A human relevant mixture of persistent organic pollutants (POPs) and perfluorooctane sulfonic acid (PFOS) enhance nerve growth factor (NGF)-induced neurite outgrowth in PC12 cells

Ajay Yadav,Mazia Amber,Denis Zosen,Nils Anders Labba,Eva Henriette Willemijn Huiberts,Johanna Samulin Erdem,Fred Haugen,Hanne Friis Berntsen,Shanbeh Zienolddiny,Ragnhild Elisabeth Paulsen,Erik Ropstad,Lisa Connolly,Steven Verhaegen

doi:10.1016/j.toxlet.2020.12.007

Abstract

Disruption of neurite outgrowth is a marker for neurotoxicity. Persistent organic pollutants (POPs) are potential developmental neurotoxicants. We investigated their effect on neurite outgrowth in PC12 rat pheochromocytoma cells, in absence or presence of nerve growth factor (NGF), an inducer of neuronal differentiation. Cells were exposed for 72 h to a defined mixture of POPs with chemical composition and concentrations based on blood levels in the Scandinavian population. We also evaluated perfluorooctane sulfonic acid (PFOS) alone, the most abundant compound in the POP mixture. Only higher concentrations of POP mixture reduced tetrazolium salt (MTT) conversion. High-content analysis showed a decrease in cell number, but no changes for nuclear and mitochondrial cellular health parameters. Robust glutathione levels were observed in NGF-differentiated cells. Live imaging, using the IncuCyte ZOOM platform indicated ongoing cell proliferation over time, but slower in presence of NGF. The pollutants did not inhibit neuritogenesis, but rather increased NGF-induced neurite length. PFOS induced neurite outgrowth to about 50 % of the level seen with the POP mixture. Neither the POP mixture nor PFOS affected neurite length in the absence of NGF. Our observations indicate that realistic complex mixtures of environmental pollutants can affect neuronal connectivity via NGF-induced neurite outgrowth.

Highlights

Normal development of the central nervous system requires the coordinated ontogeny of proliferation, migration, differentiation, synaptogenesis, gliogenesis, myelination and apoptosis (Barone et al, 2000)
Various cell lines have been used to study the effects of chemicals on neurons (Radio and Mundy, 2008), including the pheochromocytoma PC12 cell line which has been widely used as a model for the study of neuronal differentiation and neurite outgrowth (Fujita et al, 1989)
Since neuritogenesis is a marker for neuronal differentiation, and neurite outgrowth is very sensitive to neurotoxicants we investigated the effects of persistent organic pollutants (POPs) on neurite dynamics in the PC12 cell model

Summary

Introduction

Normal development of the central nervous system requires the coordinated ontogeny of proliferation, migration, differentiation, synaptogenesis, gliogenesis, myelination and apoptosis (Barone et al, 2000). Perturbations of these processes through exposure to environmental toxicants during development, could affect structures and functions of the brain, leading to lifelong impairment (Grandjean and Landrigan, 2006). Many of the neurodevelopmental processes that occur in vivo for example cell differentiation, neurite outgrowth, and synaptogenesis, can be followed using in vitro neuronal systems. Serum-withdrawal together with stimulation with nerve growth factor (NGF) results in the cells undergoing one doubling and induction of neurite outgrowth (Greene, 1978)

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Conclusion