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Scalable production of siRNA-encapsulated extracellular vesicles for the inhibition of KRAS-mutant cancer using acoustic shock waves 利用声波冲击波大规模生产用于抑制 KRAS 突变癌症的 siRNA 包裹细胞外囊泡
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-26 DOI: 10.1002/jev2.12508
Hyo Kyeong Kim, Yujeong Choi, Kyoung Hwa Kim, Yeongju Byun, Tae Hee Kim, Jae Hwan Kim, Shung Hyun An, DaeHo Bae, Myeong Kwan Choi, Minyoung Lee, Gwansuk Kang, Jihwa Chung, Seok-Hyun Kim, Kihwan Kwon

Extracellular vesicles (EVs) have emerged as a potential delivery vehicle for nucleic-acid-based therapeutics, but challenges related to their large-scale production and cargo-loading efficiency have limited their therapeutic potential. To address these issues, we developed a novel “shock wave extracellular vesicles engineering technology” (SWEET) as a non-genetic, scalable manufacturing strategy that uses shock waves (SWs) to encapsulate siRNAs in EVs. Here, we describe the use of the SWEET platform to load large quantities of KRASG12C-targeting siRNA into small bovine-milk-derived EVs (sBMEVs), with high efficiency. The siRNA-loaded sBMEVs effectively silenced oncogenic KRASG12C expression in cancer cells; they inhibited tumour growth when administered intravenously in a non-small cell lung cancer xenograft mouse model. Our study demonstrates the potential for the SWEET platform to serve as a novel method that allows large-scale production of cargo-loaded EVs for use in a wide range of therapeutic applications.

细胞外囊泡(EVs)已成为一种潜在的核酸类治疗药物递送载体,但其大规模生产和货物装载效率方面的挑战限制了其治疗潜力。为了解决这些问题,我们开发了一种新颖的 "冲击波细胞外囊泡工程技术"(SWEET),作为一种非遗传、可扩展的生产策略,利用冲击波(SWs)将 siRNA 封装在 EVs 中。在这里,我们描述了如何利用 SWEET 平台将大量 KRASG12C 靶向 siRNA 高效地装载到小型牛乳衍生 EVs(sBMEVs)中。加载了 siRNA 的 sBMEVs 能有效抑制癌细胞中致癌基因 KRASG12C 的表达;在非小细胞肺癌异种移植小鼠模型中静脉注射这些 siRNA 能抑制肿瘤生长。我们的研究表明,SWEET 平台有可能成为一种新方法,大规模生产载货 EVs,用于广泛的治疗应用。
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引用次数: 0
Benchmarking transcriptome deconvolution methods for estimating tissue- and cell-type-specific extracellular vesicle abundances 用于估算组织和细胞类型特异性细胞外囊泡丰度的转录组去卷积方法基准测试
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-25 DOI: 10.1002/jev2.12511
Jannik Hjortshøj Larsen, Iben Skov Jensen, Per Svenningsen

Extracellular vesicles (EVs) contain cell-derived lipids, proteins and RNAs; however, determining the tissue- and cell-type-specific EV abundances in body fluids remains a significant hurdle for our understanding of EV biology. While tissue- and cell-type-specific EV abundances can be estimated by matching the EV's transcriptome to a tissue's/cell type's expression signature using deconvolutional methods, a comparative assessment of deconvolution methods' performance on EV transcriptome data is currently lacking. We benchmarked 11 deconvolution methods using data from four cell lines and their EVs, in silico mixtures, 118 human plasma and 88 urine EVs. We identified deconvolution methods that estimated cell type-specific abundances of pure and in silico mixed cell line-derived EV samples with high accuracy. Using data from two urine EV cohorts with different EV isolation procedures, four deconvolution methods produced highly similar results. The three methods were also concordant in their tissue- and cell-type-specific plasma EV abundance estimates. We identified driving factors for deconvolution accuracy and highlighted the importance of implementing biological knowledge in creating the tissue/cell type signature. Overall, our analyses demonstrate that the deconvolution algorithms DWLS and CIBERSORTx produce highly similar and accurate estimates of tissue- and cell-type-specific EV abundances in biological fluids.

