Relationship between toxicity of selected insecticides and expression of stress proteins (HSP, GRP) in cultured human cells: effects of commercial formulations versus pure active molecules

Three carbamate (formetanate, methomyl, pyrimicarb) and one pyrethroid (bifenthrin) insecticides were investigated both as pure chemicals and as commercial formulations in order to unveil possible toxic effects of additives and solvents present in the commercial formulations and to evaluate the cellular stress response as a defense mechanism. Toxic effects were evaluated on A549 cells, derived from a human lung carcinoma, by measuring (1) threshold concentrations leading to a decrease of the growth rate (LOEC), (2) sublethal concentrations (SC) which arrested growth without killing the cells, and (3) expression levels of several stress proteins, i.e., HSP27, HSP72/73, HSP90, GRP78, and GRP94. As compared to the pure active molecule, LOEC appeared at lower concentrations when using the commercial formulations, i.e., Dicarzol (formetanate), Lannate20 (methomyl) and Talstar or Kiros EV (bifenthrin). Propylene glycol and propylene glycol monomethyl ether, respectively, present in Talstar and kiros, do not account for the high toxicity of these commercial formulations and do not potentiate the toxicity of bifenthrin. Additive but not synergistic adverse effects were observed when cells are exposed to a mixture of 4 different commercial formulations. Our results show that the concentrations of active molecules recommended in flori-cultural general use or for spray preparations are much higher than SC concentrations, as determined on A549 pulmonary cells. GRP78 was up-regulated by all the insecticides, commercial preparations being more efficient to trigger the stress reaction. This suggests that insecticides and additives present in commercial formulations disrupt ER functions. Conversely, HSP72/73 was found to be down-regulated by all the insecticides. This seems to be related with a decrease of protein synthesis in the cytosol, as a result of the ER unfolded protein response. Indeed, tunicamycin, known to inhibit N-linked glycosylation in the ER, was found to induce a similar inverse correlation between GRP78 overexpression and HSP72/73 under-expression. Expression of GRP94 was found to be increased and HSP27 lowered by the highest concentrations of bifenthrin commercial formulations. Methomyl and Lannate20 only induced an under-expression of HSP90.