Altered calcium regulation by thermosensitive transient receptor potential channels in etoposide-resistant WERI-Rb1 retinoblastoma cells

Differences in transient receptor potential (TRP) and cannabinoid receptor type 1 (CB1) expression levels can serve as prognostic factors for retinoblastoma (RB) tumor progression. We hypothesized in RB tissue that such differences are also indicators of whether or not they are sensitive to etoposide. Accordingly, we compared in malignant etoposide-sensitive and etoposide-resistant WERI-Rb1 cells TRPV1, TRPM8 and TRPA1 subtype and CB1 gene expression pattern levels and accompanying functional activity using quantitative real-time RT-PCR, immunohistochemistry, immunofluorescence microscopy, calcium imaging as well as patch-clamp technology. Gene expression patterns were evaluated in enucleated human RB tissues (n=4). Both etoposide-resistant and etoposide-sensitive WERI-Rb1 cells expressed all of the aforementioned channels based on responses to known activators and thermal challenges. However, TRPA1 was absent in the etoposide-resistant counterpart. Even though both types of RB cells express TRPV1 as well as TRPM8 and CB1, the capsaicin (50μM) (CAP)-induced Ca ²⁺ rise caused by TRPV1 activation was prompt and transient only in etoposide-resistant RB cells (n=8). In this cell type, the inability of CB1 activation (10μM WIN) to suppress Ca ²⁺ responses to CAP (50μM; n=4) may be attributable to the absence of TRPA1 gene expression. Therefore, using genetic approaches to upregulate TRPA1 expression could provide a means to induce etoposide sensitivity and suppress RB cell tumorigenesis.