Mesenchymal Stem Cell Regulation of Macrophage Phagocytosis; Quantitation and Imaging

Jodi F. Evans; Anthony E. Ricigliano; Anthony V. Morante; Emily Martinez; Darlisy Vargas; Jonah Thyagaraj

doi:10.3791/62729

Regulação de células-tronco mesenquimais da fagocitose macrófago; Quantitação e Imagem

JoVE Journal
Immunology and Infection

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JoVE Journal Immunology and Infection

Mesenchymal Stem Cell Regulation of Macrophage Phagocytosis; Quantitation and Imaging

Regulação de células-tronco mesenquimais da fagocitose macrófago; Quantitação e Imagem

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09:10 min

July 16, 2021

DOI: 10.3791/62729-v

Jodi F. Evans¹, Anthony E. Ricigliano¹, Anthony V. Morante¹, Emily Martinez¹, Darlisy Vargas¹, Jonah Thyagaraj¹

¹Molloy College

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Overview

This protocol outlines a method for quantifying and visualizing the regulation of macrophage phagocytosis by mesenchymal stem cells (MSCs) using pH-sensitive fluorescent particles. The technique aims to enhance understanding of MSCs' role in innate immunity.

Key Study Components

Area of Science

Cell Biology
Immunology
Stem Cell Research

Background

Mesenchymal stem cells (MSCs) can influence immune responses.
Macrophages play a crucial role in phagocytosis and innate immunity.
Understanding cell contact mechanisms is vital for therapeutic applications.
Fluorescent imaging can provide insights into cellular interactions.

Purpose of Study

To elucidate how MSCs regulate macrophage phagocytosis.
To develop a high-throughput method for analyzing cell interactions.
To apply the technique to various co-culture models.

Methods Used

Co-culture of MSCs and macrophages.
Use of pH-sensitive fluorescent bioparticles.
Fluorescence imaging to visualize phagocytosis.
Washing and quenching steps to ensure accurate results.

Main Results

Demonstrated the ability of MSCs to regulate macrophage activity.
Provided a reliable method for quantifying phagocytosis.
Showed the applicability of the technique to other phagocytes.
Highlighted the importance of cell contact in immune regulation.

Conclusions

The protocol offers a valuable tool for studying MSC and macrophage interactions.
Insights gained could inform therapeutic strategies targeting innate immunity.
Future applications may extend to other immune cell types.

Frequently Asked Questions

What is the significance of using pH-sensitive fluorescent particles?

They allow for visualization of phagocytosis specifically in acidic environments, enhancing the accuracy of the results.

Can this method be applied to other types of immune cells?

Yes, the technique can be adapted for use with neutrophils and other phagocytes.

What are the key steps in the co-culture method?

The key steps include co-culturing MSCs with macrophages, introducing fluorescent particles, and performing fluorescence imaging.

Who demonstrated the procedure in the study?

Ethan Chetkof, an undergraduate in the lab, demonstrated the procedure.

What are the potential applications of this research?

This research could inform therapeutic strategies for modulating immune responses in various diseases.

How does this study contribute to our understanding of innate immunity?

It sheds light on the mechanisms by which MSCs influence macrophage function, a key aspect of innate immunity.

Apresentado aqui é um protocolo para quantificar e produzir imagens dinâmicas de células-tronco mesenquimais (MSC) regulação mediada de fagocitose macrófago (MΦ) de partículas de levedura não opssonizada (zymosan) que são conjugadas a uma molécula fluorescente sensível ao pH.

Este método de cocultura ajudará a responder às principais perguntas em torno dos mecanismos de contato celular por trás da regulação de células-tronco mesenquimais da fagocitose de macrófago e, portanto, iluminará o papel do MSC na regulação da imunidade inata. Esta técnica pode ser aplicada em análises de alto rendimento. As biopartículas conjugadas aos corantes sensíveis ao pH fluorescem apenas em ambientes ácidas phagolysosome.

Por isso, lavar e saciar são necessários. Este método também pode ser aplicado a uma variedade de modelos de co-cultura que incluem neutrófilos e outros fagocitos. Demonstrando o procedimento será Ethan Chetkof, um estudante atualmente no meu laboratório.

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