Abstract
Lead (Pb) is one of the most important environmental pollutant metals accumulating in the atmosphere, water, foods, and in organisms living in contaminated areas. Skin is one of the main targets of Pb toxicity based on its ability of direct penetration upon exposure. The underlying cell damaging pathomechanisms have not been revealed in detail. Herein, we focus on Pb-induced oxidative and nitrosative stress that has not been previously thoroughly investigated. We investigated these effects in order to elucidate the pathomechanisms and as well to identify potential biological markers that may indicate Pb toxicity. Human immortalized keratinocytes (HaCaT cells) were exposed to Pb (100 μM) either for 5 minutes or 6 hours. Pb-induced cellular damage was evaluated by immunocytochemistry analysis of multiple signalling cascades, e.g. apoptosis, Akt, MAPK, NOS, nitrotyrosine and 8-isoprostane formation, detection of nitrosative stress using Diaminofluorescein (DAF-FM) and oxidative stress using 3'-(p aminophenyl) fluorescein (APF). We found that Pb exposure resulted in significantly enhanced NO and ROS production in HaCaT cells. Pb led to enhanced eNOS-phosphorylation at Ser1177, and Ser116 residues but not Thr495. AKT phosphorylation but not MAP kinases were enhanced by Pb In addition, Pb induced apoptosis as shown by Caspase-3 activation and PARP cleavage. Our results suggest that Pb mediates its toxic effect in keratinocytes through oxidative and nitrosative stress which is accompanied by differential changes of eNOS phosphorylation and apoptosis. These data significantly contribute to understanding of underlying mechanisms of Pb-induced cellular damage.
Highlights
Lead (Pb) is one of the most important metals that pollute the natural environment due to man’s impact
Pb increased nitric oxide (NO) production: The intracellular production of free NO-radicals was measured by the NO-sensitive fluorescent dye (DAFFM DA) during a 5 minutes period of BaP exposure
The DAF-FM fluorescence showed a time dependent significant increase in NO release after Pb exposure throughout the whole treatment period compared to the control (H2O) group, it showed a fair acceleration throughout the whole period (Figure 1)
Summary
Lead (Pb) is one of the most important metals that pollute the natural environment due to man’s impact. As Pb cannot be degraded, it accumulates in the atmosphere, water, foods, and in organisms living in contaminated areas [1]. Environmental accumulation with Pb has accelerated due to its dose relationship to industrialization, major sources of lead exposure are dust, water, paint, cosmetics, folk remedies, and food supplements [2]. Prolonged exposure to Pb may cause reproductive impairment, hypertension, and nephropathy. Pb slows nerve conduction, alters calcium homeostasis, inhibits enzymes, and stimulates synthesis of binding proteins [3]. The persistence of Pb in animals and humans and the associated health risk is a highly relevant topic of current concern
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