Increased TNF-alpha-induced apoptosis in lymphocytes from aged humans: Changes in TNF-alpha receptor expression and activation of caspases

Aging is characterized by increased T cell lymphopenia, T cell dysfunction, and increased serum TNF levels. In this study, we have examined the role o f TNF-induced apoptosis in T cell deficiency in lymphocytes from aged human s. The constitutive expression of TNF receptors (TNFRI and TNFRII) and the adapter molecules, including TNFR-associated death domain protein (TRADD), TNFR-associated factor 2 (TRAF-2), and receptor interacting protein (RIP), were analyzed both at the protein level by flow cytometry or Western blotti ng, and at the mRNA level using quantitative PCR or Northern blotting in ly mphocytes from aged and young subjects. The susceptibility of T cells to un dergo TNF-induced apoptosis was analyzed using terminal deoxynucleotidyltra nsferase-mediated UTP-end-labeling (TUNEL) and DNA ladder assays, Caspase ( caspase-8 and caspase-3) activation was compared between aged and young sub jects using Western blotting and colorimetric assays. In lymphocytes from a ged humans, there was an increased susceptibility of CD4(+) and CD8(+) T ce lls to undergo TNF-alpha-induced apoptosis, as observed by TUNEL assay and DNA fragmentation ladder assay, Increased TNF-alpha-induced apoptosis was a lso observed in both CD45RA(+) and CD45RO(+) T cells from aging subjects. A n increased constitutive expression of TNFRI and TRADD and decreased expres sion of TNFRII and TRAF-2 were observed in lymphocytes from aged as compare d with young controls. In addition, there was an early and increased activa tion of caspases (caspase-8 and caspase-3) involved in TNFR/TNF signaling p athway, as evident by early cleavage of caspase-8, poly(ADP-ribose) polymer ase (PARP), and caspase-3 substrate DEVD-p-nitroamilide NA. These data sugg est that an increased TNF-alpha-induced apoptosis may play a role in T cell deficiency associated with human aging.