细胞外囊泡(EV)含有细胞衍生的脂质、蛋白质和 RNA;然而,确定体液中组织和细胞类型特异的 EV 丰度仍然是我们了解 EV 生物学的一个重大障碍。虽然组织和细胞类型特异性的 EV 丰度可以通过使用去卷积方法将 EV 的转录组与组织/细胞类型的表达特征相匹配来估算,但目前还缺乏对去卷积方法在 EV 转录组数据上的性能的比较评估。我们使用来自四种细胞系及其 EV、硅学混合物、118 人血浆和 88 尿液 EV 的数据,对 11 种去卷积方法进行了基准测试。我们确定了一些去卷积方法,这些方法能高精度地估算纯细胞系和硅学混合细胞系衍生 EV 样本的细胞类型特异性丰度。使用来自两个尿液EV队列的数据和不同的EV分离程序,四种解卷积方法得出了高度相似的结果。这三种方法对组织和细胞类型特异性血浆 EV 丰度的估计也是一致的。我们确定了去卷积准确性的驱动因素,并强调了在创建组织/细胞类型特征时应用生物学知识的重要性。总之,我们的分析表明,解卷积算法 DWLS 和 CIBERSORTx 对生物液体中组织和细胞类型特异性 EV 丰度的估计高度相似且准确。
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引用次数: 0
Extracellular vesicles derived from melanoma cells induce carcinoma-associated fibroblasts via miR-92b-3p mediated downregulation of PTEN 来自黑色素瘤细胞的细胞外囊泡通过 miR-92b-3p 介导的 PTEN 下调诱导癌相关成纤维细胞。
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-24 DOI: 10.1002/jev2.12509
Stefanie Kewitz-Hempel, Nicola Windisch, Gerd Hause, Lutz Müller, Cord Sunderkötter, Dennis Gerloff

In melanoma, carcinoma-associated fibroblasts (CAFs) are important cellular components in the tumour microenvironment due to their potential to promote tumour growth and metastatic spread of malignant cells. Melanoma cells have the ability to affect non-tumour cells in the microenvironment by releasing extracellular vesicles (EVs). The mechanisms responsible for reprogramming normal dermal fibroblasts (NHDFs) into CAFs remain incompletely understood. However, it is likely thought to be mediated by melanoma-specific miRNAs, which are transported by EVs derived from melanoma cells. Therefore, we wondered if one of the most enriched miRNAs in EVs secreted by melanoma cells, miR-92b-3p, is involved in the conversion of normal fibroblasts into CAFs. We observed that melanoma cell-derived EVs indeed delivered miR-92b-3p into NHDFs and that its accumulation correlated with CAF formation, as demonstrated by enhanced expression of CAF marker genes and increased proliferation and migration. Overexpression of miR-92b-3p in NHDFs revealed similar results, while EVs deficient of miR-92b-3p did not induce a CAF phenotype. As a target we identified PTEN, whose repression led to increased expression of CAF markers. We thus provide a novel pathway of intercellular communication by which melanoma cells control the transformation of CAFs by virtue of EV-transported miRNAs.

