Decoding the Role of Bacterial Outer Membrane Vesicle and Small RNAs in Host Inflammation

解读细菌外膜囊泡和小RNA在宿主炎症中的作用

• 批准号: 10291354
• 负责人: Leigh Greathouse
• 金额: $ 42万
• 依托单位: BAYLOR UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-06 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10291354
• 关键词: Acute; Affect; Anti-Inflammatory Agents; Bacteria; Bacteroides fragilis; Biological Markers; Cells; Chemicals; Chronic; Chronic Disease; Clinical; Colitis; Colon; Colon Carcinoma; Communication; Communication Tools; Country; Development; Diarrhea; Disease; Equilibrium; Escherichia coli; Genetic Transcription; Homeostasis; Human; Immune; Immune response; Immune system; In Vitro; Individual; Inflammation; Inflammatory; Inflammatory Response; Innate Immune Response; Knowledge; Life; Membrane; Messenger RNA; Microbe; Modeling; Molecular; Mus; Nucleic Acids; Pathogenicity; Pathway interactions; Pattern recognition receptor; Pharmaceutical Preparations; Play; Prevention; Process; Production; RNA; Receptor Cell; Research; Research Personnel; Resolution; Ribosomal RNA; Role; Signal Transduction; Small RNA; System; TLR7 gene; Toll-Like Receptor Pathway; Toll-like receptors; Transfection; Transfer RNA; Vesicle; Work; bioinformatics tool; chronic inflammatory disease; curative treatments; cytokine; exosome; extracellular; extracellular vesicles; gut microbiome; in silico; microbial; pathogen; pathogenic Escherichia coli; prevent; systemic inflammatory response; tool; trafficking; treatment response; undergraduate student; uptake

PROJECT SUMMARY Chronic inflammation is associated with most non-communicable diseases, which affect over 40% of individuals in the U.S.. While drugs are available that target suppression of inflammatory effector pathways, none target resolution of chronic inflammation, in part due to a lack of understanding of how to re-establish immune homeostasis. Bacteria in the colon play a key role in regulating the balance between acute and chronic inflammation, and understanding how to re-establish immune homeostasis is paramount in preventing chronic inflammatory disease development. Enterotoxigenic Bacteroides fragilis (ETBF) is representative of one species that is responsible for diarrhea, colitis and colon cancer. Yet, the commensal non-toxigenic strain of B. fragilis (NTBF) appears to prevent inflammation. Our research group and others have documented a major communication tool of bacteria are extracellular small RNAs, which can dampen or active the immune system. Little is known, however, about the contribution of sRNA species in governing microbe-host communication. Recently, bacterial small RNA (sRNA) species were identified within outer membrane vesicles (OMVs) that are shed by bacteria. Small RNAs represent a critical trigger governing whether host cells respond with pro- or anti-inflammatory signals. Intriguingly, OMVs from the commensal B. fragilis and E. coli prevent chemically induced murine colitis, while those from pathogenic E. coli induce systemic inflammation. Preliminary evidence from our lab indicate that OMVs from ETBF and NTBF differentially activate the TLR pathway, and their OMVs carry distinct sRNA cargo. Results from our preliminary research demonstrate that OMV-derived small RNAs are differentially abundant in OMVs from ETBF as compared to NTBF. We also show that OMVs from ETBF specifically activate TLR7 as compared to OMVs from NTBF. In this study, we hypothesize that the differential inflammatory response to OMVs from toxic and non-toxigenic Bacteroides fragilis is governed, in part, by sRNA engagement with TLRs, and that this mechanism partially explains the prevention or development of chronic inflammation. The purpose of this proposal is to determine if OMVs or OMV-derived sRNA from the pathogenic ETBF either prevent or induce chronic inflammation as compared to the commensal strain NTBF. Our specific aims will 1) Elucidate the differential activation of the innate immune response between commensal and enterotoxigenic OMVs from B. fragilis using an established tri-culture system, 2) Identify the mechanism of immune- stimulating intracellular uptake and sub-cellular localization from commensal vs. enterotoxigenic B. fragilis OMVs into host cells., and 3) Determine the commensal vs. enterotoxigenic B. fragilis small RNA host targets in silico using bioinformatics tools and validate in vitro using tri-culture and nanolipovesicle transfection. Confirmation of this hypothesis would resolve an outstanding gap in the field to explain how OMVs from commensal vs pathogenic strains induce or prevent inflammation.

