Interleukins 2 and 12 activate natural killer cytolytic responses of peripheral blood mononuclear cells from patients with head and neck squamous cell carcinoma.

OBJECTIVES: To examine the capacity of interleukin-2 (IL-2) and interleukin-12 (IL-12) to modulate the cytolytic activity of peripheral blood mononuclear cells against squamous cell carcinoma, and to determine whether peripheral blood mononuclear cells from healthy donors respond differently to IL-12 than do peripheral blood mononuclear cells from patients with head and neck squamous cell carcinoma. DESIGN: Case-comparison study of a consecutive sample of patients with head and neck squamous cell carcinoma who were scheduled to undergo surgical excision. PARTICIPANTS: The study included 10 patients with stage III or IV carcinoma matched with 10 volunteer blood donors. INTERVENTION: Isolated peripheral blood mononuclear cells from patients and volunteers were treated with IL-2, 10 U/mL and 100 U/mL; IL-12, 1 U/mL or 10 U/mL; or a combination of IL-2 and IL-12. RESULTS: The combination of IL-2 and IL-12 consistently produced the greatest activation cytolysis than either cytokine alone at all concentrations tested. This increased activity against a squamous cell carcinoma cell line was seen in lymphocytes from volunteers and patients. CONCLUSION: Our findings suggest a new treatment regimen for the patient with head and neck cancer that uses immunomodulation with a combination of cytokines.

Effect of the psychoactive metabolite of marijuana, delta 9-tetrahydrocannabinol (THC), on the synthesis of tumor necrosis factor by human large granular lymphocytes.

The natural killer cell (NK)/3polymorphonuclear neutrophil axis has recently been identified to be important in early defense against the opportunistic fungi, Candida albicans. Repression of this system is therefore likely to contribute to susceptibility to opportunistic infections. delta 9-Tetrahydrocannabinol (THC), an active constituent of marijuana, has been reported to be immunosuppressive at concentrations that exceed attainable plasma levels. In this report, we examine the possibility that human large granular lymphocytes (LGL) can be immunosuppressed by exposure to THC at physiologically relevant concentrations and probed two functions associated with LGL, i.e., cytokine production and tumoricidal activity. We find that these low levels of THC inhibit tumor necrosis factor-alpha (TNF) induction from LGL by C. albicans and are dependent upon THC dose (0.005-5.0 micrograms/ml) and length of exposure (0.05-3.0 hr). Northern blot analysis indicates that the downregulation of TNF production from LGL by THC resides at the mRNA level. Moreover, exposure of LGL to physiological THC concentrations (0.01-2.0 micrograms/ml) diminishes their cytolytic activity against K562 tumor cells.

The 19-kilodalton adenovirus E1B transforming protein inhibits programmed cell death and prevents cytolysis by tumor necrosis factor alpha.

The adenovirus E1A and E1B proteins are required for transformation of primary rodent cells. When expressed in the absence of the 19,000-dalton (19K) E1B protein, however, the E1A proteins are acutely cytotoxic and induce host cell chromosomal DNA fragmentation and cytolysis, analogous to cells undergoing programmed cell death (apoptosis). E1A alone can efficiently initiate the formation of foci which subsequently undergo abortive transformation whereby stimulation of cell growth is counteracted by continual cell death. Cell lines with an immortalized growth potential eventually arise with low frequency. Coexpression of the E1B 19K protein with E1A is sufficient to overcome abortive transformation to produce high-frequency transformation. Like E1A, the tumoricidal cytokine tumor necrosis factor alpha (TNF-alpha) evokes a programmed cell death response in many tumor cell lines by inducing DNA fragmentation and cytolysis. Expression of the E1B 19K protein by viral infection, by transient expression, or in transformed cells completely and specifically blocks this TNF-alpha-induced DNA fragmentation and cell death. Cosegregation of 19K protein transforming activity with protection from TNF-alpha-mediated cytolysis demonstrates that both activities are likely the consequence of the same function of the protein. Therefore, we propose that by suppressing an intrinsic cell death mechanism activated by TNF-alpha or E1A, the E1B 19K protein enhances the transforming activity of E1A and enables adenovirus to evade TNF-alpha-dependent immune surveillance.

Adenovirus E3 14.7K protein functions in the absence of other adenovirus proteins to protect transfected cells from tumor necrosis factor cytolysis.

A 14,700-kDa protein (14.7K) encoded by the E3 region of adenovirus has been shown to protect adenovirus-infected mouse C3HA cells from lysis by tumor necrosis factor (TNF) (L. R. Gooding, L. W. Elmore, A. E. Tollefson, H. A. Brady, and W. S. M. Wold, Cell 53:341-346, 1988). These infected cells are sensitized to TNF by expression of the adenovirus E1A proteins (P. Duerksen-Hughes, W. S. M. Wold, and L. R. Gooding, J. Immunol. 143:4193-4200, 1989). In this study we show that 14.7K suppresses TNF cytolysis independently of adenovirus infection. Mouse C3HA and C127 cells were transfected with the 14.7K gene controlled by the mouse metallothionein promoter, and permanent 14.7K-expressing cell lines were tested for sensitivity to TNF cytolysis. Transfected cells which were sensitized to TNF either by inhibitors of protein synthesis, microfilament-destabilizing agents, or adenovirus infection were found to be resistant to TNF cytolysis. Two monoclonal antibodies were isolated and used to quantitate 14.7K in transfected and infected cells. Enzyme-linked immunosorbent assay (ELISA) analysis with these monoclonal antibodies and 14.7K immunoblots showed that 14.7K expression can be induced with cadmium in C3HA and C127 transfectants. The 14.7K induction correlated with a dose-dependent decrease in sensitivity to TNF cytotoxicity. The 14.7K protein does not substantially alter cell surface TNF receptor numbers or affinity on C3HA mouse fibroblasts, as determined by Scatchard analysis of 125I-TNF binding. The 14.7K protein also does not alter TNF signal transduction in general, because TNF induction of cell surface class I major histocompatibility complex molecules on 14.7K transfectants was unmodified. Our findings indicate that the adenovirus 14.7K protein functions as a specific inhibitor of TNF cytolysis in the absence of other adenovirus proteins and thus is a unique tool to study the mechanism of TNF cytotoxicity.

Induction of the heat shock response protects cells from lysis by tumor necrosis factor.

A minority of transformed cell lines are directly susceptible to lysis by TNF, whereas many cells can be made sensitive to TNF by treatment with inhibitors of protein synthesis. Other groups have shown that exposure to TNF induces in many cells a transcription/translation dependent response that protects the cell from TNF lysis. Heat shock proteins are involved in protecting cells from the lethal affects of heat and other metabolic poisons. In this report, we test the possibility that heat shock proteins are also involved in protecting cells from lysis by TNF. We find that after induction of the cellular heat shock response by either heat or arsenite treatment, both spontaneously TNF-sensitive cells and those cells made sensitive by inhibition of protein synthesis are nearly completely protected from TNF cytolysis. The heat-treated cells retained most of their capacity to bind TNF, suggesting that heat shock functions at a postreceptor binding phase of the lytic process. Mouse C3HA fibroblasts are also made sensitive to TNF lysis by treatment with cytochalasin E. We have previously found that elicitation of the cell's TNF-protective response by exposure to TNF suppresses killing of C3HA by subsequent treatment with TNF plus cytochalasin E. In contrast, we report here that induction of the heat shock response did not provide significant protection to C3HA from killing by TNF in the presence of cytochalasin E. Thus, although induction of heat shock proteins does protect cells from TNF, they appear to act by a mechanism distinct from that elicited by TNF itself.