Optical assessment of early cell responses to nanotoxicological effects
Vladimir A. Bogatyrev, 1 Daniil S. Chumakov, 1 Aram Akopiyan, 3 Timofey E. Pylaev, 1,2 Lev A. Dykman 1 1 Institute of Biochemistry and Physiology of Plants and Microorganisms, RAS, Saratov, Russia 2 Saratov State Medical University, Saratov, Russia 3 Saratov State University, Saratov, Russia
Abstract
The previously developed microplate test system for assessing the toxicity of nanomaterials for the microalga Dunaliella salina, based on the change in the light attenuation parameter at a wavelength of 680 nm (Ã680) as a result of chlorophyll photodestruction, was supplemented with colorimetric and fluorimetric indicators that allow assessing the physiological state of cultures at the early stages of cytotoxic activity.
One of the first reactions of cells to the effect of nanoparticles is a change in oxidative metabolism, accompanied by hyperproduction of reactive oxygen species and activation of antioxidant defense systems. The consequence of this may be the initiation of programmed cell death. A convenient model for studying changes in the physiological state is unicellular algae, in particular, extremophilic microalgae of the genus Dunaliella. The physiological state can be indicated by the photosynthetic apparatus, and the measured parameter is the chlorophyll fluorescence, in particular the Kautsky effect, the change in the chlorophyll luminescence that occurs when the chloroplast, previously adapted to darkness, is illuminated. To put it very simply, in a physiologically active green cell, a large proportion of the absorbed light energy is spent on photosynthesis; accordingly, the difference in the fluorescence intensity of plants adapted to darkness and light is greater than in depressed or dormant cells.
An adequate measure of the chlorophyll content can be the fluorescence intensity of cultures fixed with glutaraldehyde. In this case, the distorting effect of colloidal gold preparations can be minimized, since the maxima of chlorophyll fluorescence excitation fall into the transparency windows of gold sols.
Speaker
Vladimir A. Bogatyrev
IBPPM RAS
Russia
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