Comparative neuronal differentiation of self-renewing neural progenitor cell lines obtained from human induced pluripotent stem cells

Verpelli, Chiara; Carlessi, Luigi; Bechi, Giulia; Poli, Elena Fusar; Orellana, Daniel; Heise, Christopher E.; Franceschetti, Silvana; Mantegazza, Renato; Mantegazza, Massimo; Delia, Domenico; Sala, Carlo

doi:10.3389/fncel.2013.00175

Cited by 30 publications

(33 citation statements)

References 53 publications

(59 reference statements)

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“…As previously reported (34), iPSC- were relatively immature at 10 weeks in culture (Figure 5). Qualitative analysis suggested that iPSC-neurons bearing the 15q11.2(BP1-BP2) deletion had altered dendritic morphology (Figure 5).…”

Section: Resultssupporting

confidence: 87%

“…PSD-95 labeling showed a punctate staining in iPSC-neurons from a control individual, similar to observed in postmortem human studies (38). In addition, consistent with other iPSC studies, PSD-95 appeared to be more diffuse in neurites which may represent relative immaturity in the iPSC-neurons (34). To test this possibility, further agnostic, unbiased analyses of dendritic architecture are needed in iPSC-neurons differentiated for longer periods.…”

Section: Discussionsupporting

confidence: 89%

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Genetic and Morphological Features of Human iPSC-Derived Neurons with Chromosome 15q11.2 (BP1-BP2) Deletions

Das¹,

Tapias

D’Aiuto³

et al. 2015

Complex Psychiatry

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Background: Copy number variation on chromosome 15q11.2 (BP1-BP2) causes a deletion of CYFIP1, NIPA1, NIPA2 and TUBGCP5. Furthermore, it also affects brain structure and elevates the risk for several neurodevelopmental disorders that are associated with dendritic spine abnormalities. In rodents, altered cyfip1 expression changes dendritic spine morphology, motivating analyses of human neuronal cells derived from induced pluripotent stem cells (iPSCs; iPSC-neurons). Methods: iPSCs were generated from a mother and her offspring, both carrying the 15q11.2 (BP1-BP2) deletion, and a non-deletion control. Gene expression in the deletion region was estimated using quantitative real-time PCR assays. Neural progenitor cells (NPCs) and iPSC-neurons were characterized using immunocytochemistry. Results:CYFIP1, NIPA1, NIPA2 and TUBGCP5 gene expression was lower in iPSCs, NPCs and iPSC-neurons from the mother and her offspring in relation to control cells. CYFIP1 and PSD-95 protein levels were lower in iPSC-neurons derived from the copy number variant-bearing individuals using Western blot analysis. Ten weeks after differentiation, iPSC-neurons appeared to show dendritic spines, and qualitative analysis suggested that dendritic morphology was altered in 15q11.2-deletion subjects compared with control cells. Conclusions: The 15q11.2 (BP1-BP2) deletion is associated with a reduced expression of four genes in iPSC-derived neuronal cells; it may also be associated with altered iPSC-neuron dendritic morphology.

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Section: Resultssupporting

confidence: 87%

Section: Discussionsupporting

confidence: 89%

Genetic and Morphological Features of Human iPSC-Derived Neurons with Chromosome 15q11.2 (BP1-BP2) Deletions

Das¹,

Tapias

D’Aiuto³

et al. 2015

Complex Psychiatry

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“…As primary neurons have been shown to facilitate the maturation of iPSC-derived neurons (Weick et al 2011;Verpelli et al 2013;Su et al 2015), we cultured iPSC-derived hNPCs together with primary rat cortical neurons. To identify iPSC-differentiated neurons among the co-culture system, we selectively expressed a fluorescent protein, tdTomato, behind a neuron-specific Synapsin 1 (Syn1) promoter in the hNPCs.…”