在黑色素瘤中,癌相关成纤维细胞(CAFs)是肿瘤微环境中的重要细胞成分,因为它们具有促进肿瘤生长和恶性细胞转移扩散的潜力。黑色素瘤细胞能够通过释放细胞外囊泡 (EV) 影响微环境中的非肿瘤细胞。将正常真皮成纤维细胞(NHDFs)重编程为 CAFs 的机制尚不完全清楚。不过,人们认为这可能是由黑色素瘤特异性 miRNAs 介导的,而这些 miRNAs 是由黑色素瘤细胞的 EVs 运输的。因此,我们想知道黑色素瘤细胞分泌的EVs中最富集的miRNA之一miR-92b-3p是否参与了正常成纤维细胞向CAFs的转化。我们观察到,黑色素瘤细胞衍生的EV确实将miR-92b-3p传递到了NHDFs中,其积累与CAF的形成有关,这表现在CAF标记基因的表达增强、增殖和迁移增加。在NHDFs中过表达miR-92b-3p也显示了类似的结果,而缺乏miR-92b-3p的EVs不会诱导CAF表型。我们确定了 PTEN 作为靶点,它的抑制会导致 CAF 标志物的表达增加。因此,我们提供了一种新的细胞间通信途径,黑色素瘤细胞通过 EV 运送的 miRNA 控制 CAF 的转化。
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引用次数: 0
Enhancing protective immunity against bacterial infection via coating nano-Rehmannia glutinosa polysaccharide with outer membrane vesicles 通过外膜囊泡包裹纳米地黄多糖,增强对细菌感染的保护性免疫。
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-24 DOI: 10.1002/jev2.12514
Yee Huang, Jiaying Sun, Xuemei Cui, Xuefeng Li, Zizhe Hu, Quanan Ji, Guolian Bao, Yan Liu

With the coming of the post-antibiotic era, there is an increasingly urgent need for safe and efficient antibacterial vaccines. Bacterial outer membrane vesicles (OMVs) have received increased attention recently as a potential subunit vaccine. OMVs are non-replicative and contain the principle immunogenic bacterial antigen, which circumvents the safety concerns of live-attenuated vaccines. Here, we developed a novel nano-vaccine by coating OMVs onto PEGylated nano-Rehmannia glutinosa polysaccharide (pRL) in a structure consisting of concentric circles, resulting in a more stable vaccine with improved immunogenicity. The immunological function of the pRL-OMV formulation was evaluated in vivo and in vitro, and the underlying mechanism was studied though transcriptomic analysis. The pRL-OMV formulation significantly increased dendritic cell (DC) proliferation and cytokine secretion. Efficient phagocytosis of the formulation by DCs was accompanied by DC maturation. Further, the formulation demonstrated superior lymph node targeting, contributing to a potent mixed cellular response and bacterial-specific antibody response against Bordetella bronchiseptica infection. Specifically, transcriptomic analysis revealed that the immune protection function correlated with T-cell receptor signalling and Th1/Th2/Th17 differentiation, among other markers of enhanced immunological activity. These findings have implications for the future application of OMV-coated nano-carriers in antimicrobial immunotherapy.

随着后抗生素时代的到来,人们对安全高效的抗菌疫苗的需求日益迫切。作为一种潜在的亚单位疫苗,细菌外膜囊泡 (OMV) 近来受到越来越多的关注。OMVs 不具有复制性,含有主要的免疫原性细菌抗原,从而避免了减毒活疫苗的安全性问题。在此,我们开发了一种新型纳米疫苗,将 OMV 包覆在 PEG 化纳米地黄多糖(pRL)上,形成同心圆结构,从而获得了一种更稳定、免疫原性更强的疫苗。我们在体内和体外评估了 pRL-OMV 配方的免疫功能,并通过转录组分析研究了其潜在机制。pRL-OMV制剂显著增加了树突状细胞(DC)的增殖和细胞因子的分泌。DC对制剂的高效吞噬伴随着DC的成熟。此外,该制剂还表现出卓越的淋巴结靶向性,有助于产生针对支气管败血波氏杆菌感染的强效混合细胞反应和细菌特异性抗体反应。具体来说,转录组分析表明,免疫保护功能与 T 细胞受体信号传导和 Th1/Th2/Th17 分化以及其他免疫活性增强的标记相关。这些发现对未来在抗菌免疫疗法中应用 OMV 涂层纳米载体具有重要意义。
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引用次数: 0
Plasma-derived extracellular vesicles (EVs) as biomarkers of sepsis in burn patients via label-free Raman spectroscopy 通过无标记拉曼光谱将血浆源性细胞外囊泡 (EV) 作为烧伤患者败血症的生物标记物
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-19 DOI: 10.1002/jev2.12506
Hannah J. O'Toole, Neona M. Lowe, Vishalakshi Arun, Anna V. Kolesov, Tina L. Palmieri, Nam K. Tran, Randy P. Carney

