Photobiological and thermal effects of photoactivating UVA light doses on cell cultures

Abstract

While near-ultraviolet light has been widely used to photoactivate fluorophores and caged compounds in cells, little is known of the long-term biological effects of this light. UVA (315–400 nm) photoactivating light has been well characterized in short-term cell studies and is now being employed in higher doses to control longer-duration phenomena (e.g. gene expression). Annexin V-Cy5/propidium iodideapoptosis flow cytometry assays were used to determine responses of HeLa cells to doses of UVA light up to 23.85 J cm^–2. Cells seeded at low densities had higher percentages of apoptosis and necrosis and were also more susceptible to UVA damage than cells seeded at higher densities. The dose to induce apoptosis and death in 50% of the cells (dose_1/2) was determined for two different commercially available UVA light sources: 7.6 J cm^–2 for the GreenSpot photocuring system and 2.52 J cm^–2 for the BlakRay lamp. All BlakRay doses tested had significant cellular responses, whereas no significant cellular responses were found for doses below 1.6 J cm^–2 from the GreenSpot light source. A temperature control and measurement system was used to determine direct heating from the UVA sources and also the effect that cooling cell cultures during photoexposure has on minimizing cell damage. Cooling during the BlakRay photoexposure significantly reduced the percentage of necrotic cells, but there was no significant difference for cooling during photoactivation with the GreenSpot. Differences in cell responses to similar UVA doses of different intensities suggest that photoduration should be considered along with total dose and thermal conditions in photoactivation studies.