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The in vitro toxicity of the reactive oxygen species generating enzyme xanthine oxidoreductase (XOR) to human peripheral blood lymphocytes was studied after stimulation with phytohaemoagglutinin or anti-CD3/CD28 antibodies. Apoptosis and necrosis were induced by the XOR/hypoxanthine system in a time- and concentration-dependent manner. CD8+ lymphocytes showed a higher sensitivity than CD4+ cells to the XOR/hypoxanthine system. The occurrence of apoptosis was demonstrated by annexin-V binding to injured cell membrane, which was the most precocious alteration observed, followed by the increment of transglutaminase activity, which was significant at the lowest XOR concentration used. Nuclear damage was assessed by the increased hypodiploid nuclei and by DNA migration on gel electrophoresis, which turned to an apoptotic pattern before the occurrence of cell membrane necrotic lesions. Apoptosis was induced by XOR activity proportionally to substrate concentration and was prevented by the competitive enzyme inhibitor, allopurinol. The hydrogen peroxide scavenging enzyme, catalase, gave a higher protection than superoxide dismutase from the toxicity caused by the XOR/hypoxanthine system. Necrosis occurs in a variable percentage indicating that reactive oxygen species may trigger both apoptosis and necrosis in proliferating human lymphocytes, mostly depending on XOR concentration.

We assayed the cytotoxicity of conjugates in which the plant toxin saporin (SAP) was linked to human basic fibroblast growth factor (bFGF) and urokinase-type plasminogen activator (uPA). Both bFGF and uPA play an important role in angiogenesis, and their cell surface receptors are expressed at high levels in cancer cells. The conjugates were tested on a Kaposi's sarcoma-derived cell line, IST-KS2, as a model of an actively proliferating component of Kaposi's sarcoma lesions which are highly vascularized. Both bFGF and uPA were very effective at targeting saporin to KS cells, leading to cell killing even at low concentrations of the conjugates. The bFGF–SAP and uPA–SAP mitotoxins were also highly toxic toward the permanent, endothelium-derived EAhy926 cell line. EAhy926 cells were very sensitive to two immunotoxins containing an anti-endothelium monoclonal antibody coupled to saporin or ricin A chain, whereas IST-KS2 cells were not affected by these conjugates, in agreement with immuno cytochemical...