Abstract
Modelling rare neurogenetic diseases to develop new therapeutic strategies is highly challenging. The use of human-induced pluripotent stem cells (hiPSCs) is a powerful approach to obtain specialized cells from patients. For hereditary peripheral neuropathies, such as Charcot–Marie–Tooth disease (CMT) Type II, spinal motor neurons (MNs) are impaired but are very difficult to study. Although several protocols are available to differentiate hiPSCs into neurons, their efficiency is still poor for CMT patients. Thus, our goal was to develop a robust, easy, and reproducible protocol to obtain MNs from CMT patient hiPSCs. The presented protocol generates MNs within 20 days, with a success rate of 80%, using specifically chosen molecules, such as Sonic Hedgehog or retinoic acid. The timing and concentrations of the factors used to induce differentiation are crucial and are given hereby. We then assessed the MNs by optic microscopy, immunocytochemistry (Islet1/2, HB9, Tuj1, and PGP9.5), and electrophysiological recordings. This method of generating MNs from CMT patients in vitro shows promise for the further development of assays to understand the pathological mechanisms of CMT and for drug screening.
Highlights
Peripheral nerves are critical for the functioning of the nervous system, as they forward information to the spinal cord and encephalon and provide the periphery with adapted signals
Based on an extensive review of the literature and results obtained in our laboratory, we developed a robust and reproducible protocol to improve motor neurons (MNs) differentiation of human-induced pluripotent stem cells (hiPSCs) obtained from Charcot–Marie–Tooth disease (CMT) patients
We launched this study to define a robust protocol to obtain and differentiate cells obtained from CMT patients into MNs
Summary
Peripheral nerves are critical for the functioning of the nervous system, as they forward information to the spinal cord and encephalon and provide the periphery (muscles, organs, skin, blood vessels) with adapted signals. Peripheral nerve illnesses constitute an important source of medical problems and a large group of neurological diseases of various origins. Brain Sci. 2020, 10, 407; doi:10.3390/brainsci10070407 www.mdpi.com/journal/brainsci. Brain Sci. 2020, 10, 407 peripheral neuropathies, such as Charcot–Marie–Tooth disease (CMT) disease, have a prevalence of 1:2500 and often affect patients during their entire lives. The use of next-generation sequencing has tremendously improved the molecular diagnosis of these diseases in recent years by efficiently determining the mutations involved. Even when a gene mutation is identified, the molecular and cellular pathways involved in the pathophysiology remain difficult to decipher. Because of the numerous mutations and various genes involved, animal models are of limited utility and are highly difficult to study, aside from the potential ethical problems. In vitro cellular models appear to be a promising path for the expeditious development of therapeutic strategies
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a callDisclaimer: All third-party content on this website/platform is and will remain the property of their respective owners and is provided on "as is" basis without any warranties, express or implied. Use of third-party content does not indicate any affiliation, sponsorship with or endorsement by them. Any references to third-party content is to identify the corresponding services and shall be considered fair use under The CopyrightLaw.
