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Assessing Retinal Microglial Phagocytic Function In Vivo Using a Flow Cytometry-based Assay
Published on: October 18, 2016
Assessing Retinal Microglial Phagocytic Function In Vivo Using a Flow Cytometry-based Assay
Salome Murinello1, Stacey K Moreno2, Matthew S Macauley3
1Department of Cell and Molecular Biology, The Scripps Research Institute; smuri@scripps.edu.
Abstract:
Microglia are the tissue resident macrophages of the central nervous system (CNS) and they perform a variety of functions that support CNS homeostasis, including phagocytosis of damaged synapses or cells, debris, and/or invading pathogens. Impaired phagocytic function has been implicated in the pathogenesis of diseases such as Alzheimer's and age-related macular degeneration, where amyloid-β plaque and drusen accumulate, respectively. Despite its importance, microglial phagocytosis has been challenging to assess in vivo. Here, we describe a simple, yet robust, technique for precisely monitoring and quantifying the in vivo phagocytic potential of retinal microglia. Previous methods have relied on immunohistochemical staining and imaging techniques. Our method uses flow cytometry to measure microglial uptake of fluorescently labeled particles after intravitreal delivery to the eye in live rodents. This method replaces conventional practices that involve laborious tissue sectioning, immunostaining, and imaging, allowing for more precise quantification of microglia phagocytic function in just under six hours. This procedure can also be adapted to test how various compounds alter microglial phagocytosis in physiological settings. While this technique was developed in the eye, its use is not limited to vision research.
Insights
Researchers developed a new flow cytometry method to easily measure microglial phagocytosis in vivo. This technique precisely quantifies microglial function, aiding research into neurodegenerative diseases.
Area of Science:
- Neuroscience
- Immunology
- Cell Biology
Background:
- Microglia are crucial for central nervous system (CNS) homeostasis, performing phagocytosis of cellular debris and pathogens.
- Impaired microglial phagocytosis is linked to neurodegenerative diseases like Alzheimer's and age-related macular degeneration.
- Assessing microglial phagocytic function in vivo has been historically challenging.
Purpose of the Study:
- To develop and validate a robust, in vivo method for quantifying microglial phagocytic potential.
- To offer a more efficient alternative to traditional, labor-intensive techniques.
Main Methods:
- Intravitreal delivery of fluorescently labeled particles into the eyes of live rodents.
- Quantification of particle uptake by retinal microglia using flow cytometry.
- Comparison with conventional immunohistochemical staining and imaging methods.
Main Results:
- The developed flow cytometry technique allows for precise and rapid (under six hours) measurement of microglial phagocytosis.
- This method significantly reduces the complexity and time associated with traditional approaches.
- The procedure is adaptable for assessing the impact of compounds on microglial phagocytosis.
Conclusions:
- This novel flow cytometry method provides a simple, robust, and efficient way to monitor and quantify in vivo microglial phagocytic function.
- The technique is applicable beyond vision research and can advance the study of CNS diseases.
- This method facilitates the investigation of therapeutic interventions targeting microglial phagocytosis.
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