Sepsis following burn trauma is a global complication with high mortality, with ∼60% of burn patient deaths resulting from infectious complications. Diagnosing sepsis is complicated by confounding clinical manifestations of the burn injury, and current biomarkers lack the sensitivity and specificity required for prompt treatment. There is a strong rationale to assess circulating extracellular vesicles (EVs) from patient liquid biopsy as sepsis biomarkers due to their release by pathogens from bacterial biofilms and roles in the subsequent immune response. This study applies Raman spectroscopy to patient plasma-derived EVs for rapid, sensitive, and specific detection of sepsis in burn patients, achieving 97.5% sensitivity and 90.0% specificity. Furthermore, spectral differences between septic and non-septic burn patient EVs could be traced to specific glycoconjugates of bacterial strains associated with sepsis morbidity. This work illustrates the potential application of EVs as biomarkers in clinical burn trauma care and establishes Raman analysis as a fast, label-free method to specifically identify features of bacterial EVs relevant to infection amongst the host background.

烧伤创伤后败血症是一种全球性并发症,死亡率很高,60%的烧伤患者死于感染并发症。脓毒症的诊断因烧伤的临床表现而变得复杂,目前的生物标志物缺乏及时治疗所需的灵敏度和特异性。由于病原体从细菌生物膜中释放出细胞外囊泡 (EV),并在随后的免疫反应中发挥作用,因此将患者液体生物样本中的循环细胞外囊泡 (EV) 作为败血症生物标志物进行评估具有很强的合理性。本研究将拉曼光谱应用于患者血浆中的 EVs,以快速、灵敏、特异地检测烧伤患者的败血症,灵敏度达 97.5%,特异度达 90.0%。此外,脓毒症和非脓毒症烧伤患者 EVs 之间的光谱差异可追溯到与脓毒症发病率相关的细菌菌株的特异性糖结合物。这项工作说明了在临床烧伤创面护理中将 EVs 作为生物标记物的潜在应用价值,并证明拉曼分析是一种快速、无标记的方法,可在宿主背景中特异性地识别与感染相关的细菌 EVs 特征。
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引用次数: 0
Therapeutic potential of mesenchymal stem cell-derived extracellular vesicles in SARS-CoV-2 and H1N1 influenza-induced acute lung injury 间充质干细胞衍生的细胞外囊泡在 SARS-CoV-2 和 H1N1 流感诱发的急性肺损伤中的治疗潜力
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-10 DOI: 10.1002/jev2.12495
Jun Ho Lee, Hyungtaek Jeon, Jan Lötvall, Byong Seung Cho

Mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) have shown anti-inflammatory potential in multiple inflammatory diseases. In the March 2022 issue of the Journal of Extracellular Vesicles, it was shown that EVs from human MSCs can suppress severe acute respiratory distress syndrome, coronavirus 2 (SARS-CoV-2) replication and can mitigate the production and release of infectious virions. We therefore hypothesized that MSC-EVs have an anti-viral effect in SARS-CoV-2 infection in vivo. We extended this question to ask whether also other respiratory viral infections could be treated by MSC-EVs. Adipose stem cell-derived EVs (ASC-EVs) were isolated using tangential flow filtration from conditioned media obtained from a multi-flask cell culture system. The effects of the ASC-EVs were tested  in Vero E6 cells in vitro. ASC-EVs were also given i.v. to SARS-CoV-2 infected Syrian Hamsters, and H1N1 influenza virus infected mice. The ASC-EVs attenuated SARS-CoV-2 virus replication in Vero E6 cells and reduced body weight and signs of lung injury in infected Syrian hamsters. Furthermore, ASC-EVs increased the survival rate of influenza A-infected mice and attenuated signs of lung injury. In summary, this study suggests that ASC-EVs can have beneficial therapeutic effects in models of virus-infection-associated acute lung injury and may potentially be developed to treat lung injury in humans.

