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Label-Free Identification of Lymphocyte Subtypes Using Three-Dimensional Quantitative Phase Imaging and Machine Learning
Published on: November 19, 2018
Morphological profiling of human T and NK lymphocytes by high-content cell imaging
Yolla German1, Loan Vulliard2, Anton Kamnev3
1Toulouse Institute for Infectious and Inflammatory Diseases (INFINITy), INSERM UMR1291, CNRS UMR5051, Toulouse III Paul Sabatier University, Toulouse, France; Ludwig Boltzmann Institute for Rare and Undiagnosed Diseases (LBI-RUD), Vienna, Austria.
Insights
High-content imaging reveals how actin remodeling shapes the immunological synapse. This method differentiates roles of ARPC1B and WASP in T-cell responses and identifies defects in immunodeficient cells.
Area of Science:
- Immunology
- Cell Biology
- Biophysics
Background:
- The immunological synapse (IS) is crucial for lymphocyte activation and function.
- Systematic quantitative approaches are needed to monitor IS dynamics and lymphocyte responses.
- High-content imaging offers a powerful tool for detailed analysis of cellular structures.
Purpose of the Study:
- To demonstrate the applicability of high-content imaging for analyzing human T and NK cells.
- To develop a pipeline for unbiased morphological profiling of the IS.
- To investigate the roles of actin cytoskeleton remodeling in IS architecture and lytic granule positioning.
Main Methods:
- Application of high-content imaging to human T and NK cells.
- Development of a pipeline for unbiased morphological profiling.
- Analysis of CD8+ T cells from immunodeficient individuals to study ARPC1B and WASP functions.
Main Results:
- Distinct actin cytoskeleton remodeling patterns were identified, influencing IS architecture and lytic granule positioning.
- Morphological profiling distinguished the roles of ARPC1B and WASP in IS assembly.
- ARPC1B-deficient lymphocytes showed uncoupled lytic granule and F-actin distribution.
Conclusions:
- Morphological profiling using high-content imaging is a scalable approach to monitor lymphocyte responsiveness.
- This method can identify complex aspects of lymphocyte micro-architecture and function.
- The study provides insights into the molecular mechanisms governing IS formation and lymphocyte effector functions.
Abstract:
The immunological synapse is a complex structure that decodes stimulatory signals into adapted lymphocyte responses. It is a unique window to monitor lymphocyte activity because of development of systematic quantitative approaches. Here we demonstrate the applicability of high-content imaging to human T and natural killer (NK) cells and develop a pipeline for unbiased analysis of high-definition morphological profiles. Our approach reveals how distinct facets of actin cytoskeleton remodeling shape immunological synapse architecture and affect lytic granule positioning. Morphological profiling of CD8+ T cells from immunodeficient individuals allows discrimination of the roles of the ARP2/3 subunit ARPC1B and the ARP2/3 activator Wiskott-Aldrich syndrome protein (WASP) in immunological synapse assembly. Single-cell analysis further identifies uncoupling of lytic granules and F-actin radial distribution in ARPC1B-deficient lymphocytes. Our study provides a foundation for development of morphological profiling as a scalable approach to monitor primary lymphocyte responsiveness and to identify complex aspects of lymphocyte micro-architecture.
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