Biochimica et biophysica acta. Biomembranes最新文献_第6页

“Rich arginine and strong positive charge” antimicrobial protein protamine: From its action on cell membranes to inhibition of bacterial vital functions "富含精氨酸、带强正电荷 "的抗菌蛋白质原胺：从对细胞膜的作用到抑制细菌的生命功能

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-04-16 DOI: 10.1016/j.bbamem.2024.184323

Momoka Ookubo , Yuka Tashiro , Kosuke Asano , Yoshiharu Kamei , Yoshikazu Tanaka , Takayuki Honda , Takeshi Yokoyama , Michiyo Honda

{"title":"“Rich arginine and strong positive charge” antimicrobial protein protamine: From its action on cell membranes to inhibition of bacterial vital functions","authors":"Momoka Ookubo , Yuka Tashiro , Kosuke Asano , Yoshiharu Kamei , Yoshikazu Tanaka , Takayuki Honda , Takeshi Yokoyama , Michiyo Honda","doi":"10.1016/j.bbamem.2024.184323","DOIUrl":"https://doi.org/10.1016/j.bbamem.2024.184323","url":null,"abstract":"<div>Protamine, an antimicrobial protein derived from salmon sperm with a molecular weight of approximately 5 kDa, is composed of 60–70 % arginine and is a highly charged protein. Here, we investigated the mechanism of antimicrobial action of protamine against Cutibacterium acnes (C. acnes) focusing on its rich arginine content and strong positive charge. Especially, we focused on the attribution of dual mechanisms of antimicrobial protein, including membrane disruption or interaction with intracellular components. We first determined the dose-dependent antibacterial activity of protamine against C. acnes. In order to explore the interaction between bacterial membrane and protamine, we analyzed cell morphology, zeta potential, membrane permeability, and the composition of membrane fatty acid. In addition, the localization of protamine in bacteria was observed using fluorescent-labeled protamine. For investigation of the intracellular targets of protamine, bacterial translation was examined using a cell-free translation system. Based on our results, the mechanism of the antimicrobial action of protamine against C. acnes is as follows: 1) electrostatic interactions with the bacterial cell membrane; 2) self-internalization into the bacterial cell by changing the composition of the bacterial membrane; and 3) inhibition of bacterial growth by blocking translation inside the bacteria. However, owing to its strong electric charge, protamine can also interact with DNA, RNA, and other proteins inside the bacteria, and may inhibit various bacterial life processes beyond the translation process.</div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-04-16","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140557458","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Bioconcentration potential of ionic liquids: New data on membrane partitioning and its comparison with predictions obtained by COSMOmic 离子液体的生物浓缩潜力：膜分配的新数据及其与 COSMOmic 预测结果的比较

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-04-06 DOI: 10.1016/j.bbamem.2024.184320

Jakub Maculewicz , Anna Białk-Bielińska , Dorota Kowalska , Piotr Stepnowski , Stefan Stolte , Stephan Beil , Agnieszka Gajewicz-Skretna , Joanna Dołżonek

{"title":"Bioconcentration potential of ionic liquids: New data on membrane partitioning and its comparison with predictions obtained by COSMOmic","authors":"Jakub Maculewicz , Anna Białk-Bielińska , Dorota Kowalska , Piotr Stepnowski , Stefan Stolte , Stephan Beil , Agnieszka Gajewicz-Skretna , Joanna Dołżonek","doi":"10.1016/j.bbamem.2024.184320","DOIUrl":"https://doi.org/10.1016/j.bbamem.2024.184320","url":null,"abstract":"<div>Ionic liquids (ILs) have recently gained significant attention in both the scientific community and industry, but there is a limited understanding of the potential risks they might pose to the environment and human health, including their potential to accumulate in organisms. While membrane and storage lipids have been considered as primary sorption phases driving bioaccumulation, in this study we used an in vitro tool known as solid-supported lipid membranes (SSLMs) to investigate the affinity of ILs to membrane lipid - phosphatidylcholine and compare the results with an existing in silico model. Our findings indicate that ILs may have a strong affinity for the lipids that form cell membranes, with the key factor being the length of the cation's side chain. For quaternary ammonium cations, increase in membrane affinity (logMA) was observed from 3.45 ± 0.06 at 10 carbon atoms in chain to 4.79 ± 0.06 at 14 carbon atoms. We also found that the anion can significantly affect the membrane partitioning of the cation, even though the anions themselves tend to have weaker interactions with phospholipids than the cations of ILs. For 1-methyl-3-octylimidazolium cation the presence of tricyanomethanide anion caused increase in logMA to 4.23 ± 0.06. Although some of our data proved to be consistent with predictions made by the COSMOmic model, there are also significant discrepancies. These results suggest that further research is needed to improve our understanding of the mechanisms and structure-activity relationships involved in ILs bioconcentration and to develop more accurate predictive models.</div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-04-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140631230","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Differential effects of theasinensins and epigallocatechin-3-O-gallate on phospholipid bilayer structure and liposomal aggregation 表没食子儿茶素-3-O-棓酸盐和表没食子儿茶素-3-O-棓酸盐对磷脂双分子层结构和脂质体聚集的不同影响

