Hydroxyapatite nanocomposite as a potential agent in osteosarcoma PDT

Using Nanoplatforms as a hauler for photosensitizers is a bespoke paradigm to improve its bioavailability and to boost the PDT efficacy. Herein, the photodynamic cytotoxicity of methylene blue (MB) loaded on hydroxyapatite nanoparticles (HA-NPs) was tested against human osteosarcoma-derived cells (Saos-2 cell line). HA-NPs and HA-NPs loaded with MB (HA-NPs-MB) were prepared by a chemical precipitation method and characterized by TEM, Zeta potential, FTIR, and XRD. TEM images revealed that HA-NPs have a rod shape with a diameter of 14-17 nm and length around 46-64 nm. FTIR and Zeta potential confirmed the adsorption of cationic MB on HA-NPs. XRD pattern was identical to the standard XRD pattern of HA-NPs. Incubation of Saos-2 cells (24 h) with HA-NPs-MB then irradiation of cells (5 min) with a diode laser (808 nm), causes a higher decrement of cell viability (determined by MTT assay) than that caused by free MB. The LC₅₀ was 57.53 µg/mL and 86.99 µg/mL for HA-NPs-MB and free MB, respectively. Thus, the nanoformulation of MB greatly reduced the dose of MB required for effective PDT. This study also investigated the mode of cell death after incubation of cells with free MB or HA-NPs-MB composite then exposure to laser radiation. The results revealed that the majority of cells died by apoptosis while a minor fraction of cells died by necrosis, especially in the case of HA-NPs-MB. Levels of caspase-3 and death receptor-4 (DR-4) were more elevated in the case of HA-NPs-MB than free MB. The effect of the prepared nanocomposite and free MB on Raw murine macrophage (RAW 264.7) viability was also examined using the MTT assay. The results indicated that HA-NPs-MB in the presence of laser has a great cytotoxic effect on macrophage cells compared to other treatments. This may present an advantage through decreasing macrophage that promotes tumor growth. In conclusion, HA-NPs-MB nanocomposite surmounts free MB and HA-NPs in destroying macrophage cells and Saos-2 cells through apoptosis in the presence of laser irradiation. This study introduces a thorough and new insight on osteosarcoma (cancer cell line Saos-2) PDT using HA-NPs-MB exploiting the biosafety of HA-NPs.