Section: Morphological and Functional Analysis Of Ipscdifferentiatedmentioning

confidence: 99%

Choline Ameliorates Disease Phenotypes in Human iPSC Models of Rett Syndrome

et al. 2016

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Rett syndrome (RTT) is a postnatal neurodevelopmental disorder that primarily affects girls. Mutations in the methyl-CpG-binding protein 2 (MECP2) gene account for approximately 95 % of all RTT cases. To model RTT in vitro, we generated induced pluripotent stem cells (iPSCs) from fibroblasts of two RTT patients with different mutations (MECP2 (R306C) and MECP2 (1155Δ32)) in their MECP2 gene. We found that these iPSCs were capable of differentiating into functional neurons. Compared to control neurons, the RTT iPSC-derived cells had reduced soma size and a decreased amount of synaptic input, evident both as fewer Synapsin 1-positive puncta and a lower frequency of spontaneous excitatory postsynaptic currents. Supplementation of the culture media with choline rescued all of these defects. Choline supplementation may act through changes in the expression of choline acetyltransferase, an important enzyme in cholinergic signaling, and also through alterations in the lipid metabolite profiles of the RTT neurons. Our study elucidates the possible mechanistic pathways for the effect of choline on human RTT cell models, thereby illustrating the potential for using choline as a nutraceutical to treat RTT.

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“…Dermal fibroblasts of a healthy, ethnically matched donor and their iPSCs derivatives were previously described by us. 18 Fibroblasts were cultured at 37 °C in a 5% CO 2 in E-MEM supplemented with 15% heat-inactivated FCS, 100 units/ml penicillin, 100 μ g/ml streptomycin, 100 μ M non-essential amino acids and 2 mM glutamine. Primary MEFs (PMEF-CFL, Millipore, Bedford, MA, USA) were cultured in DMEM/F12 supplemented with 20% heat-inactivated FCS, 100 units/ml penicillin, 100 μ g/ml streptomycin, 100 μ M non-essential amino acids and 2 mM glutamine.…”

Section: Methodsmentioning

confidence: 99%

Functional and molecular defects of hiPSC-derived neurons from patients with ATM deficiency

et al. 2014

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Loss of ataxia telangiectasia mutated (ATM) kinase, a key factor of the DNA damage response (DDR) pathway, causes the cancer predisposing and neurodegenerative syndrome ataxia-telangiectasia (A-T). To investigate the mechanisms of neurodegeneration, we have reprogrammed fibroblasts from ATM-null A-T patients and normal controls to pluripotency (human-induced pluripotent stem cells), and derived from these neural precursor cells able to terminally differentiate into post-mitotic neurons positive to >90% for β-tubulin III+/microtubule-associated protein 2+. We show that A-T neurons display similar voltage-gated potassium and sodium currents and discharges of action potentials as control neurons, but defective expression of the maturation and synaptic markers SCG10, SYP and PSD95 (postsynaptic density protein 95). A-T neurons exhibited defective repair of DNA double-strand breaks (DSBs) and repressed phosphorylation of ATM substrates (e.g., γH2AX, Smc1-S966, Kap1-S824, Chk2-T68, p53-S15), but normal repair of single-strand breaks, and normal short- and long-patch base excision repair activities. Moreover, A-T neurons were resistant to apoptosis induced by the genotoxic agents camptothecin and trabectedin, but as sensitive as controls to the oxidative agents. Most notably, A-T neurons exhibited abnormal accumulation of topoisomerase 1-DNA covalent complexes (Top1-ccs). These findings reveal that ATM deficiency impairs neuronal maturation, suppresses the response and repair of DNA DSBs, and enhances Top1-cc accumulation. Top1-cc could be a risk factor for neurodegeneration as they may interfere with transcription elongation and promote transcriptional decline.

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Comparative neuronal differentiation of self-renewing neural progenitor cell lines obtained from human induced pluripotent stem cells

Cited by 30 publications

References 53 publications

Genetic and Morphological Features of Human iPSC-Derived Neurons with Chromosome 15q11.2 (BP1-BP2) Deletions

Genetic and Morphological Features of Human iPSC-Derived Neurons with Chromosome 15q11.2 (BP1-BP2) Deletions

Choline Ameliorates Disease Phenotypes in Human iPSC Models of Rett Syndrome

Functional and molecular defects of hiPSC-derived neurons from patients with ATM deficiency

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