Ricin at Subpicomole Concentrations Induce Cytolytic Activity and Stimulate Lymphokine Secretion by Human Peripheral Blood Mononuclear Cells.

Human PBMCs were stimulated by ricin at very low concentrations (10(-14) M and lower) for 48 h at 37 degrees C. Cytolytic activity to K562 cell line and lymphokine secretion were been measured. PBMCs possessed the highest cytotoxicity (63 +/- 2.8% and 64 +/- 3.1%, respectively) to K562 cell line in the presence of 10(-15) M or 10(-16) M ricin. The high secretion of IL-1beta (2000 pg/ml), IL-2 (1300 pg/ml) and TNF-alpha (2200 pg/ml) was detected in the presence of 10(-15) M ricin. The mechanisms of the target cell death induced by PBMCs after co-incubation with various lectins: 10(-8) M phytohemagglutinin, 10(-8) M concanavalin A and ricin at 10(-15) M or 10(-16) M concentrations were compared. Internucleosomal DNA fragmentation characteristic of apoptotic mechanisms of the target cell death in all cases were observed, and the maximum of DNA fragmentation was registered in the presence of 10(-15) M ricin.

Purification and Partial Characterization of a 14 kDa Protein Enhancing Mitogen-Stimulated Proliferation of Human Peripheral Blood Mononuclear Cells.

Cytotoxicity of plasma free platelets or peripheral blood mononuclear cells (PBMCs) upon stimulation with Ca-ionophore A23187 at various (10(-9) - 10(-6) M) concentrations has been studied on human erythroid leukaemia cell line (K562). A 14 kDa protein, which did not display any cytotoxic activity to K562 cells, but stimulate PBMC proliferation has been isolated from the supernatant of plasma free platelets stimulated by 10(-7) M Ca-ionophore. The protein displayed a very slight mitogenic effect on PBMCs, but it enhanced proliferative response of PBMC stimulated by concanavalin A at suboptimal concentrations. The p14 N-terminal sequence ((1)VLERTXA(7)-) is identical to the region 99-103 residues of the human MHC class II antigen DQ-beta chain. Also, we identified this 14 kDa protein in the supernatant of PBMCs stimulated by 10(-7) M Ca-ionophore. Its N-terminal sequence (VLERTXA-) is identical to the one of p14 from the stimulated plasma-free platelet supernatant.

Stimulation of Human Platelet Cytotoxicity by Various Activators in the Absence of Antibodies: Release of Thromboxane A2 and Soluble Cytotoxic Factors. Purification and Partial Characterisation of 14 and 28 kDa Cytotoxic Proteins.

The cytolytic activity of human plasma free platelets treated with various stimulators (Ca-ionophore, PAF (platelet activating factor), PHA, and ricin) has been studied on human erythroid leukemia cell line (K562). The 14 kDa and 28 kDa platelet cytotoxic factors (PCF) have been purified from the supernatant obtained from platelets stimulated by 10(-7) M Ca-ionophore. The PCF-14 N-terminal sequence ((1)YAPQXQFGP(9)-) appeared to be homologous to the region 241-249 residues of the human C1s complement component. The PCF-28 N-terminal sequence (DVGLT-) did not display any homology to known human proteins. The PCF-14 and the PCF-28 have been detected both in the supernatants of platelets treated with various stimulators (10(-7) M Ca-ionophore A23187, or 10(-8) M PAF, or 10(-9) M PHA, or 10(-13) M ricin) and in the extracts of disrupted platelets treated by these stimulators. All of these stimulators enhanced the production of thromboxane A(2) by platelets, as evidenced from the accumulation of thromboxane B(2), a spontaneous hydrolysis product of thromboxane A(2). These results indicate that platelet cytotoxicity to the K562 cells and the release of PCF-14 and PCF-28 might be mediated by the stimulation of thromboxane A(2) synthesis, when platelets have been activated through the cycloxygenase pathway.