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-04-04 DOI: 10.1016/j.bbamem.2024.184312

Asako Narai-Kanayama , Sumio Hayakawa , Takayuki Yoshino , Futa Honda , Hiroko Matsuda , Yumiko Oishi

{"title":"Differential effects of theasinensins and epigallocatechin-3-O-gallate on phospholipid bilayer structure and liposomal aggregation","authors":"Asako Narai-Kanayama , Sumio Hayakawa , Takayuki Yoshino , Futa Honda , Hiroko Matsuda , Yumiko Oishi","doi":"10.1016/j.bbamem.2024.184312","DOIUrl":"https://doi.org/10.1016/j.bbamem.2024.184312","url":null,"abstract":"<div>(−)-Epigallocatechin-3-O-gallate (EGCg), the major catechin responsible for the health-enhancing and disease-preventive effects of green tea, is susceptible to auto-oxidation at physiological pH levels. However, whether the oxidized EGCg resulting from its oral consumption possesses any bioactive functions remains unclear. This study presents a differential analysis of intact and oxidized EGCg regarding their interactions with phosphatidylcholine liposomes, serving as a simple biomembrane model. In the presence of ascorbic acid, pre-oxidized EGCg induced liposomal aggregation in a dose-dependent manner, whereas intact EGCg did not. Toxicity evaluation using calcein-loaded liposomes revealed that liposomal aggregation is associated with minimal membrane damage. Through fractionation of the oxidized EGCg sample, the fraction containing theasinensins showed high liposomal aggregation activity. Overall, these results suggest that oxidatively condensed EGCg dimers may stimulate various cells by altering the plasma membrane in a manner different from that of EGCg monomers.</div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-04-04","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140534951","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Modification of the RTX domain cap by acyl chains of adapted length rules the formation of functional hemolysin pores 用长度适宜的酰基链修饰 RTX 结构域帽，可形成功能性溶血素孔

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-04-01 DOI: 10.1016/j.bbamem.2024.184311

Anna Lepesheva , Michaela Grobarcikova , Adriana Osickova , David Jurnecka , Sarka Knoblochova , Monika Cizkova , Radim Osicka , Peter Sebo , Jiri Masin

{"title":"Modification of the RTX domain cap by acyl chains of adapted length rules the formation of functional hemolysin pores","authors":"Anna Lepesheva , Michaela Grobarcikova , Adriana Osickova , David Jurnecka , Sarka Knoblochova , Monika Cizkova , Radim Osicka , Peter Sebo , Jiri Masin","doi":"10.1016/j.bbamem.2024.184311","DOIUrl":"https://doi.org/10.1016/j.bbamem.2024.184311","url":null,"abstract":"<div>The acylated pore-forming Repeats in ToXin (RTX) cytolysins α-hemolysin (HlyA) and adenylate cyclase toxin (CyaA) preferentially bind to β2 integrins of myeloid leukocytes but can also promiscuously bind and permeabilize cells lacking the β2 integrins. We constructed a HlyA1–563/CyaA860–1706 chimera that was acylated either by the toxin-activating acyltransferase CyaC, using sixteen carbon-long (C16) acyls, or by the HlyC acyltransferase using fourteen carbon-long (C14) acyls. Cytolysin assays with the C16- or C14-acylated HlyA/CyaA chimeric toxin revealed that the RTX domain of CyaA can functionally replace the RTX domain of HlyA only if it is modified by C16-acyls on the Lys983 residue of CyaA. The C16-monoacylated HlyA/CyaA chimera was as pore-forming and cytolytic as native HlyA, whereas the C14-acylated chimera exhibited very low pore-forming activity. Hence, the capacity of the RTX domain of CyaA to support the insertion of the N-terminal pore-forming domain into the target cell membrane, and promote formation of toxin pores, strictly depends on the modification of the Lys983 residue by an acyl chain of adapted length.</div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-04-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S0005273624000427/pdfft?md5=07a9ac943f2962b725a6f273cf3b0e95&pid=1-s2.0-S0005273624000427-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140540417","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Fumonisin B1 protects against long-chained polyunsaturated fatty acid-induced cell death in HepG2 cells – implications for cancer promotion 伏马菌素 B1 可防止长链多不饱和脂肪酸诱导的 HepG2 细胞死亡--对癌症的促进作用。

