Life Technologies

iPSC overview Patient-derived iPSCs offer exciting potential in both cell therapy and in vitro disease modeling by enabling access to cell populations that are otherwise unavailable from living donors. Efficient reprogramming of donor somatic cells to iPSCs plays a key role in the realization of this potential. We used the CytoTune®-iPS Sendai Reprogramming Kit to reprogram donor cells because it uses a non-integrating (RNA virus) and replication-deficient Sendai virus (SeV) vector [2,3], allowing resulting iPSCs to be free from the risk of potential multiple insertions or tumorigenicity. This kit has been shown to reprogram normal human foreskin fibroblasts, peripheral blood mononuclear cells, and CD34+ cells to generate integration-free iPSCs with high efficiency, and was recently used to generate iPSCs that were differentiated to clinicalgrade retinal precursor cells under xeno-free conditions [4]. In the work presented here, fibroblasts from skin biopsies of three PD donors (PD-1, PD-2, and PD-3), one Multiple Systems Atrophy (MSA) donor, and two age-matched nondiseased control individuals (Ctrl-1 and Ctrl-2) were efficiently reprogrammed to iPSCs. MSA has been described as a more severe form of PD, with a more rapid disease progression after the first appearance of symptoms [5]. Fibroblasts from PD donors had previously been genotyped, and were known to contain LRRK2, GBA, and PARK2 mutations, whereas the MSA line was from a sporadic case with no mutations in genes known to be associated with PD. The resulting iPSCs were evaluated using both cellular and genetic analysis tools to confirm successful reprogramming before moving to further studies, and demonstrated that the iPSCs were transgene-free and karyotypically normal, 8 Life Technologies | Parkinson's cell model expressed known pluripotency markers, and were able to form embryoid bodies (EBs), spheroid structures that present three germ layer lineages, thereby confirming the cells' ability to differentiate. Gene expression profiles clearly distinguished these iPSCs from their parental fibroblasts and demonstrated a high level of consistency with expression patterns from control iPSC and H9 embryonic stem cell (ESC) lines. As a quality-control procedure, each line was "genetically fingerprinted" at both the fibroblast and the iPSC stage to verify cell line identity and to provide a means of verification throughout subsequent applications. Six cell lines were generated: •Three PD donors (PD-1, PD-2, and PD-3) •One Multiple Systems Atrophy (MSA) donor •Two age-matched non-diseased control individuals (Ctrl-1 and Ctrl-2) How Does Sendai Virus Reprogram Cells?

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