Introduction to Thematic Minireview Series: Development of Human Therapeutics Based on Induced Pluripotent Stem Cell (iPSC) Technology

Rao, Mahendra S.; Gottesfeld, Joel M.

doi:10.1074/jbc.r113.543652

Cited by 8 publications

(2 citation statements)

References 6 publications

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“…Once optimized for cell number and purity, iPSC-derived cells could be used to build upon the success of a prior HCS study that uncovered small molecules which improve peroxisome assembly in PBD-ZSD patient fibroblasts [ 18 ]. Likewise, we note the emerging role of iPSCs in toxicology assays for potential liabilities of therapeutic agents [ 106 ] and the possibility of uncovering environmental exposures that could more severely impact patients with PBD-ZSD and individuals with other diseases associated with peroxisomal dysfunction. In a broader context, the results of HCS using PBD-ZSD patient-derived neural and hepatic cells could help address fundamental questions regarding the potential benefits of evaluating multiple patient cell types in drug discovery efforts for a variety of disorders.…”

Section: Discussionmentioning

confidence: 99%

Induced pluripotent stem cell models of Zellweger spectrum disorder show impaired peroxisome assembly and cell type-specific lipid abnormalities

et al. 2015

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IntroductionZellweger spectrum disorder (PBD-ZSD) is a disease continuum caused by mutations in a subset of PEX genes required for normal peroxisome assembly and function. They highlight the importance of peroxisomes in the development and functions of the central nervous system, liver, and other organs. To date, the underlying bases for the cell-type specificity of disease are not fully elucidated.MethodsPrimary skin fibroblasts from seven PBD-ZSD patients with biallelic PEX1, PEX10, PEX12, or PEX26 mutations and three healthy donors were transduced with retroviral vectors expressing Yamanaka reprogramming factors. Candidate induced pluripotent stem cells (iPSCs) were subject to global gene expression, DNA methylation, copy number variation, genotyping, in vitro differentiation and teratoma formation assays. Confirmed iPSCs were differentiated into neural progenitor cells (NPCs), neurons, oligodendrocyte precursor cells (OPCs), and hepatocyte-like cell cultures with peroxisome assembly evaluated by microscopy. Saturated very long chain fatty acid (sVLCFA) and plasmalogen levels were determined in primary fibroblasts and their derivatives.ResultsiPSCs were derived from seven PBD-ZSD patient-derived fibroblasts with mild to severe peroxisome assembly defects. Although patient and control skin fibroblasts had similar gene expression profiles, genes related to mitochondrial functions and organelle cross-talk were differentially expressed among corresponding iPSCs. Mitochondrial DNA levels were consistent among patient and control fibroblasts, but varied among all iPSCs. Relative to matching controls, sVLCFA levels were elevated in patient-derived fibroblasts, reduced in patient-derived iPSCs, and not significantly different in patient-derived NPCs. All cell types derived from donors with biallelic null mutations in a PEX gene showed plasmalogen deficiencies. Reporter gene assays compatible with high content screening (HCS) indicated patient-derived OPC and hepatocyte-like cell cultures had impaired peroxisome assembly.ConclusionsNormal peroxisome activity levels are not required for cellular reprogramming of skin fibroblasts. Patient iPSC gene expression profiles were consistent with hypotheses highlighting the role of altered mitochondrial activities and organelle cross-talk in PBD-ZSD pathogenesis. sVLCFA abnormalities dramatically differed among patient cell types, similar to observations made in iPSC models of X-linked adrenoleukodystrophy. We propose that iPSCs could assist investigations into the cell type-specificity of peroxisomal activities, toxicology studies, and in HCS for targeted therapies for peroxisome-related disorders.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-015-0149-3) contains supplementary material, which is available to authorized users.