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-03-11 DOI: 10.1016/j.bbamem.2024.184310

Sylvia Riedel , Stefan Abel , Hester-Mari Burger , Sonja Swanevelder , Wentzel C.A. Gelderblom

{"title":"Fumonisin B1 protects against long-chained polyunsaturated fatty acid-induced cell death in HepG2 cells – implications for cancer promotion","authors":"Sylvia Riedel , Stefan Abel , Hester-Mari Burger , Sonja Swanevelder , Wentzel C.A. Gelderblom","doi":"10.1016/j.bbamem.2024.184310","DOIUrl":"10.1016/j.bbamem.2024.184310","url":null,"abstract":"<div>Fumonisin B1 (FB1), a food-borne mycotoxin, is a cancer promoter in rodent liver and augments proliferation of initiated cells while inhibiting the growth of normal hepatocytes by disrupting lipid biosynthesis at various levels. HepG2 cancer cells exhibited resistance to FB1-induced toxic effects presumably due to their low content of polyunsaturated fatty acids (PUFA) even though FB1-typical lipid changes were observed, e.g. significantly increased phosphatidylethanolamine (PE), decreased sphingomyelin and cholesterol content, increased sphinganine (Sa) and sphinganine/sphingosine ratio, increased C18:1ω-9, decreased C20:4ω-6 content in PE and decreased C20:4ω-6_PC/PE ratio. Increasing PUFA content of HepG2 cells with phosphatidylcholine (PC) vesicles containing C20:4ω-6 (SAPC) or C22:6ω-3 (SDPC) disrupted cell survival, cellular redox status and induced oxidative stress and apoptosis. A partially protective effect of FB1 was evident in PUFA-enriched HepG2 cells which may be related to the FB1-induced reduction in oxidative stress and the disruption of key cell membrane constituents indicative of a resistant lipid phenotype. Interactions between different ω-6 and ω-3 PUFA, membrane constituents including cholesterol, and the glycerophospho- and sphingolipids and FB1 in this cell model provide further support for the resistant lipid phenotype and its role in the complex cellular effects underlying the cancer promoting potential of the fumonisins.</div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-03-11","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.sciencedirect.com/science/article/pii/S0005273624000415/pdfft?md5=9160b5c0d6b6eee7ea5ec7b048c33dd6&pid=1-s2.0-S0005273624000415-main.pdf","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140118646","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Perception and protection: The role of Bce-modules in antimicrobial peptide resistance 感知与保护：Bce 模块在抗菌肽耐药性中的作用。

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-03-07 DOI: 10.1016/j.bbamem.2024.184309

Benjamin J. Orlando

引用次数: 0

Coordinated regulation of phosphatidylinositol 4-phosphate and phosphatidylserine levels by Osh4p and Osh5p is an essential regulatory mechanism in autophagy Osh4p和Osh5p对4-磷酸肌醇磷脂和磷脂酰丝氨酸水平的协调调控是自噬过程中必不可少的调控机制。

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-03-02 DOI: 10.1016/j.bbamem.2024.184308

Moe Muramoto , Nanaru Mineoka , Kayoko Fukuda , Sayuri Kuriyama , Tatsunori Masatani , Akikazu Fujita

{"title":"Coordinated regulation of phosphatidylinositol 4-phosphate and phosphatidylserine levels by Osh4p and Osh5p is an essential regulatory mechanism in autophagy","authors":"Moe Muramoto , Nanaru Mineoka , Kayoko Fukuda , Sayuri Kuriyama , Tatsunori Masatani , Akikazu Fujita","doi":"10.1016/j.bbamem.2024.184308","DOIUrl":"10.1016/j.bbamem.2024.184308","url":null,"abstract":"<div>Macroautophagy (hereafter autophagy) is an intracellular degradative pathway in budding yeast cells. Certain lipid types play essential roles in autophagy; yet the precise mechanisms regulating lipid composition during autophagy remain unknown. Here, we explored the role of the Osh family proteins in the modulating lipid composition during autophagy in budding yeast. Our results showed that osh1-osh7∆ deletions lead to autophagic dysfunction, with impaired GFP-Atg8 processing and the absence of autophagosomes and autophagic bodies in the cytosol and vacuole, respectively. Freeze-fracture electron microscopy (EM) revealed elevated phosphatidylinositol 4-phosphate (PtdIns(4)P) levels in cytoplasmic and luminal leaflets of autophagic bodies and vacuolar membranes in all deletion mutants. Phosphatidylserine (PtdSer) levels were significantly decreased in the autophagic bodies and vacuolar membranes in osh4∆ and osh5∆ mutants, whereas no significant changes were observed in other osh deletion mutants. Furthermore, we identified defects in autophagic processes in the osh4∆ and osh5∆ mutants, including rare autophagosome formation in the osh5∆ mutant and accumulation of autophagic bodies in the vacuole in the osh4∆ mutant, even in the absence of the proteinase inhibitor PMSF. These findings suggest that Osh4p and Osh5p play crucial roles in the transport of PtdSer to autophagic bodies and autophagosome membranes, respectively. The precise control of lipid composition in the membranes of autophagosomes and autophagic bodies by Osh4p and Osh5p represents an important regulatory mechanism in autophagy.</div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-03-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"140027297","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Recognition and remodeling of endosomal zones by sorting nexins 分拣蛋白对内膜区的识别和重塑。

