Intracellular reactive oxygen species in monocytes generated by photosensitive chromophores activated with blue light

Abstract 

Objectives

Disinfection of the tooth pulp-canal system is imperative to successful endodontic therapy. Yet, studies suggest that 30–50% of current endodontic treatments fail from residual bacterial infection. Photodynamic therapy using red-light chromophores (630nm) to induce antimicrobial death mediated by generated reactive oxygen species (ROS) has been reported, but red-light also may thermally damage resident tissues. In the current study, we tested the hypothesis that several blue light chromophores (380–500nm) generate intracellular reactive oxygen species but are not cytotoxic to mammalian cells.

Methods

THP1 monocytes were exposed to 10μM of four chromophores (chlorin e6, pheophorbide-a, pheophorbide-a-PLL, and riboflavin) for 30min before activation with blue light (27J/cm², 60s). After activation, intracellular ROS were measured using a dihydrofluorescein diacetate technique, and cytotoxicity was determined by measuring mitochondrial activity with the MTT method.

Results

All photosensitizers produced intracellular ROS levels that were dependent on both the presence of the photosensitizer and blue light exposure. Riboflavin and pheophorbide-a-PLL produced the highest levels of ROS. Photosensitizers except riboflavin exhibited cytotoxicity above 10μM, and all except pheophorbide-a-PLL were more cytotoxic after blue light irradiation.

Significance

The current study demonstrated the possible utility of blue light chromophores as producers of ROS that would be useful for endodontic disinfection.

Keywords: ROS, Photodynamic therapy, Endodontic infection, Oxidative stress, Cytotoxicity