Assessment and Quantification of Foam Cells and Lipid Droplet-Accumulating Microglia in Mouse Brain Tissue Using
Boaz K Maiyo1, Sanna H Loppi1, Helena W Morrison2
1Department of Immunobiology, University of Arizona, Tucson, AZ, USA.
Abstract:
This paper presents a refined, user-friendly protocol for using boron-dipyrromethene (BODIPY) to assess and quantify foam cells and lipid droplet-accumulating microglia (LDAM) in mouse brain tissue. The protocol aims to enhance existing methodologies by offering precise and efficient evaluation of foam cells and LDAM burden in various neuropathological conditions linked to lipid metabolism and neuroinflammation. A notable challenge in analyzing tissue from mouse models of these neurodegenerative disorders is the interference caused by the autofluorescent molecule lipofuscin. Our protocol addresses this issue with specific steps that effectively distinguish BODIPY fluorescence from lipofuscin autofluorescence, using advanced imaging techniques and filter settings to ensure accurate and reliable analysis. By providing a straightforward and accessible method, this research aims to facilitate the broader adoption of BODIPY-based techniques for detailed foam cell and LDAM analysis in mouse brain tissue, potentially enhancing diagnostic capabilities and deepening our understanding of how these cells contribute to neurodegenerative disease mechanisms. Key features • To induce foam cell/LDAM CNS formation, this protocol was developed using brain tissue from mice subjected to permanent occlusion of the middle cerebral artery. • The protocol utilizes mouse brain tissue that is fixed in 4% PFA. • Additional markers, CD68 and Iba1, are incorporated to evaluate myeloid cell lineage. • The protocol includes a simple method for distinguishing BODIPY fluorescence from autofluorescence.
Insights
This study introduces a user-friendly boron-dipyrromethene (BODIPY) protocol for quantifying foam cells and lipid droplet-accumulating microglia (LDAM) in mouse brains. It effectively distinguishes BODIPY signals from lipofuscin autofluorescence for accurate neuroinflammation analysis.
Area of Science:
- Neuroscience
- Immunology
- Biochemistry
Background:
- Foam cells and lipid droplet-accumulating microglia (LDAM) are implicated in neuroinflammation and neurodegenerative diseases.
- Accurate quantification of these cells in mouse models is crucial for understanding disease mechanisms.
- Lipofuscin autofluorescence in brain tissue complicates accurate analysis of lipid metabolism and cellular changes.
Purpose of the Study:
- To present a refined, user-friendly protocol for assessing and quantifying foam cells and LDAM in mouse brain tissue using boron-dipyrromethene (BODIPY).
- To enhance existing methodologies for precise and efficient evaluation of foam cell and LDAM burden in neuropathological conditions.
- To provide a method that effectively distinguishes BODIPY fluorescence from lipofuscin autofluorescence.
Main Methods:
- Utilized mouse brain tissue from a middle cerebral artery occlusion model to induce foam cell/LDAM formation.
- Employed 4% paraformaldehyde (PFA) fixation for brain tissue preparation.
- Incorporated CD68 and Iba1 markers to identify myeloid cell lineage.
- Developed specific imaging techniques and filter settings to differentiate BODIPY fluorescence from lipofuscin autofluorescence.
Main Results:
- A protocol was successfully developed and validated for BODIPY-based assessment of foam cells and LDAM.
- The protocol effectively overcomes the challenge of lipofuscin autofluorescence interference.
- The method allows for accurate and reliable quantification of these specific cell populations in mouse brain tissue.
Conclusions:
- The presented BODIPY protocol offers a straightforward and accessible method for foam cell and LDAM analysis in mouse brain tissue.
- This technique can facilitate broader adoption and enhance diagnostic capabilities in neurodegenerative disease research.
- Improved quantification of foam cells and LDAM will deepen understanding of their role in neuroinflammation and disease pathogenesis.
