We used the REN human mesothelioma cell line, as an in vitro cancer cell model, and the non-malignant human mesothelial MeT5A cell line, as normal cell model. Cytosolic Ca2+ concentration was measured by the fluorescent indicator Fura-2. Immunofluorescence, Western blot, and siRNA technique were employed to assess the involvement of T-type Ca2 + channels. Cell viability was determined by the calcein assay.
REN cells transiently exposed to 1–10 μM Res showed increasing peaks of Ca2+ that were absent in Ca2+-free medium and were reduced by non-selective (Ni2+), and highly selective (NNC 55-0396) T-type Ca2+ channels antagonist, and by siRNA knockout of Cav3.2 T-type Ca2+ channel gene. Dose-dependent curve of Res-induced Ca2+ peaks showed a rightward shift in normal MeT-5 A mesothelial cells (EC50 = 4.9 μM) with respect to REN cells (EC50 = 2.7 μM). Moreover, incubation with 3 and 10 μM Res for 7 days resulted in cell growth inhibition for REN, but not for MeT-5 A cells.
Res induces Ca2+ influx, possibly mediated through T-type Ca2+ channels, with significant selectivity towards mesothelioma cells, suggesting a possible use as an adjuvant to chemotherapy drugs for mesothelioma clinical treatment.