Among sexually transmitted diseases, infection by human papillomavirus (HPV) has become one of the most important. On the other hand, though epidemiological data show that some HPV types are closely associated with cervical cancer, few reports have been found with reference to penile carcinoma because of its rare occurrence. The aim of this study was to investigate the relationship between HPV infection and penile cancer in Argentina. A retrospective study was carried out on 38 white men with penile squamous-cell carcinoma. Sixty-five archival fixed biopsies taken from 34 primary penile tumors, 25 nodal metastases, 1 skin "satellite" metastasis and 5 histologically normal lymph nodes were used as specimens. HPV detection and typing were carried out by the polymerase chain reaction (PCR) using generic primers, combined with single-stranded conformational polymorphism (SSCP) analysis. HPV DNA was found in 71% patients, corresponding 81% of them to "high risk" types, with predominance of HPV 18. Both primary tumors and metastases showed concordance of HPV occurrence and type in both lesions. In 3 patients, HPV 16 was detected not only in primary tumors and metastases, but also in histologically normal lymph nodes. Our data indicate that most penile carcinomas in Argentine patients are etiologically related to HPV, especially to "high risk" genital types. The agreement in HPV detection between primary tumors and metastases suggests a potential viral role in tumor progression. HPV detection in otherwise histologically normal lymph nodes might be useful as early marker of a metastatic process.

The new murine bladder model described resembles human bladder disease, making it a useful tool for studying the molecular mechanisms of tumor progression and metastasis, and assaying antimetastatic and anti-invasive agents.

Bacillus Calmette-Guérin (BCG) is the most effective treatment for superficial and in situ transitional bladder cancer. Although the complete mechanisms for its effect are not fully understood yet, both immunological and direct effects on tumor cells have been proposed. It has been proposed that apoptotic tumor cells could be better inducers of immunity than necrotic ones. Thus, apoptosis of bladder cancer cells could contribute to a global response to BCG. Lysosomal hydrolase cathepsin B (CB) is involved in the apoptotic process and has a key role in breast cancer cell programmed death through the activation of a pro-apoptotic protein BID. Truncated BID participates in the mitochondrial apoptotic pathway that involves the activation of pro-caspase 9. The possibility that CB can be involved in apoptosis of TCC line has not been explored yet. Therefore, we analyzed the participation of CB in BCG-induced apoptosis of human and murine TCC lines. Apoptosis was evaluated by a morphologic assay and CB activity by a substrate-specific colorimetric method. Expression of CB, BID and pro-caspase 9 was determined by Western blotting. BCG induced apoptosis of murine (MBT2, MB49) and human (T24) TCC lines. An increase in both CB activity and protein was also observed. The apoptosis of T24 and MB49 cell lines was mediated by activation of pro-caspase 9 and BID, both proteins are involved in mitochondrial apoptosis. Apoptosis and activation of pro-caspase 9 and BID were inhibited by CA-074Me (CA), a cell permeable CB inhibitor. Thus, CB is involved in BCG-induced apoptosis of TCC lines, using at least in part the mitochondrial pathway.

Bacillus Calmette-Guérin (BCG) is considered to be one of the most effective treatments for superficial and in situ bladder cancer. The exact mechanism of the antitumor activity of BCG is not completely understood. Peroxisome proliferatoractivated receptor gamma (PPARÁ) is a member of the nuclear receptor superfamily of ligand-activated transcription factors that is involved in cell growth and differentiation as well as inflammatory processes. PPARÁ is expressed in normal urothelium and a lack of expression was associated with bladder cancer progression. We analyzed whether PPARÁ is involved in the inhibition of bladder cancer cell survival by BCG. PPARÁ expression in murine MB49 and human T24 bladder cancer cells was evaluated employing immunofluorescence and inmunohistochemistry techniques. In vitro cell viability and nitric oxide (NO) production was evaluated by using MTS and Griess reagent respectively. Our results show that BCG induced the cytoplasmatic expression of PPARÁ in bladder tumor cells in vitro and in vivo. BADGE, antagonist of this receptor, abrogated in vitro BCG-mediated cell cytotoxicity. Natural agonist 15-deoxy-Δ12,14 prostaglandin J2 (15-d-PGJ2) but not rosiglitazona (RO), a synthetic agonist, induced in vitro inhibition of cell viability of both cancer cell lines and the effect was partially reversed by BADGE. We also determined whether the activation of PPARÁ could inhibit NO production, which is considered a survival factor for bladder tumor cells. Both 15-d-PGJ2 and RO significantly inhibited the NO production in T24 and MB49 cells by PPARÁindependent pathway since it was not antagonized by BADGE. Thus, our results show that BCG induces functional PPARÁ in bladder tumor cells in vivo and in vitro, being these receptors intrinsically involved in the antitumor activity of BCG.

BackgroundBacillus Calmette-Guerin (BCG) is the most effective treatment for non-muscle invasive bladder cancer. However, a failure in the initial response or relapse within the first five years of treatment has been observed in 20% of patients. We have previously observed that in vivo administration of an inhibitor of nitric oxide improved the response to BCG of bladder tumor bearing mice. It was described that this effect was due to a replacement of tumor tissue by collagen depots. The aim of the present work was to clarify the mechanism involved in this process.Methodology/Principal FindingsWe demonstrated that BCG induces NIH-3T3 fibroblast proliferation by activating the MAPK and PI3K signaling pathways and also differentiation determined by alpha-smooth muscle actin (alpha-SMA) expression. In vivo, intratumoral inoculation of BCG also increased alpha-SMA and collagen expression. Oral administration of L-NAME enhanced the pro-fibrotic effect of BCG. Peritoneal macrophages obtained from MB49 tumor-bearing mice treated in vivo with combined treatment of BCG with L-NAME also enhanced fibroblast proliferation. We observed that FGF-2 is one of the factors released by BCG-activated macrophages that is able to induce fibroblast proliferation. The involvement of FGF-2 was evidenced using an anti-FGF2 antibody. At the same time, this macrophage population improved wound healing rate in normal mice and FGF-2 expression was also increased in these wounds.Conclusions/SignificanceOur findings suggest that fibroblasts are targeted by BCG both directly and through activated macrophages in an immunotherapy context of a bladder murine model. We also described, for the first time, that FGF-2 is involved in a dialog between fibroblasts and macrophages induced after BCG treatment. The fact that L-NAME administration improves the BCG effect on fibroblasts, NO inhibition, might represent a new approach to add to the conventional BCG therapy.