CYTOTOXIC ACTIVITY OF TUMOR-NECROSIS-FACTOR-ALPHA MUTANTS - CORRELATIONS WITH FREE SPHINGOSINE LEVEL IN MURINE LIVER

Sphingosine, a highly cytotoxic product of enzymatic degradation of sp hingomyelin, is accumulated in organs of animals treated with tumor ne crosis factor ct (TNF-alpha). To determine the role of sphingosine in TNF cytotoxicity, TNF mutants differing in their toxicity to L929 cell s were produced and studied. Wild-type TNF and the mutant with deleted residues 67-71 exhibited the highest toxicity and caused a marked acc umulation of sphingosine in murine liver cells. TNF variants with sing le point mutations (E127Q, I155L, or V150I) and double mutations (V150 I plus I155L) caused moderate increases in sphingosine content and wer e significantly less toxic (compared to the wild type). The toxicity o f sphingosine, as well as its mutagenic and antimutagenic activities, were analyzed. Being highly cytotoxic, sphingosine lacked mutagenic ac tivity, but exerted antimutagenic effects in E. coli cells. The involv ement of products of enzymatic degradation of phospholipids in the act ivation of enzymes of the sphingomyelin cycle (and, consequently, in t he accumulation of sphingosine in cells of TNF-alpha-treated animals) is discussed.