An <em>In Vivo</em> Mouse Model to Measure Na&#239;ve CD4 T Cell Activation, Proliferation and Th1 Differentiation Induced by Bone Marrow-derived Dendritic Cells

Raquel Toribio-Fernandez; Virginia Zorita; Beatriz Herrero-Fernandez; Jose M. Gonzalez-Granado

doi:10.3791/58118

Un modelo de ratón In Vivo a medida ingenuo CD4 células T activación, proliferación y di...

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Immunology and Infection

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JoVE Journal Immunology and Infection

An In Vivo Mouse Model to Measure Naïve CD4 T Cell Activation, Proliferation and Th1 Differentiation Induced by Bone Marrow-derived Dendritic Cells

Un modelo de ratón In Vivo a medida ingenuo CD4 células T activación, proliferación y diferenciación de Th1 inducida por las células dendríticas derivadas de la médula ósea

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08:39 min

August 22, 2018

DOI: 10.3791/58118-v

Raquel Toribio-Fernandez¹, Virginia Zorita¹, Beatriz Herrero-Fernandez², Jose M. Gonzalez-Granado^1,2,3

¹LamImSys Lab,Centro Nacional de Investigaciones Cardiovasculares Carlos III (CNIC), ²LamImSys Lab,Instituto de Investigación Sanitaria Hospital 12 de Octubre (imas12), ³CIBER de Enfermedades Cardiovasculares

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Overview

This article presents a protocol for in vivo determination of na茂ve CD4 T cell activation, proliferation, and Th1 differentiation induced by GM-CSF bone marrow-derived dendritic cells. The protocol includes steps for bone marrow and T-cell isolation, dendritic cell generation, and adoptive transfer of both dendritic and T-cells.

Key Study Components

Area of Science

Immunology
T-cell biology
Dendritic cell function

Background

Understanding T-cell activation and differentiation is crucial for immunological research.
GM-CSF derived dendritic cells play a significant role in stimulating T-cell responses.
In vivo studies provide insights that are often not achievable through in vitro methods.
Adoptive transfer techniques are essential for studying immune responses in a controlled manner.

Purpose of Study

To establish a reliable protocol for studying T-cell activation and differentiation in vivo.
To facilitate the understanding of dendritic cell-T cell interactions.
To provide a visual demonstration of complex isolation and transfer techniques.

Methods Used

Isolation of bone marrow cells from adult mice.
Preparation of GM-CSF bone marrow-derived dendritic cells.
Adoptive transfer of dendritic cells and T-cells into recipient mice.
Assessment of T-cell activation and differentiation in vivo.

Main Results

The protocol successfully demonstrates T-cell activation and Th1 differentiation.
Visual aids significantly enhance understanding of complex procedures.
In vivo manipulation of T-cells allows for real-time observation of immune responses.
Findings contribute to the broader understanding of adaptive immunity.

Conclusions

This protocol is a valuable tool for researchers studying T-cell biology.
It bridges the gap between in vitro and in vivo studies in immunology.
Future applications may expand our understanding of immune system dynamics.

Frequently Asked Questions

What is the main advantage of this protocol?

The protocol allows for the study of T-cell activation and differentiation in an in vivo environment while enabling cell manipulation in vitro.

Why is visual demonstration important?

Visual demonstration is critical as the organ and bone isolation and adoptive transfer steps are difficult to master through text instructions alone.

What type of cells are isolated in this protocol?

The protocol involves the isolation of bone marrow cells and T-cells from adult mice.

How does GM-CSF influence dendritic cells?

GM-CSF is used to derive dendritic cells from bone marrow, which play a key role in T-cell activation.

What are the implications of this research?

The findings enhance our understanding of adaptive immunity and T-cell responses, which can inform therapeutic strategies.

Aquí, presentamos un protocolo para la determinación de en vivo de la activación de las células (células T) CD4 T ingenua, proliferación y diferenciación de Th1 inducida por el GM-CSF de la médula (BM)-derivado de las células dendríticas (DCs). Además, este protocolo describe aislamiento del BM y del T-cell, la generación de C.C. y transferencia adoptiva de DC y del T-cell.

Este método puede ayudar a responder preguntas clave en el campo de la inmunología sobre la activación, proliferación y diferenciación de las células T Th1, así como la estimulación de la inmunidad adaptativa de las células presentadoras de antígenos. La principal ventaja de esta técnica es que permite el estudio de un entorno in vivo a la vez que permite la manipulación celular in vitro. La demostración visual de este método es fundamental, ya que el aislamiento de órganos y huesos y los pasos de transferencia adoptivos son difíciles de dominar solo con instrucciones de texto.

Para el aislamiento de células de médula ósea, desinfecte las extremidades traseras de un ratón adulto con etanol al 70% y use tijeras para hacer una incisión lateral en la piel a través de las extremidades traseras. Inserte las tijeras en la cara medial de las patas y corte la piel con las tijeras hasta el comienzo de las extremidades traseras. Luego, use fórceps para separar la piel de las piernas.

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