Chronic Exposure to Dietary Sterol Glucosides is Neurotoxic to Motor Neurons and Induces an ALS–PDC Phenotype

Abstract

Epidemiological studies of the Guamanian variants of amyotrophic lateral sclerosis (ALS) and parkinsonism, amyotrophic lateral sclerosis-parkinsonism dementia complex (ALS-PDC), have shown a positive correlation between consumption of washed cycad seed flour and disease occurrence. Previous in vivo studies by our group have shown that the same seed flour induces ALS and PDC phenotypes in out bred adult male mice. In vitro studies using isolated cycad compounds have also demonstrated that several of these are neurotoxic, specifically, a number of water insoluble phytosterol glucosides of which β-sitosterol β-D-glucoside (BSSG) forms the largest fraction. BSSG is neurotoxic to motor neurons and other neuronal populations in culture. The present study shows that an in vitro hybrid motor neuron (NSC-34) culture treated with BSSG undergoes a dose-dependent cell loss. Surviving cells show increased expression of HSP70, decreased cytosolic heavy neurofilament expression, and have various morphological abnormalities. CD-1 mice fed mouse NIH-PA Author ManuscriptNIH-PA Author Manuscript NIH-PA Author Manuscript chow pellets containing BSSG for 15 weeks showed motor deficits and motor neuron loss in the lumbar and thoracic spinal cord, along with decreased glutamate transporter labelling, and increased glial fibrillary acid protein reactivity. Other pathological outcomes included increased caspase-3 labelling in the striatum and decreased tyrosine-hydroxylase labelling in the striatum and substantia nigra. C57BL/6 mice fed BSSG-treated pellets for 10 weeks exhibited progressive loss of motor neurons in the lumbar spinal cord that continued to worsen even after the BSSG exposure ended. These results provide further support implicating sterol glucosides as one potential causal factor in the motor neuron pathology previously associated with cycad consumption and ALS-PDC.

Early Exposure to Environmental Toxin Contributes to Neuronal Vulnerability and Axonal Pathology in a Model of Familial ALS

“…The 10-week starting point for SG feeding allowed time for sufficient baseline motor behavioral testing after weaning at 3 weeks. The 42 mg/kg body weight dose of SG corresponds approximately to those reported in Tabata et al [21]. However, in the previous study, toxin exposure was set at a fixed concentration and began at 5 months of age.…”

“…Cycad seed flour was found to contain a variety of neurotoxins, some of which remained in the flour after washing. Of these, we focused our attention on several steryl glucosides (SG) [18][19][20][21]. These molecules are neurotoxic to primary neuronal and astroglial cultures [18] as well as to motor neuron-derived cell lines [20].…”

“…These molecules are neurotoxic to primary neuronal and astroglial cultures [18] as well as to motor neuron-derived cell lines [20]. Moreover, synthetic SGs fed to mice caused motor deficits consistent with an ALS-PDC phenotype, notably a progressive motor neuron loss in spinal cord and later cell loss in the striatum [21,22]. Motor neuron loss in animal models is demonstrable by the cycad neurotoxins linked to Guamanian ALS and Parkinsonism [23,24].…”

Disparity of outcomes: the limits of modeling amyotrophic lateral sclerosis in murine models and translating results clinically

“…To date, primarily on account of the unknown etiology, there is a lack of models that are representative of the sporadic form of the disease. Our group has previously investigated the dietary administration of cycad flour or extracted sterol glucosides as causative agents in the purported disease cascade (91)(92)(93) On account of our increased understanding of the underlying genetic causal factors at play in ALS (Figure 1), there have been multiple additional murine models developed since the advent of the mutant SOD1 mouse. These include mice expressing mutant TDP-43 or the C9ORF72-associated hexanucleotide repeat expansions (95)(96)(97).…”

In Vivo and In Vitro Models to Study Amyotrophic Lateral Sclerosis

“…They respond to agents affecting voltage-gated ion channels, cytoskeletal organization and axonal transport similarly with primary motor neurons, but failed to reproduce synaptic connections (Durham, et al, 1993). This cell line is widely used in several studies related to ALS (Foran, et al, 2011;Tabata, et al, 2008;Vijayalakshmi, et al, 2009;Yang, et al, 2010). For example, NCS-34 cells were treated with the cerebrospinal fluid (CSF) collected from sporadic ALS patients.…”