间充质干细胞(MSC)衍生的细胞外囊泡(EVs)已在多种炎症疾病中显示出抗炎潜力。2022年3月出版的《细胞外囊泡杂志》(Journal of Extracellular Vesicles)显示,来自人类间充质干细胞的细胞外囊泡能抑制严重急性呼吸窘迫综合征冠状病毒2(SARS-CoV-2)的复制,并能减轻传染性病毒的产生和释放。因此,我们假设间充质干细胞-EVs 在体内感染 SARS-CoV-2 时具有抗病毒作用。我们将这一问题延伸至间叶干细胞-EV是否也能治疗其他呼吸道病毒感染。我们使用切向流过滤法从多层细胞培养系统获得的条件培养基中分离出了脂肪干细胞衍生的EVs(ASC-EVs)。在体外 Vero E6 细胞中测试了 ASC-EVs 的作用。此外,还为感染了 SARS-CoV-2 的叙利亚仓鼠和感染了 H1N1 流感病毒的小鼠静脉注射了 ASC-EV。ASC-EVs 可减轻 SARS-CoV-2 病毒在 Vero E6 细胞中的复制,减轻受感染叙利亚仓鼠的体重和肺损伤症状。此外,ASC-EVs 还能提高甲型流感感染小鼠的存活率,减轻肺损伤症状。总之,这项研究表明,ASC-EVs 可在病毒感染相关急性肺损伤模型中产生有益的治疗效果,并有可能开发用于治疗人类肺损伤。
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引用次数: 0
Extracellular vesicle analytical science loses a touch of creativity and kindness 细胞外囊泡分析科学失去了创造力和亲切感
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-10 DOI: 10.1002/jev2.12504
Paolo Bergese, Marcella Chiari, Alessandro Gori, Benedetta Bussolati, Pietro Parisse, the EVIta Board
<p></p><p>It is with profound sadness and sorrow that we announce the unexpected passing of our cherished friend and colleague, Marina Cretich. Marina, a renowned scientist in the field of bioanalytical chemistry, passed away on June 29th 2024, leaving behind a heritage of invaluable contributions to science. Her career was marked by a relentless pursuit of knowledge and an unwavering dedication to advancing scientific understanding.</p><p>Marina graduated in Biological Sciences with a specialization in Molecular Biology from the University of Milano in 1998 with Prof. Piergiorgio Righetti. She then joined the National Research Council of Italy (CNR), where she served as a Researcher at the Institute of Chemistry for Molecular Recognition in Milan, becoming a key figure in the Analytical Microsystem laboratory guided by Dr. Marcella Chiari. Thanks to the interdisciplinarity of her approach, she made significant strides in the progress of advanced methods and materials for bio-molecular recognition, together with high-sensitive and selective biosensors, providing new tools for detecting biomolecules with unprecedented accuracy and efficiency. Her strong focus on integrating bioanalytical techniques, microfluidics and detection tools, enabled more rapid and precise analyses, enhancing the capabilities of lab-on-a-chip devices, towards point-of-care diagnostics.</p><p>These studies paved the road to her approach to extracellular vesicles, addressing the challenges posed by their separation and analysis. Specifically, Marina was working on affinity-capturing protocols from complex samples for EV isolation and on microarray platforms for their molecular characterization (Daaboul et al., <span>2016</span>). Her interest in the field was raised by participating in the EU project INDEX, coordinated by Dr Marcella Chiari, and became true love for this fascinating and challenging area. This led Marina to establish her own research team and to find the Extracellular Vesicle Lab at SCITEC-CNR. Soon after, she was the Coordinator of the EU project MARVEL, a multi-partner project at the intersection of chemistry and technology, biology and translational medicine, which was centred around the use of membrane-sensing peptides (MSP) as enabling tools for the multiscale EV isolation (Gori et al., <span>2020</span>, Gori et al, <span>2024</span>). She was a true pioneer in this field and she consolidated a leading expertise in the area of ultrasensitive EV analysis, aiming to fill existing gaps in the clinical translation of EVs in diagnostics. Her contribution is witnessed by remarkable scientific contributions (Frigerio et al., <span>2022</span>; Musicò et al., <span>2024</span>) and, despite very recent, the concepts and technologies that she developed set the basis for an ever-increasing number of preclinical and clinical collaborations encompassing EV analysis in the fields of neurodegeneration, cancer, and heart diseases.</p><p>In recent years, she also dedic
然而,人们记住她的不仅仅是她的科学贡献,还有她的善良、慷慨、可靠以及她对所有有幸认识她的人产生的积极影响。玛丽娜是敬业和专业精神的典范,是激励我们所有人的灯塔和基石。
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引用次数: 0
The activity of the quorum sensing regulator HapR is modulated by the bacterial extracellular vesicle (BEV)-associated protein ObfA of Vibrio cholerae 霍乱弧菌的细菌胞外囊泡(BEV)相关蛋白 ObfA 可调节法定人数感应调节因子 HapR 的活性
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-10 DOI: 10.1002/jev2.12507
Stephan P. Ebenberger, Fatih Cakar, Yi-Chi Chen, Katharina Pressler, Leo Eberl, Stefan Schild

