Paraoxon and Pyridostigmine Interfere with Neural Stem Cell Differentiation

Abstract

Acetylcholinesterase (AChE) inhibition has been described as the main mechanism of organophosphate (OP)-evoked toxicity. OPs represent a human health threat, because chronic exposure to low doses can damage the developing brain, and acute exposure can produce long-lasting damage to adult brains, despite post-exposure medical countermeasures. Although the main mechanism of OP toxicity is AChE inhibition, several lines of evidence suggest that OPs also act by other mechanisms. We hypothesized that rat neural progenitor cells extracted on embryonic day 14.5 would be affected by constant inhibition of AChE from chronic exposure to OP or pyri-dostigmine (a reversible AChE blocker) during differentiation. In this work, the OP paraoxon decreased cell viability in concentrations >50 μM, as measured with the MTT assay; however, this effect was not dose-dependent. Reduced viability could not be attributed to blockade of AChE activity, since treatment with 200 μM pyri-dostigmine did not affect cell viability, even after 6 days. Although changes in protein expression patterns were noted in both treatments, the distribution of differentiated phenotypes, such as the percentages of neurons and glial cells, was not altered, as determined by flow cytometry. Since paraoxon and pyridostigmine each decreased neurite outgrowth (but did not prevent differentiation), we infer that developmental patterns may have been affected.

Neurotoxicity in Preclinical Models of Occupational Exposure to Organophosphorus Compounds

“…The 100 μM group also showed cytoplasmic volume decrease and increased number of vacuoles, characteristics of necrotic cell death (Yousefpour et al, 2006). This data was supported by a later study showing that POX exposure for 6 days at 50, 100, 150, 200, and 300 μM resulted in reduced cell viability of cultured neural stem and progenitor cells (dNPCs), as measured by MTT assay and PI (Berríos et al, 2015). Despite 200 μM POX exposures causing 50% AChE inhibition, the authors concluded that cell viability was unrelated to AChE inhibition, based on studies using pyridostigmine (PY), a reversible AChE inhibitor that did not reduce cell viability despite producing the same degree of AChE inhibition (Berríos et al, 2015).…”

“…Yousefpour et al (2006) reported decreases in the number and length of neuronal processes in hippocampal primary cells 1 week after exposure to POX at 30, 40, 50, and 100 μM. These data were supported by an additional study showing that prolonged POX exposure at 16–66 μM POX indiscriminately decreased neurite outgrowth in dNPCs, as measured by Scholl analysis (Berríos et al, 2015). Additionally, T O CP dose-dependently inhibited neurite outgrowth in SH-SY5Y cells following 24 h treatment with 200, 500, and 1000 μM, and an autophagy inhibitor prevented and reversed this growth inhibition, indicating a critical role for autophagy in T O CP-induced neurite changes at high doses (Chen et al, 2013).…”

“…DZ (Rush et al, 2010), parathion (Yousefpour et al, 2006; Berríos et al, 2015), and DDVP (Binukumar et al, 2010; Binukumar and Gill, 2010) have also been shown to induce cytotoxicity following exposures that do not inhibit ChE activity in vitro , or exposures that do not cause overt toxicity in vivo . DZ produced neuronal cell death in primary cortical neurons following 24 h of exposure to 30 and 100 μM, resulting in approximately 50% AChE inhibition (Rush et al, 2010).…”

“…Changes in length, number, or branching patterns of axons or dendrites have been linked to psychiatric manifestations in patients, including depression, anxiety, AD, and PD (Marsden, 2013; Rubia et al, 2014; Canu et al, 2015; Negrón-Oyarzo et al, 2016). Axonal length, neurite length, neurite number, and dendritic branching in neuronal culture systems have also been shown to be altered following exposure to a variety of OPs, including CPF (Howard et al, 2005; Yang et al, 2008; Speed et al, 2012), DZ (Pizzurro et al, 2014a,b), parathion (Yousefpour et al, 2006; Berríos et al, 2015), and T O CP (Chen et al, 2013; Duarte et al, 2016; Hausherr et al, 2016). For example, in primary cultures of sympathetic neurons, CPF inhibited axon outgrowth but promoted dendritic arborization (Howard et al, 2005).…”

Cognitive impairment and neuronal damage in Alzheimer's disease are malleable

“…The 100 µM group also showed cytoplasmic volume decrease and increased number of vacuoles, characteristics of necrotic cell death (Yousefpour et al 2006). This data was supported by a later study showing that POX exposure for 6 days at 50, 100, 150, 200, and 300 µM resulted in reduced cell viability of cultured neural stem and progenitor cells (dNPCs), as measured by MTT assay and PI (Berrios et al 2015).…”