TYPE-1 PROPHOSPHOLIPASE A(2) PROPEPTIDE IMMUNOREACTIVITY IS RELEASED FROM ACTIVATED GRANULOCYTES

Objective: To establish a ELISA assay to measure release of type 1-pho spholipase A(2) propeptide from activated granulocytes. Human type 1-p rophospholipase A(2) (1-proPLA(2)) is biosynthesized and stored as ina ctive zymogen. Activation involves tryptic-like cleavage at the N-term inus, with equimolar release of the heptapeptide DSGISPR. Methods: Usi ng antibodies directed to the carboxyterminus of synthetic DSGISPR we developed a sensitive solid-phase ELISA specific for the released prop eptide that accurately reports the activation of 1-proPLA(2). The pres ence of the 1-proPLA(2) precursor itself can be determined by trypsini zation of the sample and subsequent assay for free DSGISPR. Results: U sing this ELISA, we demonstrated the presence of immunoreactive DSGISP R and its 14 kDa 1-proPLA(2)-like precursor in human granulocytes, but their absence in human macrophages and lymphocytes. Stimulation of cu ltured granulocytes with 1 pM of TNF alpha or GM-CSF caused rapid rele ase of DSGISPR and precursor into the surrounding medium. The immunore active signal coeluted with standard synthetic DSGISPR on G50 Sephadex chromatography. Conclusion: Release of DSGISPR immunoreactivity appea rs to be a specific consequence of granulocyte activation of potential relevance to the clinical pathophysiology of conditions like acute lu ng injury.