Generation and cytotoxic profile of human peripheral blood CD4 + T lymphocytes

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Abstract
The effects of a variety of metabolic and anti‐tumour necrosis factor (TNF) antibodies were utilized to distinguish several different mechanisms of cytotoxicity employed by CD4+ effectors isolated from human peripheral blood lymphocytes (PBL). PBL, unseparated high buoyant density T cells and their CD4+ T cell subsets were activated with anti‐CD3 monoclonal antibody (MoAb) and interleukin‐2 (IL‐2) for 1‐5 days. CD4+ T cells activated with IL‐2/anti‐CD3 MoAb were cytotoxic when directed by a bispecific anti‐nitrophenyl (NP)‐anti‐CD3 MoAb heteroconjugate against both NP‐modified nucleated target cells (TC) and non‐nucleated sheep red blood cells (SPLBC). This CD4+ T population also lysed L929 in a TNF‐a dependent manner. Interestingly, different mechanisms of nucleated and non‐nucleated TC directed lysis by CD4+ effectors were implied by distinct patterns of sensitivity to cholera toxin (CT) and cyclosporin A (CsA). Cyclosporin A and CT inhibited CD4+ T cell directed lysis of SRBC, but not EL4. Cholera toxin, CsA or EGTA pretreatment also significantly inhibited the release of α‐N‐benzyloxycarbonyl‐l‐lysinethiobenzylester (BLT)‐esterase activity suggesting that degranulation of CD4+ effectors may be a critical step in their redirected lysis of SRBC. Overall, these findings suggested that activated human peripheral blood (PB) CD4+ effectors can lyse TC by at least three distinct mechanisms: (i) a CsA‐sensitive directed lysis of SRBC which correlates with exocytosis and presumably occurs via membrane lesions; (ii) a CsA‐insensitive directed lysis of NP‐modified nucleated TC that does not appear to involve exocytosis and is metabolically distinct; and (iii) a direct TNF‐dependent lysis of TNF‐sensitive TC. The highly prolifetative CD4+ T cell population could be propagated for at least 35 days while retaining cytotoxicity and secreting up to 80 U/mL of IL‐2. These data raise the possibility that anti‐CD3 MoAb plus IL‐2 activated CD4+ T cells may prove effective in adoptive tumour immunotherapy.