Lactadherin binding and phosphatidylserine expression on cell surface-comparison with annexin A5.
BACKGROUND
Transbilayer movement of anionic phospholipids from the inner to the outer leaflet of the plasma membrane occurs during platelet activation, red cell senescence, and apoptosis. The anionic phospholipid-binding protein, annexin A5, has been used to detect the presence of phosphatidylserine on the outer leaflet of the cell membrane. Lactadherin, a glycoprotein secreted by macrophages, binds to phosphatidylserine on apoptotic cells and promote their clearance by macrophages.


METHODS
The authors isolated and labeled lactadherin and annexin A5 with FITC and compared their ability to detect phosphatidylserine expression by flow cytometry.


RESULTS
FITC-lactadherin induced greater shift in the histogram and a higher mean fluorescence intensity than FITC-annexin A5 when platelets were activated with thrombin (0.1 unit/mL) or Ca(2+) ionophore A23187 (1 microM). Similarly, lactadherin was more sensitive in detecting phosphatidylserine in red cells induced to express phosphatidylserine. Also, in HL 60 cells undergoing apoptosis, lactadherin detected phosphatidylserine expression earlier than annexin A5. In patients with disseminated intravascular coagulation, lactadherin detected phosphatidylserine-expressing platelets in most patients, whereas under similar conditions, FITC-annexin A5 could not.


CONCLUSIONS
The authors' studies show that FITC-lactadherin is a better probe than annexin A5 in detecting phosphatidylserine-expressing activated platelets, red cells, and apoptotic cells.
Dasgupta
Swapan K
SK
Department of Pathology and Medicine (Thrombosis Research), Baylor College of Medicine, Houston, Texas, USA.
Guchhait
Prasenjit
P
Thiagarajan
Perumal
P
D000818:Animals; D017304:Annexin A5 / Q000378:metabolism*; D000954:Antigens, Surface / Q000378:metabolism*; D017209:Apoptosis; D001792:Blood Platelets / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D004912:Erythrocytes / Q000378:metabolism; D016650:Fluorescein-5-isothiocyanate; D005786:Gene Expression Regulation; D018922:HL-60 Cells; D006801:Humans; D008565:Membrane Proteins / Q000378:metabolism*; D008894:Milk Proteins / Q000378:metabolism*; D010718:Phosphatidylserines / Q000378:metabolism*; D011485:Protein Binding
D017304:Annexin A5; D000954:Antigens, Surface; D008565:Membrane Proteins; D008894:Milk Proteins; D010718:Phosphatidylserines; D016650:Fluorescein-5-isothiocyanate; D002118:Calcium