Real Time <em>In Vivo</em> Tracking of Thymocytes in the Anterior Chamber of the Eye by Laser Scanning Microscopy

Elisa Oltra; Alejandro Caicedo

doi:10.3791/58236

JoVE Journal
Biology

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JoVE Journal Biology

Real Time In Vivo Tracking of Thymocytes in the Anterior Chamber of the Eye by Laser Scanning Microscopy

生体内での実時間追跡レーザー顕微鏡による眼房内で胸腺細胞

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08:21 min

October 2, 2018

DOI: 10.3791/58236-v

Elisa Oltra^1,2, Alejandro Caicedo³

¹School of Medicine and Dentistry,Universidad Católica de Valencia San Vicente Mártir, ²Unidad Mixta CIPF-UCV,Centro de Investigación Príncipe Felipe, ³Department of Medicine,University of Miami

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Overview

This study demonstrates a protocol for longitudinal intravital real-time tracking of thymocytes using laser scanning microscopy in thymic implants within the anterior chamber of the mouse eye. This innovative approach allows for continuous monitoring of progenitor cell recruitment and the egress of mature T-cells, providing insights into the mechanisms of autoimmunity and immunodeficiency.

Key Study Components

Research Area

Immunology
Thymocyte behavior
In vivo microscopy techniques

Background

Importance of studying thymus function
Role of thymocytes in immune response
Mechanisms of autoimmunity and immunodeficiency

Methods Used

Longitudinal intravital imaging
Mouse models with thymic implants
Laser scanning microscopy

Main Results

Real-time visualization of thymocyte dynamics
Tracking of cell recruitment and egress
Insights into thymus physiology and its dependence on vascularization

Conclusions

This study highlights a novel method for observing thymus activity in a living organism.
It opens avenues for further research into immunological disorders and cell behavior in grafted tissues.

Frequently Asked Questions

What is the purpose of the thymic implant in this study?

The thymic implant allows researchers to observe the behavior of thymocytes in a living system, providing insights into their roles in immune responses.

How does laser scanning microscopy contribute to this research?

Laser scanning microscopy enables real-time visualization of cellular dynamics and interactions within the thymic graft, allowing for detailed analysis of immunological processes.

What are the implications of tracking T-cell egress?

Tracking T-cell egress is crucial for understanding the development of the immune system and the potential dysregulation in autoimmune diseases.

Can this technique be applied to other tissues?

Yes, the method can also be adapted for studying other tissues such as pancreatic islets and kidney glomeruli.

What is the significance of vascularization in this context?

Vascularization is important as it influences thymus physiology and is critical for the delivery of nutrients and immune factors necessary for thymocyte development.

What challenges are associated with thymus transplantation?

Challenges include ensuring proper engraftment, avoiding immune rejection, and maintaining tissue integrity during surgical procedures.

What advancements does this protocol represent in immunology research?

This protocol represents a significant advancement by enabling longitudinal studies of thymocyte behavior in a living organism, thereby providing unprecedented insights into immune system functioning.

プロトコルの目的は、レーザーマウス眼房内で胸腺のインプラントで顕微鏡による胸腺細胞の縦の生体リアルタイム追跡を示すことです。角膜の透明性と血管新生血管前駆細胞の動員と成熟した T 細胞の出口を継続的に記録が可能です。

この方法は、自己免疫、免疫不全、中央耐性、胸腺の不全のメカニズムに関する免疫学分野の主要な質問に答えることができます。この技術の主な利点は、血管化および移植されたマウス胸腺セグメント内の前駆細胞の動員および成熟したT細胞の出口のインビボ縦方向の記録を可能にすることです。この方法は、胸腺の発達および機能に関する洞察を提供することができるが、膵島または腎臓糸球体リなどの他の組織にも適用することができる。

胸腺を分離するには、薄層流れフードの後回りの直立位置に無菌吸収性ペーパータオルの上にマウスを置き、70%エタノールでマウス腹部をスプレーして拭きます。下腹部の表面的なV字型切開を行い、胸腔を露出させ、まっすぐな10センチメートルの解剖ハサミを使用して、腹側の正中線に沿って胸部に0.5〜1センチメートルの切開を行います。胸部の両側に皮膚を折り畳んで胸腔を露出させ、隔膜と肋骨ケージを通して2つの深い0.5〜1センチメートルの側面切開を行い、胸部前胸腔の優れた遠心部にアクセスします。

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