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Section: Discussionmentioning

confidence: 99%

Induced pluripotent stem cell models of Zellweger spectrum disorder show impaired peroxisome assembly and cell type-specific lipid abnormalities

et al. 2015

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“…And finally, it is important to provide functional activity of transplanted iPSCs. Despite all these difficulties, rapidly developing technologies of cellular reprogramming are at present among most perspective in the molecular medicine [37].…”

Section: Reverse Transcription Polymerase Chain Reaction (Rtpcr)mentioning

confidence: 99%

A Platform for Studying Molecular and Cellular Mechanisms of Parkinson’s Disease Based on Human Induced Pluripotent Stem Cells

Новосадова¹,

Nekrasov²,

Честков³

et al. 2016

Sovrem Tehnol Med

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The aim of the study was to obtain induced pluripotent stem cells (iPSCs) from patients with various forms of Parkinson's disease (PD), and to create on this basis a platform for studying the pathogenesis of the disease at the molecular and cellular level with the development of a protocol of the stem cell differentiation.Materials and Methods. iPSCs were derived from cultured skin fibroblasts, taken from five patients with various forms of PD (PARK8, PARK2, GBA-associated and sporadic forms), and reprogrammed with the help of lentiviral vectors and on the basis of Sendai virus. The obtained iPSCs clones were cultured to the stage of embryonic bodies and, after spontaneous differentiation, stained immunocytochemically. Gene expression and neural markers in these iPSCs lines were analysed using reverse transcription polymerase chain reaction.Results. The obtained iPSCs clones had a normal 46 XY karyotype, stained specifically with Oct4, Nanog, TRA-1-81 and SSEA-4 antibodies, and expressed marker genes responsible for maintaining the pluripotent condition. In the cultures of differentiated iPSCs, cells positively stained for the markers of the three primary germ layers (ectoderm, mesoderm, and endoderm) have been revealed. An effective protocol of iPSCs differentiation into dopaminergic neurons has been worked out, and confirmed by the expression of the specific markertyrosine hydroxylase enzyme.Conclusion. On the basis of explicitly characterized iPSCs from patients with various forms of PD and the developed cellular protocol, a platform for studying the pathogenesis of PD at the molecular and cellular level has been created. Obtaining cell population enriched with dopaminergic neurons opens a perspective for their application for personalized cell replacement PD therapy.

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Characterizing Pluripotent Stem Cells Using the TaqMan® hPSC ScorecardTM Panel

Fergus

Quintanilla

Lakshmipathy

2014

Methods in Molecular Biology

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Rapid technological developments for the efficient generation of footprint-free induced pluripotent stem cells (iPSC) enabled the creation of patient-specific iPSC for downstream applications in drug discovery and regenerative medicine. However, the large number of iPSCs, generated from diverse genetic backgrounds using various methods and culture conditions, created a steep challenge for rapid characterization and a demand for standardized methods. Current methods rely on a combination of in vitro and in vivo cellular analyses based on the expression of markers of self-renewal and the ability of the cells to differentiate into cell types representative of the three germ layers as a confirmation of functional pluripotency. These methods, though informative and extensively used, are not ideal for parallel analyses of large numbers of samples and hence not amenable to high-throughput environments. Recently, genetic and epigenetic expression signatures were used to define and confirm cell states, thus providing a surrogate molecular assay that can potentially replace complex in vivo cellular assays such as teratoma formation. In this chapter, we describe a molecular assay for rapid characterization and standardization of pluripotent stem cells. The TaqMan(®) hPSC Scorecard™ Panel is a comprehensive gene expression real-time PCR assay that consists of 94 individual q-PCR assays comprised of a combination of control, housekeeping, self-renewal, and lineage-specific genes. The resulting expression data set is analyzed using cloud-based analysis software that compares the expression pattern against a reference standard composed of multiple functionally validated ESC and iPSC lines. This system was successfully used to test several ESC and iPSC lines in their undifferentiated states to confirm their signatures of self renewal, as well as their terminally differentiates states, via spontaneous differentiation and directed differentiation into specific lineages, to determine the lines' differentiation potential. This genetic analysis tool, together with the flexibility to utilize varying sample inputs and preparation methods, provides a rapid method to confirm functional pluripotency of ESCs and iPSCs.

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Introduction to Thematic Minireview Series: Development of Human Therapeutics Based on Induced Pluripotent Stem Cell (iPSC) Technology

Cited by 8 publications

References 6 publications

Induced pluripotent stem cell models of Zellweger spectrum disorder show impaired peroxisome assembly and cell type-specific lipid abnormalities

Induced pluripotent stem cell models of Zellweger spectrum disorder show impaired peroxisome assembly and cell type-specific lipid abnormalities

A Platform for Studying Molecular and Cellular Mechanisms of Parkinson’s Disease Based on Human Induced Pluripotent Stem Cells

Characterizing Pluripotent Stem Cells Using the TaqMan® hPSC ScorecardTM Panel

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