Atomic Force Microscopy of Cellular Damage in Human Keratinocytes Exposed at Pesticide Solution.

Pesticides are chemicals intentionally released into the environment to cause toxic effects in selected pest species. In particular, Deltamethrin is a pyrethroid widely used for synthesizing pesticide products which are very effective in damaging the central nervous system of pests. Although the use of pesticides yields benefits, it can also cause drawbacks, such as potential toxicity to humans and other animals. Study of the exposure effects, at the concentration indicated for the utilization and even lowers, is important for the safety of those people employed in the manufacturing and utilization activities and consumers as well. To assess the possible health effects due to the action of these chemicals, investigations on cells “in vitro” are required to monitor cellular changes caused by non-cytotoxic doses of such compounds. Previously, we found biochemical and morphological damages induced to cultured human epithelial cells by exposure at Delthametrin concentration well below the cytotoxic value [1]. In this work, we analyze morphological changes induced in human cells by the exposure at a Deltamethrin-based commercial pesticide (‘‘Decaflow’’) containing formulant agents as well. In particular, our aim is to investigate the role of formulants in the toxicity of the Decaflow solution by using AFM techniques for the cellular characterization. In fact, besides the well-known topographical performance, AFM has the capability to monitor interesting mechanical surface parameters, as roughness and elastic moduli. So, we studied a mathematical method through which it is possible to filter a roughness parameter according to the size of the surface features. In this way we can estimate two roughness values: one related to the real plasmatic membrane roughness and another one related to the membrane micro-waviness caused by the underneath cytoskeleton network. As for the elastic moduli, this is an index of the cellular rigidity and so it is related to the cytoskeleton too. Cultured human keratinocyte cells have been exposed for 24 h to different Decaflow solutions containing increasing concentrations of Deltamethrin. Our investigation shows that cells exposed at Decaflow solution containing a given concentration of active ingredient are more damaged with respect to cells exposed at the same concentration of pure Deltamethrin. In particular, AFM topographies of exposed cells show morphological alteration of cell form, reduction of volume and decreasing of cell adhesion and connection structures, as lamellopodia and microvilli. The roughness values indicate an increasing in membrane roughness and contemporaneously damages in the cytoskeleton network as a consequence of the chemical action of the toxic. These damages have been confirmed by the trend of the elastic moduli, which indicates a softening of the cells with increasing pesticide concentration. Some of such injuries were detected in the case of pure Deltamethrin exposure, but to a lesser degree. The AFM technique has been confirmed a powerful technique to evaluate the state of health of the cells and its alteration induced by chemical or mechanical stresses.