Encyclopedia of Experiments: Immunology
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Transcript
Take a multiwell plate with mesenchymal stem cells, or MSCs, in selected wells.
Introduce macrophages and incubate, facilitating MSC-macrophage interaction in the co-culture wells.
Add zymosan — a yeast cell wall component — conjugated to a pH-sensitive fluorophore to all the wells.
Macrophage scavenger receptors bind to zymosan, triggering its engulfment inside a phagosome, which fuses with a lysosome to form a phagolysosome.
An acidic pH inside the phagolysosome causes the fluorophore to emit fluorescence.
In the co-culture wells, MSCs project tunneling nanotubes, or TNT, cytoplasmic extensions that bridge with macrophages. MSC mitochondria travel through the TNT to enter the macrophages, increasing the total number of functional mitochondria.
An elevated mitochondrial number augments the production of adenosine triphosphate or ATP, which fuels the energy requirement for an increase in phagocytosis.
Record the fluorescence intensity over time from the internalized fluorophore-conjugated zymosans.
Co-culture shows increased fluorescence compared to wells with only macrophages, indicating MSC-mediated enhancement of macrophage phagocytosis.
