Methods in cell biology最新文献_第4页

Bacterial predators and BALOs: Growth protocol and relation with mitochondria.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-08-24 DOI: 10.1016/bs.mcb.2024.07.003

Valerio Iebba

{"title":"Bacterial predators and BALOs: Growth protocol and relation with mitochondria.","authors":"Valerio Iebba","doi":"10.1016/bs.mcb.2024.07.003","DOIUrl":"https://doi.org/10.1016/bs.mcb.2024.07.003","url":null,"abstract":"The microbial world is characterized by mechanisms of competition and predation, akin to the animal world. However, while predation's ecological role is well-established in animals, it's less understood in bacteria due to fewer known predators and unclear phylogenetic affiliations. Nevertheless, microorganisms can prey on bacterial cells, including Bacteriophages, Protists, and Predatory Prokaryotes. These predators inhabit various habitats and may play vital roles in bacterial ecology and ecosystem regulation. Predatory interactions between host and parasite are common in nature. Predatory bacteria, such as Bdellovibrio and like organisms (BALOs), employ various strategies, including epibiotic predation and direct invasion. BALOs, which thrive in the periplasmic space of Gram-negative bacterial cells, modulate bacterial populations and could serve as preventive or therapeutic agents against Gram-negative infections. While primarily active against extracellular prey, BALOs may also target mitochondria, which are crucial for cellular processes. The relationship between intracellular bacteria and host mitochondria, including morphology, function, and apoptosis, warrants further exploration. Protocols for growing, propagating, and detecting predatory activities of BALOs, particularly Bdellovibrio bacteriovorus, are provided to assess their presence and activities against potential prey.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"194 ","pages":"151-167"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143586227","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

Monitoring the mitochondrial localization of mycobacterial proteins.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-11-18 DOI: 10.1016/bs.mcb.2024.10.018

Krishnaveni Mohareer, Jayashankar Medikonda, Sriram Yandrapally, Anushka Agarwal, Sharmistha Banerjee

引用次数: 0

Quantification of mitochondrial reactive oxygen species in macrophages during sepsis.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-02-24 DOI: 10.1016/bs.mcb.2024.01.005

Kanmani Suganya, Paulraj Kanmani, Guochang Hu

{"title":"Quantification of mitochondrial reactive oxygen species in macrophages during sepsis.","authors":"Kanmani Suganya, Paulraj Kanmani, Guochang Hu","doi":"10.1016/bs.mcb.2024.01.005","DOIUrl":"https://doi.org/10.1016/bs.mcb.2024.01.005","url":null,"abstract":"Sepsis is the leading causes of death globally, arising from an imbalanced host response to severe infection. It leads to multi-organ failure and poor outcomes in septic patients due to compromised glucose and lipid oxidation, reduced oxygen consumption, elevated levels of circulating substrates, and impaired mitochondrial function. Mitochondria, essential cellular organelles, play a vital role in regulating various cellular activities and the host immune response to infection. Pathogens, particularly bacteria, often disrupt mitochondrial functions to dysregulate host immunity. Additionally, the mitochondrial function is closely associated with most host immune responses, making mitochondria crucial in maintaining host homeostasis during infection. The intrinsic inflammatory response triggered by pathogens in sepsis impairs mitochondrial function, resulting in excessive production of mitochondrial reactive oxygen species (ROS) and subsequently damage to multiple organs. Here, we present a simple protocol for assessing mitochondrial ROS levels in bone marrow-derived macrophages (BMDMs) isolated from mice. We observed a higher level of ROS generation in lipopolysaccharide (LPS)-treated BMDMs, indicating the effectiveness and efficiency of our designed protocol for assessing mitochondrial ROS generation in vitro.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"194 ","pages":"59-75"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143586249","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

A screening system to determine the effect of bacterial metabolites on MAdCAM-1 expression by transformed endothelial sinusoidal cells.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-02-26 DOI: 10.1016/bs.mcb.2024.01.007

Ai-Ling Tian, Marion Leduc, Marine Fidelle, Laurence Zitvogel, Guido Kroemer, Oliver Kepp

