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评估感染黄病毒的活培养细胞中线粒体呼吸变化的实验方案。

Protocol to assess changes in mitochondrial respiration in living cultured cells infected with flaviviruses.

作者信息

Barragan Torres Viviana Andrea, Chatel-Chaix Laurent

机构信息

Centre Armand-Frappier Santé Biotechnologie, Institut National de la Recherche Scientifique, Laval, QC H7V 1B7, Canada.

出版信息

STAR Protoc. 2025 Aug 28;6(3):104051. doi: 10.1016/j.xpro.2025.104051.

DOI:10.1016/j.xpro.2025.104051
PMID:40880229
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC12410553/
Abstract

Mitochondria are essential organelles involved in energy production, making them prime targets for flaviviruses, such as dengue and Zika viruses, to enhance viral replication. Here, we present a protocol to measure multiple respiratory parameters in living hepatocarcinoma cells infected with flaviviruses in combination with pharmacological treatments or genetic expression modulation. We detail steps for infecting cells with flaviviruses and measuring the oxygen consumption rates using Seahorse technology. For complete details on the use and execution of this protocol, please refer to Freppel et al..

摘要

线粒体是参与能量产生的重要细胞器,这使其成为登革热和寨卡病毒等黄病毒增强病毒复制的主要目标。在此,我们提供了一个实验方案,用于测量感染黄病毒的活肝癌细胞中的多个呼吸参数,并结合药物治疗或基因表达调控。我们详细介绍了用黄病毒感染细胞以及使用海马技术测量氧消耗率的步骤。有关此实验方案的使用和执行的完整详细信息,请参考弗雷佩尔等人的研究。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/fccd1c73a197/gr6.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/4ce7382c767c/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/141ad0b17dca/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/5d957e8e1781/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/a99204ff021f/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/d1cdb767803e/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/7625bf1843b4/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/fccd1c73a197/gr6.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/4ce7382c767c/fx1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/141ad0b17dca/gr1.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/5d957e8e1781/gr2.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/a99204ff021f/gr3.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/d1cdb767803e/gr4.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/7625bf1843b4/gr5.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/8e3d/12410553/fccd1c73a197/gr6.jpg

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本文引用的文献

1
Dengue virus and Zika virus alter endoplasmic reticulum-mitochondria contact sites to regulate respiration and apoptosis.登革热病毒和寨卡病毒会改变内质网与线粒体的接触位点,以调节呼吸作用和细胞凋亡。
iScience. 2024 Dec 14;28(1):111599. doi: 10.1016/j.isci.2024.111599. eCollection 2025 Jan 17.
2
Extracellular flux assay (Seahorse assay): Diverse applications in metabolic research across biological disciplines.细胞外通量分析( Seahorse 分析):在跨生物学科的代谢研究中的多种应用。
Mol Cells. 2024 Aug;47(8):100095. doi: 10.1016/j.mocell.2024.100095. Epub 2024 Jul 18.
3
Zika virus modulates mitochondrial dynamics, mitophagy, and mitochondria-derived vesicles to facilitate viral replication in trophoblast cells.
寨卡病毒调节线粒体动力学、线粒体自噬和线粒体衍生小泡,以促进滋养层细胞中的病毒复制。
Front Immunol. 2023 Sep 14;14:1203645. doi: 10.3389/fimmu.2023.1203645. eCollection 2023.
4
A protocol to assess cellular bioenergetics in flavivirus-infected cells.评估黄病毒感染细胞细胞生物能量学的方案。
STAR Protoc. 2022 Apr 11;3(2):101297. doi: 10.1016/j.xpro.2022.101297. eCollection 2022 Jun 17.
5
Flaviviridae and mitochondria: Everything you always wanted to know about their relationship but were afraid to ask.黄病毒科与线粒体:你一直想了解但又不敢问的关于它们关系的一切。
Virologie (Montrouge). 2022 Feb 10. doi: 10.1684/vir.2022.0926.
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Dengue virus induces mitochondrial elongation through impairment of Drp1-triggered mitochondrial fission.登革病毒通过损害动力相关蛋白1(Drp1)引发的线粒体分裂来诱导线粒体延长。
Virology. 2017 Jan;500:149-160. doi: 10.1016/j.virol.2016.10.022. Epub 2016 Nov 4.
7
High-throughput screening using dengue virus reporter genomes.使用登革病毒报告基因基因组进行高通量筛选。
Methods Mol Biol. 2013;1030:205-19. doi: 10.1007/978-1-62703-484-5_17.