项目总结 慢性炎症与大多数非传染性疾病有关，这些疾病影响到超过40%的 在美国的个人。虽然有针对炎症效应通路抑制的药物可用， 没有一个目标是解决慢性炎症，部分原因是缺乏对如何重建 免疫动态平衡。结肠中的细菌在调节急性和非急性之间的平衡方面起着关键作用。 慢性炎症，了解如何重建免疫稳态是至关重要的 预防慢性炎症性疾病的发展。产肠毒素脆弱类杆菌(ETBF)是 代表一种引起腹泻、结肠炎和结肠癌的物种。然而，这种共生关系 非产毒脆弱杆菌菌株(NTBF)似乎可以预防炎症。我们的研究小组和其他人 已经证明细菌的一个主要交流工具是细胞外小RNA，它可以抑制 或者激活免疫系统。然而，关于sna物种在治理中的贡献，人们知之甚少。 微生物与宿主的交流。最近，细菌小RNA(Srna)物种被鉴定在外源。 由细菌脱落的膜泡(OMV)。小RNA代表着一个关键的触发因素 宿主细胞是否以促炎或抗炎信号作出反应。有趣的是，来自共生的OMV 脆弱芽孢杆菌和大肠杆菌可预防化学诱导的小鼠结肠炎，而致病性大肠杆菌可诱导 全身炎症。我们实验室的初步证据表明，ETBF和NTBF的OMV 不同地激活TLR通路，它们的OMV携带不同的sRNA货物。结果来自我们的 初步研究表明，OMV来源的小RNA在OMV和OMV中不同地丰富 ETBF与NTBF的比较。我们还表明，ETBF的OMV与TLR7相比，可以特异性地激活TLR7 从NTBF到OMV。在这项研究中，我们假设OMVS的不同炎症反应 从有毒和无毒的脆弱类杆菌中分离出来的部分原因是sRNA与TLRs的接触，以及 这一机制部分解释了慢性炎症的预防或发展。目的 这项建议的目的是确定OMVS或OMV衍生的来自致病ETBF的sRNA是否可以防止 或引起慢性炎症，与共生菌株NTBF相比。我们的具体目标将是1) 阐明共生和肠毒素在先天免疫反应中的不同激活 脆弱芽孢杆菌OMVS利用已建立的三代培养体系，2)鉴定免疫机制-- 共生与肠毒素脆弱芽孢杆菌刺激细胞内摄取和亚细胞定位 OMVS进入宿主细胞，以及3)确定共系膜与肠毒素产生性脆弱杆菌小RNA宿主 利用生物信息学工具在硅胶中打靶，并使用三培养和纳米卵泡进行体外验证 转染法。这一假说的确认将解决该领域的一个悬而未决的空白，以解释 来自共生菌和致病株的OMVS可以诱导或预防炎症。

项目成果

Epigenetic memory acquired during long-term EMT induction governs the recovery to the epithelial state.

在长期 EMT 诱导过程中获得的表观遗传记忆控制着上皮状态的恢复。

• DOI: 10.1098/rsif.2022.0627
• 发表时间: 2023
• 期刊: Journal of the Royal Society, Interface
• 作者: Jain,Paras;Corbo,Sophia;Mohammad,Kulsoom;Sahoo,Sarthak;Ranganathan,Santhalakshmi;George,JasonT;Levine,Herbert;Taube,Joseph;Toneff,Michael;Jolly,MohitKumar
• 通讯作者: Jolly,MohitKumar

A preparation of bacterial outer membrane with osmium tetroxide and uranyl acetate co-stain enables improved structural determination by transmission electron microscopy.

用四氧化锇和乙酸双氧铀共染色制备细菌外膜，可以改进透射电子显微镜的结构测定。

• DOI: 10.1093/jmicro/dfad027
• 发表时间: 2023
• 期刊: Microscopy (Oxford, England)
• 作者: Sheikh,Aadil;Zechmann,Bernd;Sayes,ChristieM;Taube,JosephH;Greathouse,KLeigh
• 通讯作者: Greathouse,KLeigh