{"title":"A screening system to determine the effect of bacterial metabolites on MAdCAM-1 expression by transformed endothelial sinusoidal cells.","authors":"Ai-Ling Tian, Marion Leduc, Marine Fidelle, Laurence Zitvogel, Guido Kroemer, Oliver Kepp","doi":"10.1016/bs.mcb.2024.01.007","DOIUrl":"https://doi.org/10.1016/bs.mcb.2024.01.007","url":null,"abstract":"Mucosal addressin cell adhesion molecule 1 (MAdCAM-1) expression in high endothelial venules is regulated by bacterial metabolites emanating from the gut and the interaction of MAdCAM-1 with α4β7 integrin mediates lymphocyte diapedesis into gut-associated secondary lymphoid tissues. MAdCAM-1 thus controls the abundance of circulating immunosuppressive T cells that can reach malignant tissue and compromise the therapeutic efficacy of anticancer immunotherapy. Here we describe a biosensor-based phenotypic assessment that facilitates the high throughput screening (HTS)-compatible assessment of MAdCAM-1 regulation in response to exposure to bacterial metabolites. This screening routine encompasses high endothelial venule cells expressing green fluorescent protein (GFP) under the control of the MAdCAM-1 promoter combined with robot-assisted bioimaging and a multistep image analysis pipeline. Altogether this system facilitates the discovery of bacterial composites that control anticancer immunity via the sequestration of Th17-specific regulatory T cells (Treg17) in the gut.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"194 ","pages":"119-133"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143586222","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

Monitoring cellular dynamics upon infection using a holotomography-based approach.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2025-01-10 DOI: 10.1016/bs.mcb.2024.12.003

Ilaria Nunzi, Gloria D'Achille, Nada Dhaouadi, Fabio Marcheggiani, Caterina Licini, Mariangela Di Vincenzo, Monia Orciani, Gianluca Morroni, Saverio Marchi

引用次数: 0

Visualizing mitochondrial electron transport chain complexes and super-complexes during infection of human macrophages with Legionella pneumophila.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-03-07 DOI: 10.1016/bs.mcb.2024.01.003

Mariatou Dramé, Daniel Schator, Carmen Buchrieser, Pedro Escoll

{"title":"Visualizing mitochondrial electron transport chain complexes and super-complexes during infection of human macrophages with Legionella pneumophila.","authors":"Mariatou Dramé, Daniel Schator, Carmen Buchrieser, Pedro Escoll","doi":"10.1016/bs.mcb.2024.01.003","DOIUrl":"https://doi.org/10.1016/bs.mcb.2024.01.003","url":null,"abstract":"The ultrastructure of mitochondria is pivotal for their respiratory activity. Thus, the regulation of the assembly of the super-complexes (SCs) of the mitochondrial electron transport chain (ETC) might be a core aspect of macrophage immunometabolism during bacterial infection. In order to study the impact of infection by Legionella pneumophila on the configuration of mitochondrial complexes and SCs in human macrophages, we have adapted and combined different methods such as cell sorting of infected cells, magnetic isolation of highly pure and functional mitochondria, quality control of mitochondrial purity by flow cytometry, and BN-PAGE (Blue-Native Polyacrylamide Gel Electrophoresis) coupled to Western Blot using near-infrared (NIR) fluorescence. The here presented protocol uses infected and non-infected human macrophage-like THP-1 cells and GFP-expressing L. pneumophila, but the method can be used to analyze the configuration of ETC complexes and SCs also in other mammalian cells and infected with different intracellular bacteria expressing a fluorescent protein.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"194 ","pages":"19-42"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143586259","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

Assessment of adhering and invading properties of Escherichia coli strains.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-09-20 DOI: 10.1016/bs.mcb.2024.08.011