Vibrio cholerae, a facultative human pathogen and causative agent of the severe diarrheal disease cholera, transits between the human intestinal tract and aquatic reservoirs. Like other bacterial species, V. cholerae continuously releases bacterial extracellular vesicles (BEVs) from its surface, which have been recently characterised for their role during in vivo colonisation. However, between epidemic outbreaks, V. cholerae persists in the biofilm mode for extended periods in aquatic reservoirs, which enhances environmental fitness and host transition. In this study, we investigated the effect of V. cholerae BEVs on biofilm formation, a critical feature for ex vivo survival. In contrast to BEVs from planktonic cultures, our results show that physiological concentrations of BEVs from dynamic biofilm cultures facilitate V. cholerae biofilm formation, which could be linked to a proteinaceous factor. Comparative proteomic analyses of planktonic- and biofilm-derived BEVs identified a previously uncharacterised outer membrane protein as an abundant component of dynamic biofilm-derived BEVs, which was found to be responsible for the BEV-dependent enhancement of biofilm production. Consequently, this protein was named outer membrane-associated biofilm facilitating protein A (ObfA). Comprehensive molecular studies unravelled ObfA as a negative modulator of HapR activity. HapR is a key transcriptional regulator of the V. cholerae quorum sensing (QS) cascade acting as a potent repressor of biofilm formation and virulence. Consistently, obfA mutants not only exhibited reduced biofilm production but also reduced colonisation fitness. Surprisingly, our results demonstrate that ObfA does not affect HapR through the canonical QS system but via the Csr-cascade altering the expression of the small regulatory RNAs CsrC and CsrD. In summary, this study elucidates a novel intraspecies BEV-based communication in V. cholerae that influences biofilm formation and colonisation fitness via a new regulatory pathway involving HapR, Csr-cascade and the BEV-associated protein ObfA.

