Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disorder marked by the loss of motor neurons (MNs) in the brain and spinal cord, leading to fatally debilitating weakness. Because this disease predominantly affects MNs, we aimed to characterize the distinct expression profile of that cell type in order to elucidate underlying disease mechanisms and identify novel targets that inform on MN health during ALS disease timecourse. microRNAs (miRNAs) are short, non-coding RNAs that can shape the expression profile of a cell and, consequently, often exhibit cell type enriched expression. To determine MN-enriched miRNA expression, we utilized Cre recombinase-dependent miRNA tagging and affinity purification in mice. By defining the in vivo miRNA expression of MNs, all neurons, astrocytes, and microglia, we then focused on MN-enriched miRNAs via a comparative analysis, and found they may functionally distinguish MNs postnatally from other spinal neurons. Characterizing the levels of the MN-enriched miRNAs in CSF harvested from ALS models of MN disease demonstrated that one miRNA (miR-218) tracked with MN loss and was responsive to an ALS therapy in rodent models. Thus, we have employed cellular expression profiling tools to define the distinct miRNA expression of MNs, which is likely to enrich future studies of MN disease. This approach enabled development of a novel, drug-responsive marker of MN disease in ALS rodents.
Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disease in which motor neurons (MNs) in the brain and spinal cord are selectively lost. In order to develop tools to understand the distinct expression profiles of MNs and ultimately, monitor MN disease progression, we identified small regulatory RNAs (miRNAs) that were highly enriched or exclusive in MNs. The signal for one of these MN-enriched miRNAs is detectable in spinal tap biofluid from an ALS rat model, where its levels change as disease progresses, suggesting it may be a clinically useful marker of disease status. Furthermore, rats treated with an ALS therapy have restored expression of this MN RNA marker, making it a MN-specific and drug-responsive marker for ALS rodents.
The authors declare no other competing financial interests.
We thank Gregory F. Wu and Conrad C. Weihl for helpful comments and critique on this work. We thank Pak Chan, Stanford University, for providing the hSOD1 WT rats. We thank Klaus-Armin Nave, Max-Planck, for the Cnp1 Cre mouse. We thank the Division of Neuropathology, Department of Pathology and Immunology for assistance with autopsy tissue acquisition. We thank Amber Salter, PhD for help with statistical analyses. We also thank Elena Fisher for assistance with manuscript preparation. This work was supported by Project5 for ALS to T.M.M.; the National Institute of Neurological Disorders and Stroke [K08NS074194 and R01NS078398 to T.M.M., F31NS077781 to E.D.K., F31NS092340 to M.N.L.]; the Robert Packard Center for ALS Research to T.M.M.; and the University of Missouri Spinal Cord Injuries Research Program to T.M.M., and the Hope Center for Neurological Disorders. J.D.D. is supported in part by a NARSAD Independent Investigator grant from the Brain and Behavior Research Foundation. This work was also supported by access to equipment made possible by the Hope Center for Neurological Disorders, and the Departments of Neurology and Psychiatry at Washington University School of Medicine. Data from Nikon A1Rsi Confocal were performed in part through the use of Washington University Center for Cellular Imaging (WUCCI) supported by Washington University School of Medicine, The Children's Discovery Institute of Washington University and St. Louis Children's Hospital, the Foundation for Barnes-Jewish Hospital and the National Institute for Neurological Disorders and Stroke (NS086741).