Abstract
PurposePer- and poly-fluoroalkyl-substances (PFASs) are synthetic compounds that raised concern due to their potential adverse effects on human health. Long-chain PFAS were banned by government rules in many states, and thus, new emerging PFAS were recently introduced as substitutes. Among these, Perfluoro{acetic acid, 2-[(5-methoxy-1,3-dioxolan-4-yl)oxy]}, ammonium salt (C6O4) was recently introduced to produce a range of food contact articles and literature data about this compound are scanty. The aim of this study was to evaluate the in vitro effects of exposure to C6O4, compared with PFOA and PFOS on thyroid cells.MethodsFRTL5 rat-thyroid cell lines and normal human thyroid cells (NHT) were incubated with increasing concentrations of C6O4 for 24, 48, 72, and 144 h to assess cell viability by WST-1. Cell viability was confirmed by AnnexinV/PI staining. Long-chain PFAS (PFOA and PFOS) were used at same concentrations as positive controls. The proliferation of cells exposed to C6O4, PFOA, and PFOS was measured by staining with crystal violet and evaluation of optical density after incubation with SDS. Changes in ROS production by FRTL5 and NHT after exposure to C6O4 at short (10, 20, and 30 min) and long-time points (24 h) were evaluated by cytofluorimetry.ResultsC6O4 exposure did not modify FRTL5 and NHT cell viability at any concentration and/or time points with no induction of necrosis/apoptosis. At difference, PFOS exposure reduced cell viability of FRTL5 while and NHT, while PFOA only in FRTL5. FRTL5 and NHT cell proliferation was reduced by incubation with by PFOA and PFOS, but not with C6O4. ROS production by NHT and FRTL5 cells was not modified after C6O4 exposure, at any time/concentration tested.ConclusionsThe present in vitro study constitutes the first evaluation of the potential adverse effects of the new emerging PFAS C6O4 in cultured rat and human thyroid cells, suggesting its safety for thyroid cells in vitro.
Highlights
Per- and poly-fluoroalkyl substances (PFASs) are a class of man-made chemicals, globally used as surfactants in industrial productions due to their surface active properties as well as to their thermal and chemical stability [1]
The WST-1 assay indicated that the viability of FRTL5 was not affected by C6O4, and that no damage on the mitochondrial respiration occurred following exposure to the compound
perfluorooctanoic acid (PFOA) exposure did not modify normal human thyroid cells (NHT) cell viability at all time points and concentrations (ANOVA F = 3.641, p = 0.07) (Fig. 2d–g) At difference, perfluorooctanesulfonic acid (PFOS) exposure induced a reduction of NHT cell viability, starting from different concentrations at all time points (ANOVAs 24 h F = 15.338, p < 0.05; 48 h F = 13.609 p < 0.05; 72 h F = 18.239 p < 0.05; 144 h F = 52.615 p < 0.05) (Fig. 2h–m)
Summary
Per- and poly-fluoroalkyl substances (PFASs) are a class of man-made chemicals, globally used as surfactants in industrial productions due to their surface active properties as well as to their thermal and chemical stability [1]. Among PFAS, two “Long chain PFAS” (so-called owing to the presence of 8 or more carbon atoms), perfluorooctanoic acid (PFOA) and perfluorooctanesulfonic acid (PFOS), were object of growing concerns due to their elevated persistency and bioaccumulation in the environment [2] These compounds are constantly discharged into the environment from manufacturing processes and daily usage, being persistent and with long time of degradation [1]. Further concern derives from the fact that PFOA and PFOS were reported to act as endocrine thyroid disruptors and, in some cases, as cancer promoting agents [3]; they induce oxidative stress, both in humans and in animal models [3,4,5,6] Due to their potential adverse effects for human health, longchain PFAS production was restricted or even banned in some countries [7]. On 16 April 2019, ARPAV (the Regional Agency for the Prevention and Protection of the Environment in Veneto) detected C6O4 in the River Po near certain municipalities in the Veneto region, bordering on the Emilia-Romagna region (Corbola), and in one municipality on the border between Lombardy and Emila-Romagna regions (Castelmassa), with peak concentrations of around 100 ng/l [16]
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