glia and crucial for Drosophila lifespan. By integrating multiomics data, we demonstrated that glial Ugt35b plays key roles in regulating glycerolipid and glycerophospholipid metabolism in the brain. Notably, we found that Ugt35b and Lsd-2 are co-expressed in glia and confirmed their protein interaction in vivo. Knockdown of Ugt35b significantly reduced LD formation by downregulating Lsd-2 expression, while overexpression of Lsd-2 partially rescued the shortened lifespan in glial Ugt35b RNAi flies. Our findings reveal the crucial role of glial Ugt35b in regulating LD formation to maintain brain lipid homeostasis and support Drosophila lifespan.</p>","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":"44 1","pages":"115099"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142892124","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
Pathogenic role for CD101-negative neutrophils in the type I interferon-mediated immunopathogenesis of tuberculosis. cd101阴性中性粒细胞在I型干扰素介导的结核病免疫发病机制中的致病作用。
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2024-12-17 DOI: 10.1016/j.celrep.2024.115072
Mohd Saqib, Shreya Das, Tanvir N Nafiz, Elizabeth McDonough, Poornima Sankar, Lokesh K Mishra, Ximeng Zhang, Yi Cai, Selvakumar Subbian, Bibhuti B Mishra
{"title":"Pathogenic role for CD101-negative neutrophils in the type I interferon-mediated immunopathogenesis of tuberculosis.","authors":"Mohd Saqib, Shreya Das, Tanvir N Nafiz, Elizabeth McDonough, Poornima Sankar, Lokesh K Mishra, Ximeng Zhang, Yi Cai, Selvakumar Subbian, Bibhuti B Mishra","doi":"10.1016/j.celrep.2024.115072","DOIUrl":"10.1016/j.celrep.2024.115072","url":null,"abstract":"<p><p>Neutrophils are vital for immunity against Mycobacterium tuberculosis (Mtb), the causative agent of tuberculosis (TB), yet their heterogeneous nature suggests a complex role in TB pathogenesis. Here, we identify two distinct neutrophil populations based on CD101 expression, highlighting their divergent roles in TB. CD101-negative (CD101<sup>-ve</sup>) neutrophils, which resemble immature, pro-inflammatory granulocytes, exhibit reduced Mtb phagocytosis compared to their mature, CD101-positive (CD101<sup>+ve</sup>) counterparts. Our findings reveal that type I interferons (IFN-Is) suppress neutrophil Mtb uptake and drive the recruitment of CD101<sup>-ve</sup> neutrophils to the lungs. Infiltration of these cells promotes Mtb extracellular persistence, exacerbates epithelial damage, and impairs surfactant production. Furthermore, we demonstrate that granulocyte colony-stimulating factor (G-CSF) and chemokine receptor CXCR2 are essential for the pulmonary accumulation of CD101<sup>-ve</sup> neutrophils. Our study uncovers a pathogenic role for CD101<sup>-ve</sup> neutrophils in TB and highlights the IFN-I-dependent recruitment of this functionally compromised immature neutrophil as a driver of TB immunopathogenesis.</p>","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":"44 1","pages":"115072"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142853146","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
Metabolic transcriptomics dictate responses of cone photoreceptors to retinitis pigmentosa. 代谢转录组学指示视锥光感受器对视网膜色素变性的反应。
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2024-12-21 DOI: 10.1016/j.celrep.2024.115160
Sang Joon Lee, Douglas Emery, Eric Vukmanic, Yekai Wang, Xiaoqin Lu, Wei Wang, Enzo Fortuny, Robert James, Henry J Kaplan, Yongqing Liu, Jianhai Du, Douglas C Dean
{"title":"Metabolic transcriptomics dictate responses of cone photoreceptors to retinitis pigmentosa.","authors":"Sang Joon Lee, Douglas Emery, Eric Vukmanic, Yekai Wang, Xiaoqin Lu, Wei Wang, Enzo Fortuny, Robert James, Henry J Kaplan, Yongqing Liu, Jianhai Du, Douglas C Dean","doi":"10.1016/j.celrep.2024.115160","DOIUrl":"10.1016/j.celrep.2024.115160","url":null,"abstract":"","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":"44 1","pages":"115160"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142876336","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
SLC7A5 is required for cancer cell growth under arginine-limited conditions. SLC7A5是癌细胞在精氨酸限制条件下生长所必需的。
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2025-01-03 DOI: 10.1016/j.celrep.2024.115130
Kyle N Dunlap, Austin Bender, Alexis Bowles, Alex J Bott, Joshua Tay, Allie H Grossmann, Jared Rutter, Gregory S Ducker
