Stem cells assist in the development of treatments for neurodegenerative diseases, including Alzheimer’s disease & Parkinson’s disease . iPS cells derived from patient-specific somatic cells will avoid complications relating to immune rejection. Currently, researchers are developing stem cell-based therapies for PD using induced pluripotent stem (iPS) cells. iPS cells can differentiate into cells from any of the three germ layers, including neural stem cells (NSCs). Transplantation of neural stem cells (NSCs) is an emerging therapy for treating neurological disorders by restoring neuronal function. In this study, shRNA was used to knock down the expression of mutant α-synuclein (SNCA) in iPS cells that were generated from SNCA A53T transgenic mice, and these iPS cells were differentiated to NSCs. After injecting these NSCs into SNCA A53T mice, the therapeutic effects of these cells were evaluated. Transplantation of neural stem cells produced from SNCA A53T iPS cells with knocking down SNCA not only improved SNCA A53T mice coordination abilities, balance abilities, and locomotor activities but also significantly prolonged their lifespans. An innovative therapeutic approach that combines stem cell therapy and gene therapy will be used for the treatment of Parkinson’s disease.

Keywords: Stem cells, Induced pluripotent stem (iPS) cells, Knockdown, gene editing

Here, we describe a novel therapeutic approach for the treatment of familial PD. We collected the fibroblasts from SNCA A53T mice and reprogrammed them into iPS cells. We then generated iPS-shSNCA cells by knocking down the expression of mutated SNCA in these iPS cells. Finally, we differentiated iPS-shSNCA cells into neural stem cells (NSCs) and transplanted these NSCs into cortical areas of SNCA A53T transgenic mice to evaluate the therapeutic effects of NSCs.

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Fig. 1: Generation of iPS cells from mouse embryonic fibroblasts.

Fig. 2: Differentiation of the iPS cells into neural stem cells.

Fig. 3: Evaluation of the knockdown efficiencies of SNCA shRNAs.

Fig. 4: Exploration of the therapeutic effects of NSCs and NSC-shSNCA cells on behavioral tests.

To verify the successful transplantation, the brains of each group of mice in the behavior experiment were collected and preserved in paraformaldehyde. The samples were stained for the dopaminergic neuron marker tyrosine hydroxylase (TH).

Fig. 5: Exploration of the TH-positive and apoptotic cells in PD mice brains.

Gene therapy and cellular engineering techniques have enhanced the therapeutic effects of NSC transplantations. Several strategies have been developed to silence SNCA, including RNA interference, which targets SNCA directly, or microRNAs, which target SNCA’s 3’-untranslated region of SNCA. we demonstrated the potent effects of NSCs derived from mutant SNCA knockdown-iPS cells on treating PD and provided experimental evidence for further clinical application of iPS cells in cell-based therapeutics for PD. As multiple genes were altered in PD, NSCs derived from multiple gene-edited iPS cells may be more effective in treating PD.

Fig. 6: Schematic illustration of genetically edited iPS cell-derived NSCs rescue Parkinson’s disease.

The knockdown of mutant SNCA in iPS cells can provide a suitable source of NSCs for the treatment of Parkinson’s disease caused by mutations in SNCA. This innovative therapeutic approach that combines stem cell therapy and gene therapy will be used for the treatment of Parkinson’s disease.