Plasma requirement for the aggregation of rabbit platelets by an aggregating material derived from SV40-transformed...

Certain tumors require platelets for metastases, and many of these aggregate platelets in vitro. We have studied their in vitro interaction by extracting a PAM from SV40-transformed 3T3 fibroblasts. The preparation is enriched with membrane vesicles and requires an intact sedimentable sialolipoprotein for activity. PAM aggregates platelets after a lag period (J Lab Clin Med 93:332, 1979) and requires plasma as a cofactor(s). Two plasma components have been identified with the use of PRP or GFP. The first component shortens the platelet aggregation lag period after preincubation of PAM with plasma at 37 degrees C for 10 min prior to its addition to PRP or GFP and is labile to heating at 56 degrees C for 30 min. However, the activated PAM (formed by incubation with plasma at 37 degrees C) is stable at 56 degrees C for 30 min. This labile factor appears to be a component(s) of the complement alternative pathway, since it is inactivated by treatment of plasma with cobra venom or zymosan; and guinea pig PRP deficient in C' 4 can be aggregated by PAM. The second component is a plasma factor that is stable to heating at 56 degrees C. Activated PAM can be sedimented at 100,000 x g. The sediment, when suspended in Veronal buffer, pH 7.4, does not aggregate GFP, however, addition of plasma heated at 56 degrees C restores the platelet aggregation response. Thus a material extracted from SV40 3T3 fibroblasts aggregates platelets in vitro in the presence of two factors: (1) a component(s) of the alternative complement pathway that activates PAM and shortens the platelet aggregation lag period and (2) a heat-stable factor that is required for activated PAM to aggregate platelets.