Temperature-dependent and time-dependent effects of hyperthermia mediated by dextran-coated La0.7Sr0.3MnO3: in vitro studies

Abstract

Reihaneh Haghniaz, Rinku D Umrani, Kishore M Paknikar Centre for Nanobioscience, Agharkar Research Institute, Pune, Maharashtra, India Background: The purpose of this study was to investigate the therapeutic efficacy of dextran-coated (Dex) La0.7Sr0.3MnO3 (LSMO) nanoparticles-mediated hyperthermia at different temperatures (43°C, 45°C, and 47°C) based on cell killing potential and induction of heat shock proteins in a murine melanoma cell (B16F1) line. Methods: LSMO nanoparticles were synthesized by a citrate-gel method and coated with dextran. B16F1 cells were exposed to the Dex-LSMO nanoparticles and heated using a radiofrequency generator. After heating, the morphology and topology of the cells were investigated by optical microscopy and atomic force microscopy. At 0 hours and 24 hours post heating, cells were harvested and viability was analyzed by the Trypan blue dye exclusion method. Apoptosis and DNA fragmentation were assessed by terminal deoxynucleotidyl transferase-dUTP nick end labeling (TUNEL) assay and agarose gel electrophoresis, respectively. An enzyme-linked immunosorbent assay was used to quantify heat shock protein levels. Results: Our data indicate that cell death and induction of heat shock proteins in melanoma cells increased in a time-dependent and temperature-dependent manner, particularly at temperatures higher than 43°C. The mode of cell death was found to be apoptotic, as evident by DNA fragmentation and TUNEL signal. A minimum temperature of 45°C was required to irreversibly alter cell morphology, significantly reduce cell viability, and result in 98% apoptosis. Repeated cycles of hyperthermia could induce higher levels of heat shock proteins (more favorable for antitumor activity) when compared with a single cycle. Conclusion: Our findings indicate a potential use for Dex-LSMO-mediated hyperthermia in the treatment of melanoma and other types of cancer. Keywords: hyperthermia, Dex-LSMO nanoparticles, heat shock proteins, melanoma, apoptosis