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

Gerçek zamanlı timositleri mikroskobu tarama lazer göz ön odasında Vivo içinde Takibi

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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

Gerçek zamanlı timositleri mikroskobu tarama lazer göz ön odasında Vivo içinde Takibi

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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.

Lazer fare gözün ön odasında timik implantlar mikroskobu tarama timositleri boyuna intravital gerçek zamanlı izleme göstermek için protokol hedefidir. Kornea saydamlığını ve vaskülarizasyon greft sürekli progenitör hücre işe alım ve olgun T hücreli çıkış kaydetmek için izin verir.

Bu yöntem, immünoloji alanında otoimmünite, immün yetmezlik, merkezi tolerans ve timus involution mekanizmaları hakkında anahtar soruların cevapyardımcı olabilir. Bu tekniğin en büyük avantajı, vaskülarize ve aşılı fare timus segmentleri içinde ata alımı nın ve olgun T-hücre çıkışLarının in vivo longitudinal kaydına izin vermedir. Bu yöntem timus gelişimi ve fonksiyonu içine fikir sağlayabilir rağmen, aynı zamanda pankreas adacıkları veya böbrek glomeruli gibi diğer dokulara uygulanabilir.

Timus izole etmek için, bir laminar akış kaputve sprey ve% 70 etanol ile fare karın silin dorsal dik pozisyonda steril emici kağıt havlu üzerine fare yerleştirin. Torasik boşluğu ortaya çıkarmak için alt karın yüzeysel V şeklinde bir kesi yapmak ve ventral orta hat boyunca göğüs 0,5 ila bir santimetrelik kesi yapmak için düz 10 santimetrelik diseksiyon makası bir çift kullanın. Göğüs boşluğu ortaya çıkarmak için göğüs her iki tarafında deri katlayın ve ön torasik kavitede üstün mediasten erişmek için diyafram ve göğüs kafesi ile iki ek daha derin 0.5 ila bir santimetre lateral kesiler yapmak.

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