霍乱弧菌是一种面性人类病原体,也是严重腹泻病霍乱的致病菌,它在人类肠道和水生水库之间传播。与其他细菌物种一样,霍乱弧菌不断从其表面释放细菌胞外囊泡(BEVs),这些囊泡在体内定植过程中的作用最近已得到证实。然而,在流行病爆发的间歇期,霍乱弧菌会在水生水库中长时间以生物膜模式存活,从而提高环境适应性和宿主转换能力。在这项研究中,我们调查了霍乱弧菌 BEV 对生物膜形成的影响,生物膜是霍乱弧菌体内外存活的关键特征。与来自浮游生物培养物的 BEVs 不同,我们的研究结果表明,来自动态生物膜培养物的生理浓度的 BEVs 会促进霍乱弧菌生物膜的形成,这可能与蛋白质因子有关。对浮游生物和生物膜衍生的 BEV 进行的蛋白质组学比较分析发现,动态生物膜衍生的 BEV 中含有一种以前未定性的外膜蛋白,它是 BEV 依赖性增强生物膜生成的原因。因此,这种蛋白质被命名为外膜相关生物膜促进蛋白 A(ObfA)。综合分子研究发现,ObfA 是 HapR 活性的负调制剂。HapR 是霍乱弧菌法定量感应(QS)级联的一个关键转录调节因子,对生物膜的形成和毒力起着强有力的抑制作用。一致的是,obfA 突变体不仅生物膜生成减少,而且定殖能力也降低。令人惊讶的是,我们的研究结果表明,ObfA 并不是通过典型的 QS 系统影响 HapR,而是通过 Csr 级联改变小调控 RNA CsrC 和 CsrD 的表达。总之,本研究阐明了霍乱弧菌中一种新的种内基于 BEV 的交流,这种交流通过涉及 HapR、Csr 级联和 BEV 相关蛋白 ObfA 的新调控途径影响生物膜的形成和定殖适应性。
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引用次数: 0
The 8-oxoguanine DNA glycosylase-synaptotagmin 7 pathway increases extracellular vesicle release and promotes tumour metastasis during oxidative stress 在氧化应激过程中,8-氧鸟嘌呤DNA糖基化酶-synaptotagmin 7途径会增加细胞外囊泡的释放并促进肿瘤转移。
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-05 DOI: 10.1002/jev2.12505
Ying Ma, Jiarong Guo, Haipeng Rao, Jingyu Xin, Xinyi Song, Rui Liu, Shan Shao, Jiajia Hou, Liyu Kong, Zhigang Hu, Lingfeng He, Feiyan Pan, Zhigang Guo

Reactive oxygen species (ROS)-induced oxidative DNA damages have been considered the main cause of mutations in genes, which are highly related to carcinogenesis and tumour progression. Extracellular vesicles play an important role in cancer metastasis. However, the precise role of DNA oxidative damage in extracellular vesicles (EVs)-mediated cancer cell migration and invasion remains unclear. Here, we reveal that ROS-mediated DNA oxidative damage signalling promotes tumour metastasis through increasing EVs release. Mechanistically, 8-oxoguanine DNA glycosylase (OGG1) recognises and binds to its substrate 8-oxo-7,8-dihydroguanine (8-oxoG), recruiting NF-κB to the synaptotagmin 7 (SYT7) promoter and thereby triggering SYT7 transcription. The upregulation of SYT7 expression leads to increased release of E-cadherin-loaded EVs, which depletes intracellular E-cadherin, thereby inducing epithelial-mesenchymal transition (EMT). Notably, Th5487, the inhibitor of DNA binding activity of OGG1, blocks the recognition and transmission of oxidative signals, alleviates SYT7 expression and suppresses EVs release, thereby preventing tumour progression in vitro and in vivo. Collectively, our study illuminates the significance of 8-oxoG/OGG1/SYT7 axis-driven EVs release in oxidative stress-induced tumour metastasis. These findings provide a deeper understanding of the molecular basis of cancer progression and offer potential avenues for therapeutic intervention.

活性氧(ROS)诱导的 DNA 氧化损伤被认为是基因突变的主要原因,而基因突变与癌变和肿瘤进展高度相关。细胞外囊泡在癌症转移中发挥着重要作用。然而,DNA氧化损伤在细胞外囊泡介导的癌细胞迁移和侵袭中的确切作用仍不清楚。在这里,我们揭示了 ROS 介导的 DNA 氧化损伤信号通过增加 EVs 释放促进肿瘤转移。从机理上讲,8-氧鸟嘌呤 DNA 糖基化酶(OGG1)识别并与其底物 8-氧代-7,8-二氢鸟嘌呤(8-oxoG)结合,将 NF-κB 募集到突触柄蛋白 7(SYT7)启动子上,从而触发 SYT7 的转录。SYT7 表达的上调导致 E-cadherin 负载的 EVs 释放增加,从而消耗细胞内的 E-cadherin,从而诱导上皮-间质转化(EMT)。值得注意的是,OGG1 DNA结合活性抑制剂Th5487能阻断氧化信号的识别和传递,减轻SYT7的表达并抑制EVs的释放,从而阻止肿瘤在体外和体内的进展。总之,我们的研究阐明了 8-oxoG/OGG1/SYT7 轴驱动的 EVs 释放在氧化应激诱导的肿瘤转移中的重要作用。这些发现加深了人们对癌症进展分子基础的理解,并为治疗干预提供了潜在途径。
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引用次数: 0
An aptamer-guided fluorescence polarisation platform for extracellular vesicle liquid biopsy 用于细胞外囊泡液体活检的适配体引导荧光极化平台。
IF 15.5 1区 医学 Q1 CELL BIOLOGY Pub Date : 2024-09-02 DOI: 10.1002/jev2.12502
Cuong Viet Pham, Rocky Chowdhury, Shweta Patel, Satendra Kumar Jaysawal, Yingchu Hou, Huo Xu, Lee Jia, Yu-mei Zhang, Xiaowei Wang, Wei Duan, Dongxi Xiang

