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Biology
Isolamento de células endoteliais do lúmen de artérias carótidas de camundongos para experimentos...
Isolamento de células endoteliais do lúmen de artérias carótidas de camundongos para experimentos...
JoVE Journal
Biology
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JoVE Journal Biology
Isolation of Endothelial Cells from the Lumen of Mouse Carotid Arteries for Single-Cell Multi-Omics Experiments

Isolamento de células endoteliais do lúmen de artérias carótidas de camundongos para experimentos multi-ômicos unicelulares

Full Text
3,821 Views
09:12 min
October 4, 2021

DOI: 10.3791/63128-v

Sandeep Kumar*1, Aitor Andueza*1, Nicolas Villa-Roel1, Juyoung Kim1, Dong-Won Kang1, Hanjoong Jo1,2

1Wallace H. Coulter Department of Biomedical Engineering,Georgia Institute of Technology and Emory University, 2Division of Cardiology,Georgia Institute of Technology and Emory University

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Please note that some of the translations on this page are AI generated. Click here for the English version.

Overview

This study presents a novel protocol for isolating endothelial cells and nuclei from mouse carotid arteries under various flow conditions, enabling detailed examination of molecular pathways related to atherosclerosis. The method enhances the ability to perform single-cell omics experiments, facilitating the study of endothelial cell responses to disturbed blood flow.

Key Study Components

Research Area

  • Single-cell omics
  • Endothelial cell biology
  • Atherosclerosis research

Background

  • Endothelial cells are difficult to study due to their low abundance in blood vessels.
  • The protocol significantly improves the yield of endothelial enriched samples.
  • Understanding molecular pathways influenced by blood flow dynamics is crucial for therapeutic development.

Methods Used

  • Isolation of endothelial cells and nuclei from carotid arteries of mice.
  • In vivo experimentation with various flow conditions.
  • Utilization of enzymatic digestion and subsequent cell processing techniques.

Main Results

  • Successful extraction and analysis of single endothelial cells for gene expression profiling.
  • Identification of responses to both acute and chronic disturbed flow conditions.
  • Establishment of potential therapeutic targets for atherosclerosis.

Conclusions

  • This method demonstrates a robust approach for studying endothelial cells in a single-cell context.
  • Findings have significant implications for understanding vascular biology and developing treatments for vascular diseases.

Frequently Asked Questions

What is the significance of isolating endothelial cells?
Isolating endothelial cells allows for targeted studies of their functions and responses in various physiological and pathological environments.
How does disturbed flow affect endothelial cells?
Disturbed blood flow can lead to changes in gene expression that contribute to the development of vascular diseases, such as atherosclerosis.
What techniques are used in this study?
The study employs enzymatic digestion, centrifugation, and single-cell sequencing techniques to analyze isolated cells.
Can this method be applied to other vascular tissues?
Yes, the method can potentially be adapted to isolate endothelial cells from other blood vessels.
What are potential applications of this research?
The research can inform the development of new therapies targeting endothelial function in various cardiovascular conditions.
What are some challenges in carotid artery isolation?
Challenges include the meticulous dissection required to avoid damage and contamination during the isolation process.
What is the role of flow conditions in vascular biology?
Flow conditions are crucial in determining endothelial cell behavior, which influences vascular health and disease.

Apresentamos um método para isolar células e núcleos endoteliais do lúmen de artérias carótidas de camundongos expostas a condições de fluxo estáveis ou perturbadas para a realização de experimentos ômicos unicelulares.

Em um vaso sanguíneo, o número de células endoteliais é muito baixo, o que dificulta o estudo. Nosso protocolo elimina essa limitação e nos permite estudar novas vias moleculares e terapêuticas. Nosso protocolo nos permite obter amostras enriquecidas endoteliais e estudar como o fluxo perturbado agudo e crônico afeta esse tipo de célula.

A capacidade de estudar as células endoteliais em mais detalhes torna possível definir genes-alvo e, portanto, potenciais terapêuticas para a aterosclerose visando as células endoteliais. O isolamento da artéria carótida é complicado e requer paciência e experiência. Para começar, prepare o animal injetando 0,05 miligramas por quilograma de buprenorfina por via subcutânea.

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