Abstract
Astrocytes are key players in the progression of amyotrophic lateral sclerosis (ALS). Previously, gene expression profiling of astrocytes from the pre-symptomatic stage of the SOD1G93A model of ALS has revealed reduced lactate metabolism and altered trophic support. Here, we have performed microarray analysis of symptomatic and late-stage disease astrocytes isolated by laser capture microdissection (LCM) from the lumbar spinal cord of the SOD1G93A mouse to complete the picture of astrocyte behavior throughout the disease course. Astrocytes at symptomatic and late-stage disease show a distinct up-regulation of transcripts defining a reactive phenotype, such as those involved in the lysosome and phagocytic pathways. Functional analysis of hexosaminidase B enzyme activity in the spinal cord and of astrocyte phagocytic ability has demonstrated a significant increase in lysosomal enzyme activity and phagocytic activity in SOD1G93A vs. littermate controls, validating the findings of the microarray study. In addition to the increased reactivity seen at both stages, astrocytes from late-stage disease showed decreased expression of many transcripts involved in cholesterol homeostasis. Staining for the master regulator of cholesterol synthesis, SREBP2, has revealed an increased localization to the cytoplasm of astrocytes and motor neurons in late-stage SOD1G93A spinal cord, indicating that down-regulation of transcripts may be due to an excess of cholesterol in the CNS during late-stage disease possibly due to phagocytosis of neuronal debris. Our data reveal that SOD1G93A astrocytes are characterized more by a loss of supportive function than a toxic phenotype during ALS disease progression and future studies should focus upon restorative therapies.
Highlights
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by progressive loss of motor neurons and resulting muscular weakness leading to death on average within 2–3 years of symptom onset
The transcriptomics study performed here is the first to complete an analysis of astrocytes from multiple stages of disease in the SOD1G93A mouse model of amyotrophic lateral sclerosis (ALS)
SOD1G93A astrocytes continue to decrease the expression of transcripts involved in the support of neurons such as those involved in maintaining homeostasis and neuronal support; steroid and cholesterol biosynthesis, neurotransmitter receptors and potassium and sodium ion channels important for sensing alterations in the synaptic space
Summary
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by progressive loss of motor neurons and resulting muscular weakness leading to death on average within 2–3 years of symptom onset. 10% of cases of ALS are familial (fALS) and multiple causative genes have been identified (Goodall et al, 2012). The most common known genetic cause of ALS is a mutation in the C9ORF72 gene (Dejesus-Hernandez et al, 2011; Renton et al, 2011), implicated in approximately 7% of sporadic cases (sALS) and 43% of fALS (Cooper-Knock et al, 2012). Mutations in SOD1 (Cu/Zn superoxide dismutase (1) are the second most common genetic cause of ALS, and account for 10–20% of fALS (Cudkowicz et al, 1997). Transgenic mice expressing mutant human SOD1 (mSOD1) are widely used as an animal model of ALS, allowing for detailed study of motor neurons and the surrounding glia during disease progression (Gurney et al, 1994; Turner and Talbot, 2008). Using the SOD1G93A model of ALS originally developed by Gurney et al (1994) we have developed the model on a homogeneous background featuring highly consistent disease progression (Mead et al, 2011), which allows analysis of discrete time-points in disease
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