Vesicles generated during storage of red cells are rich in the lipid raft marker stomatin.
BACKGROUND
The release of vesicles by red blood cells (RBCs) occurs in vivo and in vitro under various conditions. Vesiculation also takes place during RBC storage and results in the accumulation of vesicles in RBC units. The membrane protein composition of the storage-associated vesicles has not been studied in detail. The characterization of the vesicular membrane might hint at the underlying mechanism of the storage-associated changes in general and the vesiculation process in particular.


METHODS
Vesicles from RBCs that had been stored for various periods were isolated and RBCs of the same RBC units were used to generate calcium-induced microvesicles. These two vesicle types were compared with respect to their size with atomic force microscopy, their raft protein content with detergent-resistant membrane (DRM) analysis, and their thrombogenic potential and activity with annexin V binding and thrombin generation, respectively.


RESULTS
The storage-associated vesicles and the calcium-induced microvesicles are similar in size, in thrombogenic activity, and in membrane protein composition. The major differences were the relative concentrations of the major integral DRM proteins. In storage-associated vesicles, stomatin is twofold enriched and flotillin-2 is threefold depleted.


CONCLUSIONS
These data indicate that a stomatin-specific, raft-based process is involved in storage-associated vesiculation. A model of the vesiculation process in RBCs is proposed considering the raft-stabilizing properties of stomatin, the low storage temperature favoring raft aggregation, and the previously reported storage-associated changes in the cytoskeletal organization.
Salzer
Ulrich
U
Department of Vascular Biology and Thrombosis Research, Medical University of Vienna, Vienna, Austria. ulrich.salzer@meduniwien.ac.at
Zhu
Rong
R
Luten
Marleen
M
Isobe
Hirotaka
H
Pastushenko
Vassili
V
Perkmann
Thomas
T
Hinterdorfer
Peter
P
Bosman
Giel J C G M
GJ
D000255:Adenosine Triphosphate / Q000378:metabolism; D000465:Algorithms; D001793:Blood Preservation / Q000379:methods; D022162:Cytoplasmic Vesicles / Q000378:metabolism*; D004912:Erythrocytes / Q000166:cytology / Q000378:metabolism*; D005434:Flow Cytometry; D005947:Glucose / Q000378:metabolism; D006801:Humans; D006863:Hydrogen-Ion Concentration; D021962:Membrane Microdomains / Q000378:metabolism*; D008565:Membrane Proteins / Q000378:metabolism*; D018625:Microscopy, Atomic Force; D012964:Sodium / Q000378:metabolism
D008565:Membrane Proteins; C036860:STOM protein, human; D000255:Adenosine Triphosphate; D012964:Sodium; D005947:Glucose