Generation of tumor-specific cytotoxic T lymphocyte and prolongation of the survival of tumor-bearing mice using interleukin-18-secreting fibroblasts loaded with an epitope peptide.

There is currently much interest in generating cytotoxic T lymphocyte (CTL) responses against tumor antigens as a therapy for cancer. In this study mouse fibroblasts (H-2(b)) were genetically modified to express a costimulatory B7.1 and a mature interleukin (IL)-18, and then loaded with an ovalbumin (OVA) epitope (SIINFEKL, H-2K(b) restricted) as a model antigen, and tested for the induction of OVA-specific CTLs in C57BL/6 mice (H-2(b)). The genetically modified fibroblasts lacking either IL-18 or B7.1 were also constructed. Immunization with the IL-18/B7.1-transfected fibroblasts induced strong cytotoxic activities against OVA-expressing EL4 (EG7) tumor cells, but not against other H-2(b) tumor cells such as EL4, C1498, and B16F1 cells. The magnitude of the cytotoxic response in mice with the IL-18/B7.1-transfected fibroblasts was significantly higher than the response in mice immunized with any other cell constructs. CD8(+) T cells with OVA-specific cytotoxic activities were predominant in mice immunized with the IL-18/B7.1-transfected fibroblasts. Furthermore, treatment with the IL-18/B7.1-transfected fibroblasts significantly prolonged the survival period of EG7 tumor-bearing mice. Anti-tumor CTL immunity by the IL-18/B7.1-transfected fibroblasts could be induced without the help of host antigen-presenting cells (APCs) and NK1.1(+) cells, whereas partially decreased by the depletion of CD4(+) T cells at the inductive stage. These results support the ability of IL-18/B7.1 gene transfer to enhance the antigen-presenting capacity of fibroblasts for inducing antigen-specific CTL response.

Induction of persistent in vivo resistance to Mycobacterium avium infection in BALB/c mice injected with interleukin-18-secreting fibroblasts.

Interferon-gamma (IFN-gamma) is closely associated with the generation of cell-mediated immunity and resistance to intracellular parasites. Interleukin-18 (IL-18) is known to strongly induce IFN-gamma production by T cells and natural killer (NK) cells. To determine whether the paracrine secretion of IL-18 can efficiently stimulate the resistance to Mycobacterium avium complex (MAC) infection, 3T3 fibroblasts were stably transfected to secrete bioactive IL-18 and their effects on MAC infection were investigated in genetically susceptible BALB/c mice, compared with that of free recombinant IL-18. Immunization with IL-18-secreting fibroblasts (3T3/IL-18) during intranasal infection with MAC resulted in a significant decrease in bacterial load of lung during the entire 8-week observation period, while rIL-18 reduced the bacterial load at initial 1 week but not by 8 weeks postinfection. Immunization with the 3T3/IL-18 cells induced and maintained significantly higher levels of cytotoxic activity and nitric oxide production by lung cells than those of rIL-18 immunization. Furthermore, lung cells in mice injected with the 3T3/IL-18 cells showed persistent production of IFN-gamma throughout the 8-week period, suggesting that the 3T3/IL-18 cells induced the resistance to MAC infection via IFN-gamma production. This work suggests that IL-18-secreting fibroblasts may serve as a vehicle for paracrine secretion of IL-18 in immunotherapy of MAC infection.

Enhanced interleukin-4 production in CD4+ T cells and elevated immunoglobulin E levels in antigen-primed mice by bisphenol A and nonylphenol, endocrine disruptors: involvement of nuclear factor-AT and Ca2+.

Bisphenol A (BPA) and p-nonylphenol (NP) are representative endocrine disruptors (EDs) that may have adverse effects on human health. The influence of these compounds on allergic immune responses remains unclear. In this study, we have examined the effects of BPA and NP on production of interleukin-4 (IL-4), a pro-inflammatory cytokine closely associated with allergic immune responses. Both BPA and NP significantly enhanced IL-4 production in keyhole limpet haemocyanin (KLH)-primed CD4+ T cells in a concentration-dependent manner. Treatment with BPA or NP in vivo resulted in significant increase of IL-4 production in CD4+ T cells and of antigen-specific immunoglobulin E (IgE) levels in the sera of KLH-primed mice. Furthermore, BPA and NP enhanced the activation of IL-4 gene promoter in EL4 T cells transiently transfected with IL-4 promoter/reporter constructs, and the enhancing effect mapped to a region in the IL-4 promoter containing binding sites for nuclear factor (NF)-AT. Activation of T lymphocytes by phorbol 12-myristate 13-acetate/ionomycin resulted in markedly enhanced binding activities to the NF-AT site, which significantly increased upon addition of BPA or NP, as demonstrated by the electrophoretic mobility shift assay, indicating that the transcription factor NF-AT was involved in the enhancing effect of BPA and NP on IL-4 production. The enhancement of IL-4 production by BPA or NP was significantly reduced by nitrendipine, a blocker of Ca2+ influx, and by FK506, a calcineurin inhibitor. FK506 inhibited the NF-AT-DNA binding activity and IL-4 gene promoter activity enhanced by BPA or NP. These results represent the first report describing possible enhancement of allergic response by EDs through increasing IL-4 production in CD4+ T cells and antigen-specific IgE levels in the sera via the stimulation of Ca2+/calcineurin-dependent NF-AT activation.