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-02-24 DOI: 10.1016/j.bbamem.2024.184305

Michael Overduin, Rakesh Bhat

{"title":"Recognition and remodeling of endosomal zones by sorting nexins","authors":"Michael Overduin, Rakesh Bhat","doi":"10.1016/j.bbamem.2024.184305","DOIUrl":"10.1016/j.bbamem.2024.184305","url":null,"abstract":"<div>The proteolipid code determines how cytosolic proteins find and remodel membrane surfaces. Here, we investigate how this process works with sorting nexins Snx1 and Snx3. Both proteins form sorting machines by recognizing membrane zones enriched in phosphatidylinositol 3-phosphate (PI3P), phosphatidylserine (PS) and cholesterol. This co-localized combination forms a unique “lipid codon” or lipidon that we propose is responsible for endosomal targeting, as revealed by structures and interactions of their PX domain-based readers. We outline a membrane recognition and remodeling mechanism for Snx1 and Snx3 involving this code element alongside transmembrane pH gradients, dipole moment-guided docking and specific protein-protein interactions. This generates an initial membrane-protein assembly (memtein) that then recruits retromer and additional PX proteins to recruit cell surface receptors for sorting to the trans-Golgi network (TGN), lysosome and plasma membranes. Post-translational modification (PTM) networks appear to regulate how the sorting machines form and operate at each level. The commonalities and differences between these sorting nexins show how the proteolipid code orchestrates parallel flows of molecular information from ribosome emergence to organelle genesis, and illuminates a universally applicable model of the membrane.</div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-02-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139970861","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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

Phosphorus-31: A table-top method for 3D B1-field amplitude and phase measurements 磷-31：三维 B1 场振幅和相位测量的桌面方法。

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-02-24 DOI: 10.1016/j.bbamem.2024.184307

V. Brandejsky , O. Leinhard Dahlqvist , E. Lund , P. Lundberg

引用次数: 0

Spectroscopic behavior of bufotenine and bufotenine N-oxide: Solvent and pH effects and interaction with biomembrane models 布福滕宁和布福滕宁 N-氧化物的光谱行为：溶剂和 pH 值的影响以及与生物膜模型的相互作用。

IF 3.4 3区生物学

Biochimica et biophysica acta. Biomembranes Pub Date : 2024-02-24 DOI: 10.1016/j.bbamem.2024.184304

Gustavo Almeida e Silva , Vinícius Galvão Wakui , Lucília Kato , Cássia A. Marquezin

{"title":"Spectroscopic behavior of bufotenine and bufotenine N-oxide: Solvent and pH effects and interaction with biomembrane models","authors":"Gustavo Almeida e Silva , Vinícius Galvão Wakui , Lucília Kato , Cássia A. Marquezin","doi":"10.1016/j.bbamem.2024.184304","DOIUrl":"10.1016/j.bbamem.2024.184304","url":null,"abstract":"<div>Bufotenine is a fluorescent analog of Dimethyltryptamine (DMT) that has been widely studied due to its psychedelic properties and biological activity. However, little is known about its spectroscopic properties in different media. Thus, we present in this work, for the first time, the spectroscopic behavior of bufotenine and bufotenine N-oxide by means of their fluorescence properties. Both molecules exhibit changes in optical absorption and emission spectra with variations in pH of the medium and in different solvents. Assays in the presence of biomembranes models, like micelles and liposomes, were also performed. In surfactants titration experiments, the spectral shift observed in fluorescence shows the interaction of both molecules with pre-micellar structures and with micelles. Steady state anisotropy measurements show that both bufotenine and bufotenine N-oxide, in the studied concentration range, interact with liposomes without causing changes in the fluidity of the lipid bilayer. These results can be useful in studies that aim at searching for new compounds, inspired by bufotenine and bufotenine N-oxide, with relevant pharmacological activities and also in studies that use these molecules as markers of psychiatric disorders.</div>","PeriodicalId":8831,"journal":{"name":"Biochimica et biophysica acta. Biomembranes","volume":null,"pages":null},"PeriodicalIF":3.4,"publicationDate":"2024-02-24","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"139970862","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"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