Valerio Iebba

{"title":"Assessment of adhering and invading properties of Escherichia coli strains.","authors":"Valerio Iebba","doi":"10.1016/bs.mcb.2024.08.011","DOIUrl":"https://doi.org/10.1016/bs.mcb.2024.08.011","url":null,"abstract":"Gastrointestinal infections, caused by Enterobacteriaceae, pose a major global health challenge, resulting in significant morbidity and mortality. Enhanced adherence and invasion properties are widespread among enteric pathogenic species, particularly those linked to invasive infections such as some pathovars of Escherichia coli or pathogens like Shigella and Salmonella. Pathogenic E. coli strains are categorized into various pathotypes, including diarrheagenic E. coli (DEC) and extraintestinal pathogenic E. coli (ExPEC). Notably, Enteroinvasive E. coli (EIEC) and Adherent-invasive E. coli (AIEC) demonstrate significant invasive properties. EIEC, similar to Shigella, invades intestinal epithelial cells causing dysentery-like illness, while AIEC persists in the gut epithelium, potentially contributing to chronic inflammatory bowel diseases (IBD). Techniques like cell culture assays are vital for assessing E. coli's adherence and invasion capabilities, with specific virulence factors such as fimbriae and type III secretion systems (T3SS) playing crucial roles. Comparatively, Shigella and Salmonella also utilize T3SS for epithelial cell invasion, but with distinct effector proteins and mechanisms. Understanding these differences is crucial for diagnosis and treatment, as advanced molecular diagnostics improve the identification of invasive E. coli strains. Potential therapeutic interventions targeting fimbrial adherence, T3SS and effector proteins offer promising avenues for developing antivirulence drugs. Here are provided protocols for studying the adherence and invasion properties of E. coli and other Enterobacteriaceae to enhance diagnostic methods, ultimately improving the management of enteric infections.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"194 ","pages":"169-190"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143586224","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

Quantification of intracellular and mitochondrial ATP content in macrophages during lipopolysaccharide-induced inflammatory response.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-02-26 DOI: 10.1016/bs.mcb.2024.01.006

Paulraj Kanmani, Guochang Hu

{"title":"Quantification of intracellular and mitochondrial ATP content in macrophages during lipopolysaccharide-induced inflammatory response.","authors":"Paulraj Kanmani, Guochang Hu","doi":"10.1016/bs.mcb.2024.01.006","DOIUrl":"https://doi.org/10.1016/bs.mcb.2024.01.006","url":null,"abstract":"Sepsis, a condition characterized by systemic infection that becomes aggravated and dysregulated, is a significant cause of mortality in critically ill patients. Emerging evidence suggests that severe sepsis is often accompanied by alterations in cell metabolism, particularly mitochondrial dysfunction, resulting in multiorgan failure. Normally, metabolically active cells or tissues exhibit higher levels of mitochondrial turnover, respiration, and adenosine triphosphate (ATP) synthesis. However, during sepsis, these processes become overwhelmed or dysregulated, leading to impaired ATP production in mitochondria. Here, we present two straightforward protocols for quantifying ATP production from mitochondria in bone marrow-derived macrophages (BMDMs). Our workflow facilitates the easy isolation of BMDMs and mitochondria from BMDMs treated with lipopolysaccharide (LPS), the major cell wall component of Gram-negative bacteria. We quantified intracellular and mitochondrial ATP production in macrophages in vitro using this protocol. The results indicated a decrease in mitochondrial ATP content in BMDMs in response to LPS. With minimal adjustments, this method can be adapted for use with various human and mouse primary cells and cell lines.","PeriodicalId":18437,"journal":{"name":"Methods in cell biology","volume":"194 ","pages":"77-92"},"PeriodicalIF":0.0,"publicationDate":"2025-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"143586247","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

Gauging antigen recognition by human primary T-cells featuring orthotopically exchanged TCRs of choice.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-04-15 DOI: 10.1016/bs.mcb.2024.03.003

Vanessa Mühlgrabner, Angelika Plach, Johannes Holler, Judith Leitner, Peter Steinberger, Loïc Dupré, Janett Göhring, Johannes B Huppa

引用次数: 0

Isolation and characterization of primary NK cells and the enrichment of the KIR2DL1⁺ population.

4区生物学

Methods in cell biology Pub Date : 2025-01-01 Epub Date: 2024-03-02 DOI: 10.1016/bs.mcb.2023.09.001

Batel Sabag, Abhishek Puthenveetil, Mira Barda-Saad

引用次数: 0