Methylene blue-mediated Photodynamic Therapy in human retinoblastoma cell lines

Abstract

Retinoblastoma is a malignant tumor of the retinal precursor cells and one of the rarest types of pediatric tumor, often occurring in the earliest years of life. Symptoms are conditioned by tumor size and location; one of the most recurrent symptoms is a white reflex in the pupillary area, called leukocoria or cat's eye reflex. In the present work, we studied the in vitro effectiveness of Photodynamic treatment (Pdt) in two types of human retinoblastoma, Y79 and WERI-Rb cell lines, using methylene blue (MB), a photosensitizer (PS) from the phenothiazine group. The two cell lines were incubated with varying concentrations of MB (3, 7, 10, 15, 20, 25, 30, 40, and 50 μM), in the absence of light (dark cytotoxicity) and, in the presence of 664 nm laser light (phototoxicity) with fluences of 1, 1.5, 3, 5, 7, 10, and 15 J/cm². The Y79 cell line showed higher cellular uptake values for MB than the WERI-Rb cell line. After three hours of incubation, the Y79 and WERI-Rb took up 48% and 34% of the total photosensitizer present in the medium, respectively. Using MTT assay, the results showed that the Y79 cell line was more affected by the photo treatment as demonstrated by the combination of MB concentration and light doses compared with WERI-Rb cell line. The results were correlated with the more pronounced singlet oxygen emission observed in Y79 cells. While MB does show efficacy for eradication of retinoblastoma in vitro, only studies in appropriate animal models will reveal whether the selectivity of photokilling at tolerable drug and light doses is sufficient to suggest clinical trials.

Keywords: Cellular uptake; Methylene blue; Photodynamic Therapy; Phototoxicity; Retinoblastoma cell lines.