ANTIOXIDANT TREATMENT OF THYMIC ORGAN-CULTURES DECREASES NF-KAPPA-B AND TCF1(ALPHA) TRANSCRIPTION FACTOR ACTIVITIES AND INHIBITS ALPHA-BETA-T-CELL DEVELOPMENT

Using electrophoretic mobility shift assays (EMSA), we have recently s hown that nuclear extracts of 14-day mouse fetal thymocytes contain ab undant NF-kappaB transcription factor activity. To determine the funct ional role of NF-kappaB in early thymocyte development, we have expose d fetal thymus organ cultures to inhibitors of NF-kappaB activation, n amely the antioxidants N-acetyl-L-cysteine and butylated hydroxyanisol e. Both compounds caused a dose-dependent arrest of thymocyte differen tiation toward alphabeta, but not gammadelta, T cells. This was associ ated with a profound decrease in nuclear content of NF-kappaB and TCF1 (alpha) transcription factor activity, as determined by EMSA. In contr ast, NF-Y was affected less strongly, and cyclic AMP-response-element- binding protein levels remained essentially unchanged by antioxidants. To test the idea that alphabeta T cell development is correlated with NF-kappaB and TCF1(alpha) activity, we conducted additional experimen ts in a submersion culture system in which the generation of alphabeta T cells can be manipulated. Standard submersion culture supports gamm adelta but not alphabeta T cell development. Under these conditions, E MSA showed that transcription factor activities were similar to those seen in the presence of antioxidants. Importantly, when the generation of alphabeta T cells in submersion culture was restored by elevating oxygen concentrations, there was a dramatic increase in TCF1(alpha) ac tivity, and both NF-kappaB and NF-Y returned to control levels. Taken together, these results strongly suggest that NF-kappaB and TCF1(alpha ), presumably in concert with other transcription factors, play an imp ortant role in the development of alphabeta T cells.