Apoptotic cells were shown to induce dendritic cell immune tolerance. We applied a proteomic approach to identify molecules that are secreted from apoptotic monocytes, and thus may mediate engulfment and immune suppression. Supernatants of monocytes undergoing apoptosis were collected and compared using sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and differentially expressed proteins were identified using tandem mass spectrometry. Thrombospondin-1 (TSP-1) and its cleaved 26-kDa heparin-binding domain (HBD) were identified. We show that TSP-1 is expressed upon induction of monocyte apoptosis in a caspase-dependent pattern and the HBD is cleaved by chymotrypsinlike serine protease. We further show that CD29, CD36, CD47, CD51, and CD91 simultaneously participate in engulfment induction and generation of an immature dendritic cell (iDC) tolerogenic and phagocytic state. We conclude that apoptotic cell TSP-1, and notably its HBD, creates a signalosome in iDCs to improve engulfment and to tolerate engulfed material prior to the interaction with apoptotic cells. IntroductionIn recent years, it has become apparent that upon induction of apoptosis, apoptotic cells play an active role in their own engulfment by signaling professional phagocytes and/or antigenpresenting cells, without triggering an inflammatory or autoimmune response. [1][2][3][4][5] This process seems to play an important role in homeostasis, resolution of inflammation, and peripheral tolerance induction. 4,[6][7][8] Apoptotic cells have been shown to signal the innate immune system in a variety of ways. "Eat me" signals on apoptotic cells serve as markers for phagocytes to specifically recognize these cells and subsequently ingest them. Such signals can appear on apoptotic cell membranes. Direct signals include alteration in cell surface phospholipid composition, 9 changes in cell surface glycoprotein expression, distinct alterations in cell surface charge, 10,11 or expression of specific molecules. 12 Alternatively, certain serum or phagocyte-derived proteins can opsonize an apoptotic cell surface and signal phagocytes to engulf the opsonized cells. 4,[13][14][15][16][17] Viable cells actively express "do not eat me" signals by restriction of phosphatydilserine to the inner leaflet of their membrane, or "stay away" signals using CD31 expression. 18 Recently, attention has been given not only to apoptotic cell membrane changes and phagocyte receptors, but also to the release of a membrane-derived phospholipid, lysophosphatidylcholine, which acts as a "find me" signal that is important for phagocytic cell recruitment. 19 Most of these mechanisms suggest efficient identification and clearance of cells undergoing apoptosis, with noninflammatory and nonautoimmune consequences.We decided to further explore whether apoptotic cells can actively express and secrete molecules that have a physiological significance for their own engulfment and for the environmental immune suppression. We examined whether apoptosis-induced immune suppress...

Background: The storage of red blood cells (RBC) is associated with impairment of their properties that can induce a circulatory risk to recipients. In a preceding study (2009), we reported that post-storage rejuvenation (RJ) of stored RBC (St-RBC) efficiently reduced the storage-induced RBC/endothelial cell interaction, while only partially reversing the level of intracellular Ca²⁺, reactive oxygen species, and surface phosphatidylserine. In the present study, we examined the RJ effectiveness in repairing St-RBC mechanical properties. Methods: RBC, stored in CPDA-1 without pre-storage leukoreduction, were subjected to post-storage RJ, and the deformability, osmotic fragility (OF), and mechanical fragility (MF) of the rejuvenated St-RBC (St-RBC_Rj) were compared to those of untreated St-RBC and of freshly-collected RBC (F-RBC). Results: 5-week storage considerably increased OF and MF, and reduced the deformability of St-RBC. All alterations were only partially (40-70%) reversed by RJ, depending on the extent of the damage: the greater the damage, the lesser the relative effect of RJ. Conclusion: The findings of the present and preceding studies suggest that different St-RBC properties are differentially reversed by RJ, implying that some of the changes occur during storage and are irreversible.

Poststorage RBC rejuvenation treatment is effective in reversing the storage-induced RBC/EC interaction. This provides further documentation for the potential clinical benefit of poststorage rejuvenation.

This study provides, for the first time in humans, direct evidence that the deformability of transfused PRBC is a potent effector of transfusion outcome. Currently, PRBC are supplied primarily by the first-in-first-out criteria, while their functionality is ignored. The testing of PRBC hemodynamic quality would introduce a new paradigm into blood banking, which would contribute substantially to improving transfusion therapy.

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Liver injury is exacerbated by the transfusion of stored blood, primarily due to the change in the rheological properties of RBC. This data call for clinical studies in patients undergoing liver resection or transplantation.

Background: Red blood cells (RBCs) undergo a natural aging process occurring in the blood circulation throughout the RBC lifespan or during routine cold storage in the blood bank. The aging of RBCs is associated with the elevation of mechanical fragility (MF) or osmotic fragility (OF) of RBCs, which can lead to cell lysis. The present study was undertaken to identify RBC properties that characterize their susceptibility to destruction under osmotic/mechanical stress. Methods: RBCs were isolated from freshly donated blood or units of packed RBCs (PRBCs) and suspended in albumin-supplemented phosphate-buffered saline (PBS). In addition, PRBCs were separated by filtration through a microsphere column into two fractions: enriched with rigid (R-fraction) and deformable (D-fraction) cells. The RBCs were subjected to determination of deformability, MF and OF, moreover, the level of cell surface phosphatidylserine (PS) and the stomatin level in isolated RBC membranes were measured. Results: In the RBC population, the cells that were susceptible to mechanical and osmotic stress were characterized by low deformability and increased level of surface PS. The OF/MF was higher in the R-fraction than in the D-fraction. Stomatin was depleted in destroyed cells and in the R-fraction. Conclusion: RBC deformability, the levels of surface PS, and membrane stomatin can be used as markers of RBC fragility.