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Sphingomyelinase-induced adhesion of eryptotic erythrocytes to endothelial cells.
- Source :
-
American journal of physiology. Cell physiology [Am J Physiol Cell Physiol] 2012 Nov 01; Vol. 303 (9), pp. C991-9. Date of Electronic Publication: 2012 Sep 05. - Publication Year :
- 2012
-
Abstract
- Eryptosis, the suicidal erythrocyte death, leads to cell shrinkage and cell membrane scrambling with phosphatidylserine exposure at the cell surface. Eryptotic erythrocytes adhere to the vascular wall by binding of phosphatidylserine to the CXC chemokine ligand 16 (CXCL16). Stimulators of eryptosis include increased cytosolic Ca(2+) activity, energy depletion, and activation of ceramide-producing sphingomyelinase. The present study explored whether sphingomyelinase triggers erythrocyte adhesion to endothelial cells. To this end, human erythrocytes were exposed for 6 h to bacterial sphingomyelinase (1-10 mU/ml) and phosphatidylserine exposure was estimated from fluorescent annexin-V-binding, cell volume from forward scatter in FACS-analysis, erythrocyte adhesion to human umbilical vein endothelial cells (HUVEC) from trapping of labeled erythrocytes in a flow chamber under flow conditions at arterial shear rates, and CXCL16 protein abundance utilizing Western blotting and FACS analysis of fluorescent antibody binding. As a result, sphingomyelinase (≥1 mU/ml) triggered cell shrinkage, phosphatidylserine exposure and erythrocyte adhesion to HUVEC, effects blunted by Ca(2+) removal. Adhesion was significantly blunted by phosphatidylserine-coating annexin-V (5 μl/ml), following addition of neutralizing antibodies against endothelial CXCL16 (4 μg/ml) and following silencing of the CXCL16 gene with small interfering RNA. Pretreatment of HUVEC with sphingomyelinase upregulated CXCL16 protein abundance. Six hours pretreatment of HUVEC with sphingomyelinase (10 mU/ml) or C6-ceramide (50 μM) augmented erythrocyte adhesion following a 30-min treatment with Ca(2+) ionophore ionomycin (1 μM) or following energy depletion by 48-h glucose removal. Thus exposure to sphingomyelinase or C6-ceramide triggers eryptosis followed by phosphatidylserine- and CXCL16-sensitive adhesion of eryptotic erythrocytes to HUVEC.
- Subjects :
- Annexin A5 physiology
Antibodies, Neutralizing pharmacology
Apoptosis physiology
Calcium pharmacology
Calcium Ionophores pharmacology
Cell Adhesion drug effects
Cell Adhesion physiology
Cell Size
Cells, Cultured
Ceramides pharmacology
Chemokine CXCL16
Chemokines, CXC antagonists & inhibitors
Chemokines, CXC genetics
Chemokines, CXC physiology
Erythrocytes physiology
Gene Silencing
Glucose pharmacology
Human Umbilical Vein Endothelial Cells drug effects
Human Umbilical Vein Endothelial Cells physiology
Humans
Ionomycin pharmacology
Phosphatidylserines physiology
Receptors, Scavenger antagonists & inhibitors
Receptors, Scavenger genetics
Receptors, Scavenger physiology
Apoptosis drug effects
Erythrocytes drug effects
Sphingomyelin Phosphodiesterase pharmacology
Subjects
Details
- Language :
- English
- ISSN :
- 1522-1563
- Volume :
- 303
- Issue :
- 9
- Database :
- MEDLINE
- Journal :
- American journal of physiology. Cell physiology
- Publication Type :
- Academic Journal
- Accession number :
- 22954799
- Full Text :
- https://doi.org/10.1152/ajpcell.00239.2012