Hemolysis of human red blood cells induced by the combination of diethyldithiocarbamate (DDC) and divalent metals: Modulation by anaerobiosis, certain antioxidants and oxidants

The objective of the present communication is to describe the role played b y combinations between diethydithiocarbamate (DDC) and divalent metals in h emolysis of human RBC. RBC which had been treated with DDC (10-50 mu M) wer e moderately hemolyzed (about 50%) upon the addition of subtoxic amounts of Cu2+ (50 mu M). However, a much stronger and a faster hemolysis occurred e ither if mixtures of RBC-DDC were immediately treated either by Co2+ (50 mu M) or by a premixture of Cu2+ and Co2+ (CU:CO) (50 mu M). While Fe2+ and Ni2+, at 50 mu M, initiated 30-50% hemolysis when combined w ith DDC (50 mu M), on a molar basis, Cd2+ was at least 50 fold more efficie nt than any of the other metals in the initiation of hemolysis by DDC. On t he other hand, neither Mn2+ nor Zn, had any hemolysis-initiating effects. C o2+ was the only metal which totally blocked hemolysis if added tb DDC prio r to the addition of the other metals. Hemolysis by mixtures of DDC +(Cu:Co) was strongly inhibited by anaerobiosi s (flushing with nitrogen gas), by the reducing agents glutathione, N-acety l cysteine, mercaptosuccinate, ascorbate, TEMPO, and N-tocopherol, by the P LA(2) inhibitor bromophenacylbromide (BrPACBr), by tetracycline as well as by phosphatidtyl choline, cholesterol and by trypan blue. However, TEMPO, B rPACBr and PC were the only agents which inhibited hemolysis induced by DDC : Cd2+ complexes. On the other hand, none of the classical scavengers of reactive oxygen spec ies (ROS) employed e.g dimethylthiomea, catalase, histidine, mannitol, sodi um benzoate, nor the metal chelators desferal and phenanthroline, had any a ppreciable inhibitory effects on hemolysis induced by DDC + (Cu:Co). DDC oxidized by H2O2 lost its capacity to act in concert either with Cu2+ o r with Cd-2 to hemolyze RBC. While either heating RBC to temperatures greater than 37 degrees C or expos ure of the cells to glucose-oxidase-generated peroxide diminished their sus ceptibility to hemolysis, exposure to the peroxyl radical from AAPH, enhanc ed hemolysis by DDC + (Gu:Co). The cyclovoltammetry patterns of DDC were drastically changed either by Cu2 +, Co2+ Or by Cd2 + suggesting a strong interaction of the metals with DDC. Also, while the absorbance spectrum of DDC at 280 nm was decreased by 50% either by CO2+ Cd2+ or by H2O2, a 90% reduction in absorbance occurred if D DC+H2O2 mixtures were treated either by Cu2+ or by Co2+ , but not by Cd2+. Taken together, it is suggested that DDC-metal chelates can induce hemolysi s by affecting the stability and the integrity of the RBC membrane, and pos sibly also of the cytoskeleton and the role played by reducing agents as in hibitors might be related to their ability to deplete oxygen which is also supported by the inhibitory effects of anaeobiosis.