Resistance of activated human Th2 cells to NO-induced apoptosis is mediated by γ-glutamyltranspeptidase

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Abstract
Activation-induced death of inflammatory cells (AICD) has an important function in immune maintenance. Type 1 Th cells are known to be more susceptible to AICD than Th2 cells. In the current study we examined whether NO-induced apoptosis also preferentially eliminates Th1 cells over Th2 cells. Naive human Th lymphocytes (CD4+CD45RO–) were activated in vitro for 1 week in the presence of IL-12 plus anti-IL-4 or IL-4 plus anti-IL-12 to generate Th1- and Th2-polarized cultures respectively. Cultures were exposed to the NO donors Spermine-nonoate (Sper) and DPTA-nonoate to study NO-induced apoptosis. We found that NO preferentially induced apoptosis in Th1-polarized cells as demonstrated by Annexin staining in the presence of 10 μM Sper (70 ± 16 versus 23 ± 4.4% in Th2 cells P < 0.01) and by DioC6 staining (38 ± 10 versus 11 ± 5% in Th2 cells, P < 0.01). The mechanism of NO-induced apoptosis in Th1/Th2-polarized cells was distinct from AICD and Fas-induced apoptosis. Differential sensitivity between Th1- and Th2-polarized cultures originated at the level of intracellular glutathione (GSH) metabolism. GSH levels were higher in Th2 cells (1.6 ± 0.2-fold Th1, P < 0.01). High intracellular GSH in Th2-polarized cells did not account for reduced susceptibility to NO per se, since the inhibition of γ-glutamyltrans-peptidase (γ-GT), which is involved in GSH import, sensitized Th2 cells to NO-induced apoptosis without GSH depletion. Therefore, higher activity of γ-GT in Th2 cells (2.1 ± 0.4-fold Th1, P < 0.001) specifically protects Th2 cells against NO-induced apoptosis. Preferential NO-induced elimination of human Th1 cells at sites of inflammation may thus select Th2 cells and contribute to immune deviation.