OBJECTIVES: T cell immunity plays a critical role in host immune surveillance of tumour cell growth and metastatic spread. This study used small hairpin (sh)RNA-mediated gene silencing to target VTCN1 (B7-H4) expression in a nonsmall-cell lung cancer (NSCLC) cell line (A549) and evaluated the effects on T cell immune activity using an in vitro coculture system. METHODS: VTCN1-specific shRNA-expressing plasmid was transfected into A549 cells. Mock transfected and empty plasmid-transfected A549 cells served as controls. VTCN1 expression in A549 cells was determined by reverse transcription-polymerase chain reaction (RT-PCR) for VTCN1 mRNA and Western blotting for B7-H4 protein. Transfected A549 cells were cocultured with Jurkat cells. Jurkat cells were examined for proliferation, apoptosis, cell cycle distribution and intracellular cytokine mRNA and protein levels. RESULTS: VTCN1-specific shRNA efficiently knocked down VTCN1 mRNA and B7-H4 protein levels in A549 cells. This downregulation led to enhanced Jurkat cell proliferation, decreased apoptosis, stimulated cell cycle progression and elevated production of interferon-γ, interleukin (IL)-10 and IL-2. CONCLUSIONS: B7-H4 negatively regulates T cell-mediated antitumour immunity in NSCLC.
OBJECTIVES: T cell immunity plays a critical role in host immune surveillance of tumour cell growth and metastatic spread. This study used small hairpin (sh)RNA-mediated gene silencing to target VTCN1 (B7-H4) expression in a nonsmall-cell lung cancer (NSCLC) cell line (A549) and evaluated the effects on T cell immune activity using an in vitro coculture system. METHODS:VTCN1-specific shRNA-expressing plasmid was transfected into A549 cells. Mock transfected and empty plasmid-transfected A549 cells served as controls. VTCN1 expression in A549 cells was determined by reverse transcription-polymerase chain reaction (RT-PCR) for VTCN1 mRNA and Western blotting for B7-H4 protein. Transfected A549 cells were cocultured with Jurkat cells. Jurkat cells were examined for proliferation, apoptosis, cell cycle distribution and intracellular cytokine mRNA and protein levels. RESULTS:VTCN1-specific shRNA efficiently knocked down VTCN1 mRNA and B7-H4 protein levels in A549 cells. This downregulation led to enhanced Jurkat cell proliferation, decreased apoptosis, stimulated cell cycle progression and elevated production of interferon-γ, interleukin (IL)-10 and IL-2. CONCLUSIONS: B7-H4 negatively regulates T cell-mediated antitumour immunity in NSCLC.