A SENSITIVE BIOIMMUNOASSAY FOR THROMBIN-CLEAVED 2-CHAIN UROKINASE-TYPE PLASMINOGEN-ACTIVATOR IN HUMAN-BODY FLUIDS

Thrombin cleaves single-chain urokinase-type plasminogen activator (sc u-PA) into a two-chain form (tcu-PA/T), which is virtually inactive in plasminogen activator assays. Little is known about the physiological importance of tcu-PA/T. To examine the occurrence of tcu-PA/T in vivo , we developed a sensitive and specific bioimmunoassay (BIA) for the a ssessment of tcu-PA/T in human body fluids. In this BIA, urokinase ant igen was immuno-immobilized in microtiter plates and treated with cath epsin C, a specific activator of tcu-PA/T, after which plasminogen act ivator activity was measured. The occurrence of tcu-PA/T was examined in the plasma of 27 healthy individuals acid of 17 sepsis patients, an d in the synovial fluid of 16 rheumatoid arthritis patients. In additi on, the concentration of urokinase antigen and scu-PA were measured in all three groups. In the plasma of the healthy individuals no measura ble amounts of tcu-PA/T could be found (< detection limit of 0.2 ng/ml ). In the plasma of almost all sepsis patients tcu-PA/T could be detec ted (median value 0.4 ng/ml). The amount of tcu-PA/T was 12% of the am ount of scu-PA and accounted for about 9% of urokinase antigen. In the synovial fluid of ail rheumatoid arthritis patients tcu-PA/T could be measured (median value 5.4 ng/ml) at a concentration which was twofol d higher than the concentration found for scu-PA. In this group tcu-PA /T contributed to about 47% of the urokinase antigen. From these data we conclude that inactivation of scu-PA by thrombin can take place in vivo under pathological conditions which involve the production of lar ge amounts of thrombin. This way thrombin may regulate fibrinolysis an d extracellular proteolysis. The BIA for tcu-PA/T can be of use for fu rther research on the physiological role of tcu-PA/T.