Apoptotic Programs Are Determined during Lineage Commitment of CD4+ T Effectors: Selective Regulation of T Effector-Memory Apoptosis by Inducible Nitric Oxide Synthase

Abstract

Lineage-committed T effectors generated in response to Ag during the inflammatory phase are destined to die during termination of the immune response. We present evidence to suggest that molecular signatures of lineage commitment are reflected in apoptotic cascades activated in CD4+ T effectors. Exemplifying this, ablation of inducible NO synthase (iNOS) protected effector-memory T (TEM) cells, but not TNaive or central-memory T cells, activated in vitro, from apoptosis triggered by cytokine deprivation. Furthermore, attrition of T effectors generated in the secondary, but not the primary, response to Ag was substantially reduced in mice, which received iNOS inhibitors. Distinct patterns of iNOS expression were revealed in wild-type TEM effectors undergoing apoptosis, and ablation of iNOS protein in primary and TEM wild-type effectors confirmed observations made in iNOS−/− cells. Describing molecular correlates of this dependence, mitochondrial damage, activation of the protein Bax, and release from mitochondria of the apoptosis-inducing factor were selectively abrogated in iNOS−/− TEM effectors. Suggesting that iNOS dependence was linked to the functional identity of T cell subsets, both iNOS induction and apoptosis were compromised in IFN-γ−/− TEM effectors, which mirrored the response patterns of iNOS−/− TEM. Collectively, these observations suggest that programs regulating deletion and differentiation are closely integrated and likely encoded during lineage commitment of T effectors.

The Linker Histone H1.2 Is an Intermediate in the Apoptotic Response to Cytokine Deprivation in T-Effectors

“…T-cells proliferate and differentiate in response to antigen, to generate lineage-committed effectors, the bulk of which die, marking termination of the immune response [ 14 ]. Key elements of this process can be recapitulated in vitro , permitting investigations into the molecular regulation of T-cell apoptosis [ 16 – 18 , 20 , 21 ]. Using this experimental system we show that cytokine withdrawal from T-effectors triggers apoptotic damage characterized by nuclear fragmentation and externalization of phosphatidylserine (PS) at the cell membrane (Figures 1(a) and 1(b) ).…”

“…Loss of mitochondrial integrity is another feature of cells undergoing apoptotic duress and is reflected in the spatial redistribution of the flavoprotein, AIF (apoptosis inducing factor) [ 22 ]. AIF is a mitochondrial intermembrane space resident protein, which is released from mitochondria as a consequence of loss of outer mitochondrial membrane integrity and translocates to the nucleus to mediate DNA damage [ 21 , 23 ]. In T-cells cultured in cytokine assessed prior to the onset of the deprivation protocol (T0), AIF is detected in the cytoplasmic fraction consistent with its localization to mitochondria ( Figure 1(c) ).…”

“…10 × 10 6 T-effectors were used for the subcellular fractionation protocol [ 21 ]. Cytoplasmic and nuclear fractions were obtained using the NE-PER nuclear and cytoplasmic extraction kit (Thermo Scientific) following manufacturer's instructions.…”

Investigation of oxidative stress in patients with multifocal motor neuropathy

“…NOS would help limit immune response and contribute to the resolution of inflammation. Various functions of NOS include, among others, the downregulation of leukocyte recruitment 23,24 ; the blockage of leukocyte adhesion and transendothelial migration 25,26 ; the inhibition of T cell expansion in lymph nodes, the restriction of Th1 responses, the stimulation of apoptotic cell death in myeloid and lymphoid cells (e.g., effector memory T cells) 26,27 .…”