The translation of discoveries on extracellular vesicle (EV) based cancer biomarkers to personalised precision oncology requires the development of robust, sensitive and specific assays that are amenable to adoption in the clinical laboratory. Whilst a variety of elegant approaches for EV liquid biopsy have been developed, most of them remain as research prototypes due to the requirement of a high level of microfabrication and/or sophisticated instruments. Hence, this study is set to develop a simple DNA aptamer-enabled and fluorescence polarisation-based homogenous assay that eliminates the need to separate unbound detection ligands from the bound species for EV detection. High specificity is achieved by immobilising EVs with one set of antibodies and subsequently detecting them with a DNA aptamer targeting a distinct EV biomarker. This two-pronged strategy ensures the removal of most, if not all, non-EV substances in the input biofluids, including soluble proteins, protein aggregates or non-vesicular particles, prior to quantifying biomarker-positive EVs. A limit of detection of 5.0 × 106 EVs/mL was achieved with a linear quantification range of 5.0 × 108 to 2.0 × 1010 EVs/mL. Facilitated by a multiple parametric analysis strategy, this aptamer-guided fluorescence polarisation assay was capable of distinguishing EVs from three different types of solid cancer cells based on quantitative differences in the levels of the same sets of biomarkers on EVs. Given the simplicity of the method and its ease of implementation in automated clinical biochemistry analysers, this assay could be exploited for future EV-based continuous and real-time monitoring of the emergence of new macro- or micro-metastasis, cancer progression as well as the response to treatment throughout different stages of cancer management in the clinic.

要将基于细胞外囊泡(EV)的癌症生物标记物的发现转化为个性化的精准肿瘤学,就必须开发出稳健、灵敏和特异的检测方法,以便在临床实验室中应用。虽然目前已开发出多种优雅的 EV 液体活检方法,但由于对微细加工和/或精密仪器的要求较高,大多数方法仍停留在研究原型阶段。因此,本研究将开发一种简单的 DNA 配体和基于荧光偏振的均质检测方法,无需将未结合的检测配体与结合的物种分开,即可进行 EV 检测。先用一组抗体固定 EV,然后再用针对不同 EV 生物标记物的 DNA 类似物检测它们,从而实现高特异性。这种双管齐下的策略可确保在量化生物标记物阳性 EV 之前,去除输入生物流体中的大部分(如果不是全部)非 EV 物质,包括可溶性蛋白质、蛋白质聚集体或非囊泡颗粒。检测限为 5.0 × 106 EVs/mL,线性定量范围为 5.0 × 108 至 2.0 × 1010 EVs/mL。在多参数分析策略的帮助下,这种由适配体引导的荧光极化测定能够根据EVs上同一组生物标记物水平的定量差异,区分来自三种不同类型实体癌细胞的EVs。鉴于该方法简单且易于在自动临床生化分析仪中实施,该测定可用于未来基于EV的连续和实时监测,以监测新的大转移或微转移的出现、癌症的进展以及在临床癌症治疗的不同阶段对治疗的反应。
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Journal of Extracellular Vesicles
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