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Granzyme B (GrB) is a serine proteinase known to be expressed by cytotoxic lymphocytes and to induce, in presence of perforin (Pf), apoptosis in target cells. Recently, GrB expression has been shown (often in absence of Pf) in nonlymphoid cells, but its function is not defined. In our study, we investigated GrB and Pf expression in bladder cancer cell lines and in urothelial carcinoma (UC) tissues by reverse transcription-polymerase chain reaction (RT-PCR), Western blot, ELISA, immunofluorescence and immunohistochemistry. We also assessed the function of GrB in UC cells; the in vitro function of GrB was examined by loss-of-function experiments. Our results revealed that GrB is expressed, in absence of Pf, in UC cells. Significant differences were found between GrB expression and both increasing pathological tumor spreading and high-grade vs. low-grade pTa tumors. Notably, GrB in UC tissues was concentrated at the cancer invasion front and was expressed in neoplastic cells undergoing epithelial-mesenchymal transition, a key event in carcinoma invasion. Indeed, GrB-positive cells also expressed Snail, N-cadherin or were negative for E-cadherin. GrB expressed in tumor cell lines was enzymatically active and capable of vitronectin cleavage, implying extracellular matrix (ECM) remodeling by GrB. Inhibition of GrB activity or Stealth RNA interference-mediated GrB gene silencing markedly suppressed bladder cancer cell invasion through matrigel. This data provides the first evidence for a role of GrB in promoting cancer cell invasion. Taken together, our findings suggest that GrB, via ECM degradation, contributes to the establishment of the UC invasive phenotype. [ABSTRACT FROM AUTHOR]

Our aim was 2-fold: to investigate the role of alpha-tocopherol supplementation on the antitumor activity of DDP and to evaluate the effect of alpha-tocopherol on the survival and neurotoxicity of DDP-treated mice. Experiments performed on the M14 human melanoma line demonstrated that alpha-tocopherol supplementation did not influence the efficacy of DDP; the inhibition of cell survival and of the in vivo tumor growth after treatment with alpha-tocopherol and DDP combination was similar to that observed after DDP alone. Conversely, alpha-tocopherol was also able to increase survival of mice treated with a high dose of DDP. While DDP alone produced death in about 70% of mice, the combination reduced deaths to about 30%. Analysis of oxidative stress markers and peroxidative damage in organs indicated that the protective effect of alpha-tocopherol was mainly related to its antioxidant activity. A significant increase in the concentration of TBARS and decreased PUFAs and catalase activity were observed after DDP treatment, while with alpha-tocopherol the levels of these markers were comparable to those observed in untreated mice. Histologic analysis performed on peripheral nerve revealed that alpha-tocopherol also protected mice from severe neurologic damage induced by DDP treatment. In conclusion, our results demonstrate that alpha-tocopherol protects against the systemic toxicity and neurotoxicity induced by DDP without interfering with its antitumor activity and suggest that this combination is a promising strategy to improve the therapeutic index of DDP-based chemotherapy., (Copyright 2003 Wiley-Liss, Inc.)