Oxidative Stress Research on Huntington's Disease Neurons Using Genetically Encoded Biosensors
Abstract
Many neurodegenerative diseases, including Huntington’s disease (HD), are associated with oxidative stress in the neurons of the brain. Genetically encoded biosensorsare useful for studying these processesin vitro. Human cell cultures expressing the biosensors can serve as a cell model for developing and testing effective agents that reduce oxidative stress. In this work, transgenes encoding biosensors of glutathione oxidative potential(Grx1-roGFP2) with cytoplasmic and mitochondrial localization were introduced into human induced pluripotent stem cells of a healthy donor and an HD patient using CRISPR/Cas9-mediated genome editing. The cells were subsequently differentiated into medium spiny neurons of the striatum. The expression of the biosensors was detected in the iPSCs, neuronal precursors and mature neurons.The obtained cells could be used to study the redox potential of glutathione in HD neurons and to screen for new drug compounds aimed at reducing oxidative stress.
Keywords: genetically encoded biosensors, Huntington’s disease, induced pluripotent stem cells, medium spiny neurons, oxidative stress, glutathione, Grx1-roGFP2
References
[1] Bilan D.S., Lukyanov S.A., Belousov V.V.Genetically Encoded Fluorescent Redox Sensors]. Bioorganic chemistry. 2015 41 (3) 259-274
[2] Lee JM, Ramos EM, Lee JH et al. CAG repeat expansion in Huntington diseasedetermines age at onset in a fully dominant fashion. Neurology. 2012 78(10):690-695. doi: 10.1212/WNL.0b013e318249f683.
[3] Grigor’eva, Elena & Malankhanova, Tuyana & Surumbayeva, Aizhan et al. Generation of induced pluripotent stem cell line, ICGi007-A, by reprogramming peripheral blood mononuclear cells from a patient with Huntington’s disease. Stem Cell Research. 2019. 34. 101382-101382. 10.1016/j.scr.2018.101382.
[4] Ustyantseva EI, Medvedev SP, Vetchinova AS, Minina JM, Illarioshkin SN, Zakian SM. A Platform for Studying Neurodegeneration Mechanisms Using Genetically Encoded Biosensors. Biochemistry (Mosc). 2019;84(3): 299–309.
[5] Pellenz S, Phelps M, Tang W, Hovde BT, Sinit RB, Fu W, Li H, Chen E, Monnat RJ Jr. New Human Chromosomal Sites with ”Safe Harbor” Potential for Targeted Transgene Insertion. Hum Gene Ther. 2019 Jul;30(7):814-828. doi: 10.1089/hum.2018.169.
[6] Grigor’eva EV, Malankhanova TB, Surumbayeva A et al. Generation of GABAergic striatal neurons by a novel iPSC differentiation protocol enabling scalability and cryopreservation of progenitor cells. Cytotechnology. 2020 72(5):649-663. doi: 10.1007/s10616-020-00406-7