Cellular source of human platelet secretory phospholipase A2.
Platelets are one source of the group II extracellular form of phospholipase A2 (sPLA2) which is involved in the amplification of local and systemic inflammation. Although sPLA2 protein has been described in human platelets, its presence in human megakaryocytes has not been yet established. We demonstrated in this study that the human erythroleukaemia (HEL) cell line, which has megakaryoblastic features, constitutively expresses sPLA2. Using an anti-rhsPLA2 monoclonal antibody (mAb BA11) and dot-blot detection, we showed that HEL cells and platelets release sPLA2 into incubation medium upon stimulation by thrombin. Similar results were obtained for sPLA2 activity detected by a spectrofluorescence assay. Enzymatic activity was abolished by mAb BA11 and by protamine. In both cell types, although released, the major part of sPLA2 remained in the cell pellet, and was probably adsorbed at non-specific membrane sites. Double labelling experiments using mAb BA11 and an anti-GPIIb antiserum revealed the presence of sPLA2 in human bone-marrow megakaryocytes. The use of reverse transcription-polymerase chain reaction conjugated with hybridization analysis demonstrated the presence of mRNA encoding for sPLA2 in platelets and HEL cells. Expression of sPLA2 in platelets and megakaryocytes at both transcriptional and post-translational levels strongly argues in favour of a megakaryocytic origin of platelet sPLA2 and rules out a role for endocytosis of the enzyme from plasma by circulating platelets.
Emadi
S
S
Unité de Pharmacologie Cellulaire, Unité Associée Institut Pasteur-INSERM U285, Paris, France.
Mirshahi
M
M
Elalamy
I
I
Nicolas
C
C
Vargaftig
B B
BB
Hatmi
M
M
D001792:Blood Platelets / Q000201:enzymology*; D006801:Humans; D015151:Immunoblotting; D004915:Leukemia, Erythroblastic, Acute / Q000378:metabolism*; D008533:Megakaryocytes / Q000378:metabolism*; D010741:Phospholipases A / Q000378:metabolism*; D054467:Phospholipases A2; D012333:RNA, Messenger / Q000378:metabolism; D014407:Tumor Cells, Cultured
D012333:RNA, Messenger; D010741:Phospholipases A; D054467:Phospholipases A2