ABSTRACT
Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a promising and novel anticancer cytokine, specifically kills numerous tumor cells by apoptosis. However, some malignancies are resistant to TRAIL treatment in clinical trials, thus limiting its therapeutic potential. In the present study, the TRAIL-resistant murine hepatocellular carcinoma cell line Hepa1-6 was used to elucidate the physiological significance of TRAIL resistance, especially with respect to the immune regulatory function of TRAIL. Hepa1-6 cells were resistant to TRAIL-induced apoptosis in vitro; however, intratumoral injection of recombinant soluble TRAIL inhibited tumor growth and prolonged survival time in tumor-bearing mice. Local TRAIL treatment decreased the number of intratumoral CD4(+)CD25(+)Foxp3(+) regulatory T cells (Tregs) but did not affect CD4(+)CD25(+)Foxp3(+) Tregs in the draining lymph nodes and spleen. Further investigation showed that TRAIL induced apoptosis of tumor-activated CD4(+)CD25(+)Foxp3(+) Tregs, but not of CD4(+)CD25(-) T cells. Moreover, mouse TRAIL receptor DR5 expression was detected on the surface of the tumor-infiltrating CD4(+)CD25(+)Foxp3(+) Tregs, but not on naïve CD4(+)CD25(+)Foxp3(+) Tregs. Interestingly, intratumoral injection of TRAIL not only decreased the number of CD4(+)CD25(+)Foxp3(+) Tregs but also increased the number of tumor-specific CD8(+) CTL and augmented their cytotoxicity to the tumor cells. These data provide the novel evidence for an immune regulatory function of TRAIL and may shed light on the clinical application of TRAIL.
Subject(s)
Apoptosis/drug effects , Liver Neoplasms, Experimental/drug therapy , Liver Neoplasms, Experimental/immunology , T-Lymphocytes, Regulatory/drug effects , TNF-Related Apoptosis-Inducing Ligand/pharmacology , Animals , Cell Growth Processes/drug effects , Cell Line, Tumor , Female , Interleukin-2 Receptor alpha Subunit/immunology , Liver Neoplasms, Experimental/pathology , Mice , Mice, Inbred BALB C , Mice, Inbred C57BL , T-Lymphocytes, Cytotoxic/drug effects , T-Lymphocytes, Cytotoxic/immunology , T-Lymphocytes, Regulatory/immunology , T-Lymphocytes, Regulatory/pathology , TNF-Related Apoptosis-Inducing Ligand/immunologyABSTRACT
OBJECTIVE: Synovial cells in rheumatoid synovium display abnormal proliferation, which leads to joint destruction. TRAIL has been described as a proapoptotic factor in fibroblast-like synoviocytes (FLS). This study was undertaken to investigate the functions of rAAV2/5-TRAIL in human FLS and in arthritic mice. METHODS: Primary human FLS were infected with rAAV2/5-TRAIL in the presence or absence of epirubicin. Transgene expression was monitored by both enzyme-linked immunosorbent assay and flow cytometry. The disease-modulating activity of epirubicin plus rAAV2/5-TRAIL was investigated in mice with collagen-induced arthritis (CIA). RESULTS: Subtoxic doses of epirubicin potentiated rAAV2/5-mediated TRAIL expression in FLS and simultaneously enhanced the sensitivity of FLS to TRAIL. Epirubicin treatment up-regulated death receptor 4 (DR-4) and DR-5 expression and down-regulated FLIP expression, thereby enhancing the activation of procaspase 3, procaspase 8, and procaspase 9. An in vivo study showed that the combination of rAAV2/5-TRAIL gene therapy and epirubicin chemotherapy provided augmented antiarthritic effects in a mouse model of CIA. The intraarticular injection of rAAV2/5-TRAIL combined with epirubicin treatment significantly reduced the severity and incidence of CIA and joint swelling in the animals. Histologic evaluations revealed that inflammatory cell infiltration, cartilage destruction, and bone erosion were significantly reduced in the joints of the mice receiving the synthetic treatment. Results of a viral genome copy number assay indicated that epirubicin dramatically augmented the expression of rAAV2/5-TRAIL without altering its tissue distribution. CONCLUSION: These results suggest that epirubicin enhances the antiarthritic effect of rAAV2/5-TRAIL and that combination treatment might be an important therapeutic alternative, with practical significance for rheumatoid arthritis.
Subject(s)
Adenoviridae/genetics , Antibiotics, Antineoplastic/therapeutic use , Arthritis, Experimental/therapy , Epirubicin/pharmacology , Genetic Therapy/methods , Synovial Membrane/drug effects , TNF-Related Apoptosis-Inducing Ligand/genetics , Adenoviridae/classification , Adenoviridae/immunology , Adult , Animals , Antibodies, Viral/biosynthesis , Arthritis, Experimental/genetics , Arthritis, Experimental/pathology , Combined Modality Therapy , Disease Models, Animal , Female , Fibroblasts/drug effects , Fibroblasts/metabolism , Gene Expression/drug effects , Gene Targeting/methods , Genetic Vectors/therapeutic use , Humans , Male , Mice , Mice, Inbred DBA , Middle Aged , Recombinant Proteins , Synovial Membrane/metabolism , Synovial Membrane/pathology , TNF-Related Apoptosis-Inducing Ligand/metabolismABSTRACT
OBJECTIVE: To investigate the mechanism of anti-death receptor 5-10 (AD5-10) combined with epirubicin in treating rheumatoid arthritis (RA). METHODS: We detected the cell viability of the fibroblast-like synoviocytes (FLS) from RA patients with MTT. The expression level of apoptosis signaling pathways protein, p53, and p21 were evaluated with Western blot. RESULTS: We found that epirubicin, at different doses, could enhance the effect of AD5-10 on FLS, promoting the apoptosis of FLS. The expression levels of caspase-3, -8, -9, c-FLIP, Bcl-2, p53, and p21 in the FLS changed after epirubicin treatment. CONCLUSION: Epirubicin may coordinate with AD5-10 in inducing FLS apoptosis through affecting the levels of p53, p21, c-FLIP, and Bcl-2.
