apoptosis and monocytes

From: DARZYNKIEWICZ ZBIGNIEW (DARZYNK@nymc.edu)
Date: Sat Apr 22 2000 - 10:17:20 EST


Maciej Simm wrote:
>
> One of the people I'm working with right now is studying apoptosis in
> periteneal macrophages using a kit with annexin V monoclonal FITC ab
> and propidium iodine.
>
> The cells are autofluorescent. The "unstained" tube has signal in up
> to 2nd log.
>



Monocytes/macrophages may often be positive (by flow cytometry) for several
apoptosis markers for the reason that they may have ingested apoptotic
bodies detached from genuine apoptotic cells. This is particularly evident
e.g. during aggressive chemotherapy of leukemias when many leukemic and
perhaps normal lymphocytes die by apoptosis. During ingestion of apoptotic
bodies most likely the plasma membrane of the latter fuses with the
macrophages' plasma membrane which leads to exposure of phosphatidylserine
on the surface and annexin V positivity of macrophages. The ingested
apoptotic bodies have fragments of chromatin and thus multiplicity of DNA
strand breaks, which makes them also positive in the TUNEL assay (e.g.see
Bedner et al., Cytometry, 35: 181-195,1999). I am guessing that such
monocytes may also be positive for for other markers e.g. these that detect
activated caspases, cleavage of PARP, etc. The only way to identify such
"false positive" cells is microscopy or Laser Scanning Cytometry, which
allows one to relocate them and examine whether they are loaded with
apoptotic bodies.
Zbigniew Darzynkiewicz



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