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

激光扫描显微术实时跟踪眼球前室胸腺细胞

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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.

该协议的目的是通过激光扫描显微镜在小鼠眼前腔的胸腺植入物中显示胸腺细胞的纵向活体实时跟踪。角膜的透明度和移植的血管化允许持续记录祖细胞的招募和成熟的 T 细胞出口。

该方法有助于回答免疫学领域有关自身免疫、免疫缺陷、中央耐受性和胸腺介入机制的关键问题。该技术的主要优点是，它允许在体内纵向记录祖主招募和成熟的T细胞出口血管化和移植小鼠胸腺段。虽然这种方法可以提供对胸腺发育和功能的洞察，也可以应用于其他组织，如胰岛或肾球蛋白。

要分离胸腺，请将鼠标放在无菌吸水纸巾上，放在层流罩的后侧直立位置，然后用 70% 乙醇喷洒和擦拭小鼠腹部。在下腹部做一个表面的V形切口，以暴露胸腔，并使用一对直10厘米解剖剪刀，使一个0.5至1厘米的切口在胸部沿腹中线。折叠胸部两侧的皮肤，露出胸腔，并通过隔膜和肋骨笼再进行两个更深的 0.5 至 1 厘米的侧切口，以进入前胸腔的优越介质。

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