Monocyte-mediated Tumoricidal Activity via the Tumor Necrosis Factor–related Cytokine, TRAIL

TRAIL (tumor necrosis factor [TNF]-related apoptosis-inducing ligand) is a molecule that displays potent antitumor activity against selected targets. The results presented here demonstrate that human monocytes rapidly express TRAIL, but not Fas ligand or TNF, after activation with interferon (IFN)-γ or -α and acquire the ability to kill tumor cells. Monocyte-mediated tumor cell apoptosis was TRAIL specific, as it could be inhibited with soluble TRAIL receptor. Moreover, IFN stimulation caused a concomitant loss of TRAIL receptor 2 expression, which coincides with monocyte acquisition of resistance to TRAIL-mediated apoptosis. These results define a novel mechanism of monocyte-induced cell cytotoxicity that requires TRAIL, and suggest that TRAIL is a key effector molecule in antitumor activity in vivo

T-cell clones from early-stage cutaneous T-cell lymphoma show no polarized Th-1 or Th-2 cytokine profile

Recent studies of the cytokine pattern in skin lesions of patients with cutaneous T-cell lymphoma (CTCL) have shown that interleukin-4 (IL-4) and Il-10, both cytokines produced by T-helper type 2 cells, dominate in these lesions. Also, in single studies, interferon-gamma (IFN-gamma), a major cytokine of Th-1-cells, has been found to be absent. Consequently, it has been hypothesized that immune-suppressive Th-2 cytokines may promote local growth of the malignant lymphocyte clone. However, there is so far no evidence for T-cells as the source of the Th-2 cytokines in CTCL skin lesions nor have these cytokines been investigated at a clonal T-cell level. We established a total of 120 T-cell clones (TCCs) from lesional skin and 54 TCCs from the blood of four patients with mycosis fungoides. Epidermal TCCs (mostly CD8-positive) and dermal TCCs (mostly CD4-positive) were stimulated by the mitogen concanavalin A and, seeking a polarized cytokine pattern, the supernatants were assessed by ELISA. We showed that the vast majority of TCCs were able to secrete IFN-gamma and IL-4. IFN-gamma-deficient TCCs occurred only in the epidermis. Some (18) TCCs were found to be either negative for IL-10 production or to produce low levels only. No significant differences were observed between blood- and skin-derived TCCs. Thus a polarized Th-2 cytokine pattern was not detectable among cultured skin-infiltrating nonmalignant T-cells (TILs) isolated from early mycosis fungoides. It therefore appears unlikely that Th-2-mediated immune suppression is a major mechanism operating in early CTCL. However, this does not exclude its role in late-stage disease

Human macrophages secrete a tumoricidal activity distinct from tumour necrosis factor-alpha and reactive nitrogen intermediates

Human macrophages, differentiated in vitro from blood monocytes, can be induced to secrete tumouricidal activity when activated by combined treatment with recombinant interferon gamma and bacterial lipopolysaccharide. We have analysed conditioned culture supernatants of activated human monocytes and in vitro differentiated macrophages cultivated under serum-free conditions for cytolytic activity against a TNF alpha-insensitive human tumour cell line and characterized this activity with respect to its relationship to TNF alpha and reactive nitrogen intermediates. Cytolytic activity was recovered in the high molecular weight fraction of culture supernatants conditioned by terminally differentiated macrophages, whereas conditioned culture supernatants of freshly isolated blood monocytes, processed under identical conditions, were devoid of significant cytolytic activity. This activity was tumour-specific, strongly affecting the human lymphoma cell line JMP, whereas freshly isolated human peripheral blood lymphocytes were not affected to a significant extent. It was inactivated by heat or trypsin treatment, but only partially inhibited by a monoclonal antibody against recombinant human TNF alpha, which completely neutralized all of the TNF alpha activity detectable in the supernatants tested. Cytolytic activity could not be reduced further even by a 1000-fold excess of anti-TNF alpha antibody, suggesting that TNF alpha has some synergistic effect on the tumouricidal activity observed, rather than being the central effector molecule. This notion was supported by enhancement of low levels of cytolytic activity by addition of recombinant human TNF alpha at concentrations not having any direct cytotoxic effect on the tumour target cells used.(ABSTRACT TRUNCATED AT 250 WORDS

Inability to culture the dominant T-cell clone from the skin of primary cutaneous T-cell lymphoma as proven by TCR gamma-chain gene sequencing

Molecular analysis of T-cell receptor (TCR) chain rearrangement has recently become an attractive tool for demonstrating the clonal origin of cutaneons T-cell lymphoma (CTCL) and for identifying the malignant clone at the molecular level. Over the past decade a number of attempts have been made to culture malignant CTCL cells using standard procedures and these attempts have resulted in several cell Lines from the peripheral blood of Sezary syndrome, mycosis fungoides and CD30(+) lymphoma patients. However, so far it has not been proven by sequence analysis that the cultured T cells truly represent the malignant cells. Aiming to functionally analyze the malignant T cells at a clonal level, we generated a total of 150 T-cell clones (TCC) from lesional skin and peripheral blood of three patients with mycosis fungoides and one patient with a CD30(+) lymphoma. Cells were grown either in the presence of autologous irradiated peripheral blood feeder cells using various conditions for T-cell stimulation by direct outgrowth or from skin specimens with various cytokine combinations. In order to identify the malignant TCC we used N-region-specific PCR and compared TCR gamma -chain sequences from clones of lesional skin with in vitro-generated TCC, With the methods employed, none of the 150 established cell lines was found to be identical to the malignant TCC which was readily detected in lesional skin, Our results indicate that standard cell culture methods are not suitable for growing low-grade CTCL cells from the skin but give rise only to benign infiltrating T cells