Identification of pathways of degeneration and protection in ALS using oculomotor neuron resistance as a tool.
Eva Hedlund, associate professor in the department of Neuroscience at Karolinska Institute in Sweden and Stefania Corti associate professor in the Neurology unit at University of Milan in Italy, work together on a project selected in 2012 by the Thierry Latran Foundation. First results were just published in Neuroscience.
Eva Hedlund presents for the Foundation the research conducted in her laboratory and her collaboration with Stefania Corti.
Neurodegenerative diseases are characterized by the selective loss of specific neuronal populations with corresponding distinct clinical features, even when the pathogenic proteins are ubiquitously expressed. The lethal motor neuron disease amyotrophic lateral sclerosis (ALS) is defined by the loss of somatic motor neurons that innervate muscles (for voluntary movement) in arms, legs, trunk and face, leading to muscle wasting. However, not all motor neurons are equally vulnerable to disease; certain groups of motor neurons are spared in ALS, including those in the oculomotor nucleus, controlling eye movement (Figure 1) and motor neurons in the Onuf’s nucleus, controlling pelvic muscles. The reasons for this differential vulnerability among motor neurons remain unknown.
Figure 1. Analysis of neuromuscular junctions in extraocular muscles and lumbricals in wild-type and SOD1G93A ALS mice. At their distal end motor neurons communicate directly with skeletal muscle; these specialized synapses are termed neuromuscular junctions (NMJs). NMJs in resistant extraocular muscles in 112-day-old early symptomatic SOD1G93A mice are fully innervated and show no signs of pathology (a,b). NMJs in vulnerable lumbrical muscles of 112-day-old SOD1G93A mice (c, d) show pathology with vacant endplates (arrowhead, d) and partly innervated endplates (* in d) compared to the fully innervated NMJs wild-type mice (c).
Experiments based on transgenic mouse ALS models and human motor neurons derived from patient fibroblasts (iPSCs) indicate that factors within motor neurons are important for initiation of degeneration in ALS. Thus, an analysis of the intrinsic properties of motor neurons displaying differential susceptibility to disease could reveal why certain motor neurons are resistant to degeneration and lead to future therapies to prevent the progressive motor neuron loss in ALS.
To elucidate molecular mechanisms of differential vulnerability in ALS we are performing RNA deep sequencing of resistant and vulnerable motor neuron groups isolated by laser capture microscopy (Figure 2)si vous manquez de place vous pouvez supprimer la fig 2 from wild-type mice and transgenic ALS mice. Through this analysis we identify all the mRNA transcripts made in a motor neuron at a particular time. This enables us to study how resistant cells modulate their transcriptome (that is, which genes are being activated) in response to an ALS-inducing mutation compared to cells that will degenerate. We expect that identified differences in mRNA transcripts will give us clues to mechanisms of motor neuron vulnerability and resistance.
Figure 2. Isolation of resistant oculomotor and vulnerable hypoglossal motor neurons using laser capture microscopy (LCM). Oculomotor (a, b) and hypoglossal (c, d) motor neurons were visualized by Histogene staining (Arcturus) and isolated by LCM.
Identified candidate transcripts and pathways, which appear preferential to resistant oculomotor motor neurons, are analyzed also in human post-mortem tissues for clinical relevance. Subsequently, we determine if these transcripts can modulate survival of motor neurons (both resistant and vulnerable) generated from pluripotent stem cells in vitro in ALS-like toxicity assays and in vivo in ALS mouse models.
We work very closely with the laboratory of Prof Stefania Corti at the University of Milano in screening candidate genes in vitro and in vivo and we have a joint grant from the Thierry Latran Foundation. Our long-term goal is to identify gene targets that could be regulated to protect vulnerable but intact motor neurons in ALS patients. That is, we hope to use some of the properties that render oculomotor motor neurons resistant to degeneration in ALS and confer these onto vulnerable motor neurons in order to protect these.