{"title":"SLC7A5 is required for cancer cell growth under arginine-limited conditions.","authors":"Kyle N Dunlap, Austin Bender, Alexis Bowles, Alex J Bott, Joshua Tay, Allie H Grossmann, Jared Rutter, Gregory S Ducker","doi":"10.1016/j.celrep.2024.115130","DOIUrl":"10.1016/j.celrep.2024.115130","url":null,"abstract":"<p><p>Tumor cells must optimize metabolite acquisition between synthesis and uptake from a microenvironment characterized by hypoxia, lactate accumulation, and depletion of many amino acids, including arginine. We performed a metabolism-focused functional screen using CRISPR-Cas9 to identify pathways and factors that enable tumor growth in an arginine-depleted environment. Our screen identified the SLC-family transporter SLC7A5 as required for growth, and we hypothesized that this protein functions as a high-affinity citrulline transporter. Using isotope tracing experiments, we show that citrulline uptake and metabolism into arginine are dependent upon expression of SLC7A5. Pharmacological inhibition of SLC7A5 blocks growth under low-arginine conditions across a diverse group of cancer cell lines. Loss of SLC7A5 reduces tumor growth and citrulline import in a mouse tumor model. We identify a conditionally essential role for SLC7A5 in arginine metabolism, and we propose that SLC7A5-targeting therapeutic strategies in cancer may be effective in the context of arginine limitation.</p>","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":"44 1","pages":"115130"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142930800","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
IL-21 shapes the B cell response in a context-dependent manner. IL-21以上下文依赖的方式塑造B细胞反应。
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2025-01-09 DOI: 10.1016/j.celrep.2024.115190
Youngjun Kim, Francesca Manara, Simon Grassmann, Kalina T Belcheva, Kanelly Reyes, Hyunu Kim, Stephanie Downs-Canner, William T Yewdell, Joseph C Sun, Jayanta Chaudhuri
{"title":"IL-21 shapes the B cell response in a context-dependent manner.","authors":"Youngjun Kim, Francesca Manara, Simon Grassmann, Kalina T Belcheva, Kanelly Reyes, Hyunu Kim, Stephanie Downs-Canner, William T Yewdell, Joseph C Sun, Jayanta Chaudhuri","doi":"10.1016/j.celrep.2024.115190","DOIUrl":"10.1016/j.celrep.2024.115190","url":null,"abstract":"<p><p>The T-cell-derived cytokine IL-21 is crucial for germinal center (GC) responses, but its precise role in B cell function has remained elusive. Using IL-21 receptor (Il21r) conditional knockout mice and ex vivo culture systems, we demonstrate that IL-21 has dual effects on B cells. While IL-21 induced apoptosis in a STAT3-dependent manner in naive B cells, it promoted the robust proliferation of pre-activated B cells, particularly IgG1<sup>+</sup> B cells. In vivo, B-cell-specific Il21r deletion impaired IgG1 responses post-immunization and disrupted progression from pre-GC to GC states. Although Il21r deficiency did not affect the proportion of IgG1<sup>+</sup> cells among GC B cells, it greatly diminished the proportion of IgG1<sup>+</sup> cells among the plasmablast/plasma cell population. Collectively, our findings suggest that IL-21 serves as a critical regulator of B cell fates, influencing B cell apoptosis and proliferation in a context-dependent manner.</p>","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":"44 1","pages":"115190"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142961982","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
Tumor-induced metabolic immunosuppression: Mechanisms and therapeutic targets. 肿瘤诱导的代谢免疫抑制:机制和治疗靶点。
IF 7.5 1区 生物学
Cell reports Pub Date : 2025-01-28 Epub Date: 2025-01-10 DOI: 10.1016/j.celrep.2024.115206
Jean-Ehrland Ricci
{"title":"Tumor-induced metabolic immunosuppression: Mechanisms and therapeutic targets.","authors":"Jean-Ehrland Ricci","doi":"10.1016/j.celrep.2024.115206","DOIUrl":"10.1016/j.celrep.2024.115206","url":null,"abstract":"<p><p>Metabolic reprogramming in both immune and cancer cells plays a crucial role in the antitumor immune response. Recent studies indicate that cancer metabolism not only sustains carcinogenesis and survival via altered signaling but also modulates immune cell function. Metabolic crosstalk within the tumor microenvironment results in nutrient competition and acidosis, thereby hindering immune cell functionality. Interestingly, immune cells also undergo metabolic reprogramming that enables their proliferation, differentiation, and effector functions. This review highlights the regulation of antitumor immune responses through metabolic reprogramming in cancer and immune cells and explores therapeutic strategies that target these metabolic pathways in cancer immunotherapy, including using chimeric antigen receptor (CAR)-T cells. We discuss innovative combinations of immunotherapy, cellular therapies, and metabolic interventions that could optimize the efficacy of existing treatment protocols.</p>","PeriodicalId":9798,"journal":{"name":"Cell reports","volume":"44 1","pages":"115206"},"PeriodicalIF":7.5,"publicationDate":"2025-01-28","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"142969727","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":1,"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
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