Objective: To develop a biorepository of human induced pluripotent stem cells for studying Parkinson’s disease(PD).

Background: Genetically engineered stem cells called “induced pluripotent stem cells” or iPSCs can be developed using blood or skin cell biopsies from patients with Parkinson’s disease(PD). iPSCs can be differentiated into dopaminergic neurons that behave similar to the PD patient(1).

Method: In this pilot study, 23 patients with PD were enrolled. Either 10 ml of blood or 3 mm of skin sample were collected after informed consent was obtained. The samples were processed using non-integrative episomal vectors system. Three episomal vectors, pCXLE-hOCT3/4-shp53, pCXLE-hSK and pCXLE-hUL were used to transfect peripheral blood mononuclear cells and human fibroblasts using the Amaxa Nucleofector as previously described (2, 3). The vectors and reprogramming factor genes were cleared from the cells as the iPSC colonies expanded. The iPSCs colonies were characterized for their pluripotency and propensity to give rise to neural lineage according to set standard we identified(4). The dopaminergic cells were identified by the expression of tyrosine hydroxylase, the rate limiting enzyme for the biosynthesis of dopamine.

Results: In this initial phase, 23 patients (14 male)(6 Hispanic) with a mean age of 75 yrs (53-82 yrs) were enrolled. Three iPSC clones were derived and characterized from each patient. The iPSC colonies expressed the pluripotency markers Oct4, Nanog, Tra1-60 and SAEA4. Neural stem cells (NSCs) were derived from these iPSCs using specific mitogenic factors(5). The multipotency of these NSCs was confirmed by the expression of neural lineages; GFAP for astrocytes, O4 for oligodendrocytes and beta-III tubulin for neurons. We are currently growing organoids out of these iPSC lines, which will be used to study the disease mechanism, perform drug screening and link genotype, phenotype to the DNA-seq and RNA-seq data.

Conclusion: The possibility to generate specialized organ-specific differentiated cells using somatic cells induced to become iPSCs opens the door for unique opportunities such as understanding cellular developmental principles, investigate the role of variant genes and epigenetic changes associated with PD, link genotype, phenotype to the DNA-seq and RNA-seq data, identify novel targets for drug screening and ultimately develop effective neuroprotective or regenerative therapies in PD.

References: 1) Daadi MM, Grueter BA, Malenka RC, Redmond DE, Jr., Steinberg GK (2012) Dopaminergic neurons from midbrain-specified human embryonic stem cell-derived neural stem cells engrafted in a monkey model of Parkinson’s disease. PLoS ONE 7: e41120. 2) Hong H., Roy-Choudhury G., Kim J., Daadi M.M. (2019). Isolation and Differentiation of Self-Renewable Neural Stem Cells from Marmoset Induced Pluripotent Stem Cells. Methods Mol. Biol. 1919: 199-204. 3) Daadi M.M. (2019). Generation of Neural Stem Cells From Induced Pluripotent Stem Cells. Methods Mol. Biol. 1919: 1-7. 4) Yang G., Hong H., Torres A., Malloy K.E., Choudhury G.R., Kim J., Daadi M.M. (2019). Reference Transcriptome for Deriving Nonhuman Primate Induced Pluripotent Stem Cells. Methods Mol. Biol. 1919: 175-186. 5) Daadi M.M. (2019). Differentiation of Neural Stem Cells Derived from Induced Pluripotent Stem Cells into Dopaminergic Neurons. Methods Mol. Biol. 1919: 89-96.

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MDS Abstracts - https://www.mdsabstracts.org/abstract/texas-biorepository-of-human-induced-pluripotent-stem-cells-for-studying-parkinsons-disease/