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Generation of an induced pluripotent stem cell line from a patient with epileptic encephalopathy caused by the CYFIP2 R87C variant

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Abstract

Induced pluripotent stem cells (iPSCs) opened the possibility to use patient cells as a model for several diseases. iPSCs can be reprogrammed from somatic cells collected in a non-invasive way, and then differentiated into any other cell type, while maintaining the donor´s genetic background. CYFIP2 variants were associated with the onset of an early form of epileptic encephalopathy. Studies with patients showed that the R87C variant seems to be one of the variants that causes more severe disease, however, to date there are no studies with a human cell model that allows investigation of the neuronal phenotype of the R87C variant. Here, we generated an iPSC line from a patient with epileptic encephalopathy caused by the CYFIP2 R87C variant. We obtained iPSC clones by reprogramming urinary progenitor cells from a female patient. The generated iPSC line presented a pluripotent stem cell morphology, normal karyotype, expressed pluripotency markers and could be differentiated into the three germ layers. In further studies, this cell line could be used as model for epileptic encephalopathy disease and drug screening studies.

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Table S1: Antibodies details; Table S2: Primers list; Figure S1: Expression of epithelial, renal epithelial, and fibroblast markers in UPC and NHDF; Figure S2: Expression of POU5F1 (OCT4), NANOG, and REX1 in ESC H1, iPSC R87C C8, and UPC. Figure S3: Expression of OCT4 and SSEA4 in ESC H1, iPSC R87C C8, and UPC. Figure S4: Expression of OCT4 and TRA-1-60 in ESC H1, iPSC R87C C8, and UPC. Figure S5: Expression of OCT4 and TRA-1-81 in ESC H1, iPSC R87C C8, and UPC. Figure S6: Expression of SSEA3 in ESC H1, iPSC R87C C8, and UPC. Figure S7: Expression of germ layers' markers in differentiated iPSC R87C C8. Figure S8: Expression of germ layers markers on ESC H1 and iPSC R87C C8 differentiation.

References

  1. Takahashi K, Yamanaka S. Induction of pluripotent stem cells from mouse embryonic and adult fibroblast cultures by defined factors. Cell. 2006;2:663–76.

    Article  Google Scholar 

  2. Mora C, Serzanti M, Consiglio A, Memo M, Dell’era P,. Clinical potentials of human pluripotent stem cells. Cell Biol Toxicol. 2017;33:351–60.

    Article  CAS  PubMed  Google Scholar 

  3. Pellock JM, Hrachovy R, Shinnar S, Baram TZ, Bettis D, Dlugos DJ, et al. Infantile spasms: a U.S. consensus report. Epilepsia. 2010;51:2175–89.

    Article  PubMed  Google Scholar 

  4. Nakashima M, Kato M, Aoto K, Shiina M, Belal H, Mukaida S, et al. De novo hotspot variants in CYFIP2 cause early-onset epileptic encephalopathy. Ann Neurol. 2018;83:794–806.

    Article  CAS  PubMed  Google Scholar 

  5. Konietzny A, Bär J, Mikhaylova M. Dendritic actin cytoskeleton: structure, functions, and regulations. Front Cell Neurosci. 2017;11:1–10.

    Article  CAS  Google Scholar 

  6. Cory GOC, Ridley AJ. Braking WAVEs. Nature. 2002;418:732–3.

    Article  CAS  PubMed  Google Scholar 

  7. Derivery E, Lombard B, Loew D, Gautreau A. The wave complex is intrinsically inactive. Cell Motil Cytoskelet. 2009;66:777–90.

    Article  CAS  Google Scholar 

  8. Schenck A, Bardoni B, Moro A, Bagni C, Mandel JL. A highly conserved protein family interacting with the fragile X mental retardation protein (FMRP) and displaying selective interactions with FMRP-related proteins FXR1P and FXR2P. Proc Natl Acad Sci USA. 2001;98:8844–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Begemann A, Sticht H, Begtrup A, Vitobello A, Faivre L, Banka S, et al. New insights into the clinical and molecular spectrum of the novel CYFIP2-related neurodevelopmental disorder and impairment of the WRC-mediated actin dynamics. Genet Med. 2021;23:543–54.

    Article  CAS  PubMed  Google Scholar 

  10. Zhang Y, Kang HR, Han K. Differential cell-type-expression of CYFIP1 and CYFIP2 in the adult mouse hippocampus. Anim Cells Syst (Seoul). 2019;23:380–3.

    Article  CAS  PubMed  Google Scholar 

  11. Pathania M, Davenport EC, Muir J, Sheehan DF, López-Doménech G, Kittler JT. The autism and schizophrenia associated gene CYFIP1 is critical for the maintenance of dendritic complexity and the stabilization of mature spines. Transl Psychiatry. 2014;4:1–11.

    Google Scholar 

  12. Kang M, Zhang Y, Kang HR, Kim S, Yi Y, Lee S, et al. The CYFIP2 p.Arg87Cys causes neurological defects and degradation of CYFIP2 running. Ann Neurol. 2023;93:155–63.

    Article  CAS  PubMed  Google Scholar 

  13. Steichen C, Si-Tayeb K, Wulkan F, Crestani T, Rosas G, Dariolli R, et al. Human induced pluripotent stem (hiPS) cells from urine samples: a non-integrative and feeder-free reprogramming strategy. Curr Protoc Hum Genet. 2017;2017:21.7.1-21.7.22.

    Google Scholar 

  14. Zhou T, Benda C, Dunzinger S, Huang Y, Ho JC, Yang J, et al. Generation of human induced pluripotent stem cells from urine samples. Nat Protoc. 2012;7:2080–9.

    Article  CAS  PubMed  Google Scholar 

  15. Moralli D, Yusuf M, Mandegar MA, Khoja S, Monaco ZL, Volpi EV. An improved technique for chromosomal analysis of human ES and iPS cells. Stem Cell Rev Rep. 2011;7:471–7.

    Article  PubMed  Google Scholar 

  16. Ahn LY, Coatti GC, Liu J, Gumus E, Schaffer AE, Miranda HC. An epilepsy-associated ACTL6B variant captures neuronal hyperexcitability in a human induced pluripotent stem cell model. J Neurosci Res. 2021;99:110–23.

    Article  CAS  PubMed  Google Scholar 

  17. Lee Y, Zhang Y, Kang H, Bang G, Kim Y, Kang HR, et al. Epilepsy- and intellectual disability-associated CYFIP2 interacts with both actin regulators and RNA-binding proteins in the neonatal mouse forebrain. Biochem Biophys Res Commun. 2020;529:1–6.

    Article  CAS  PubMed  Google Scholar 

  18. Zhang Y, Kang H, Lee Y, Kim Y, Lee B, Kim JY, et al. Smaller body size, early postnatal lethality, and cortical extracellular matrix-related gene expression changes of Cyfip2-null embryonic mice. Front Mol Neurosci. 2019;11:1–5.

    Article  Google Scholar 

  19. Lee SH, Zhang Y, Park J, Kim B, Kim Y, Lee SH, et al. Haploinsufficiency of Cyfip2 causes lithium-responsive prefrontal dysfunction. Ann Neurol. 2020;88:526–43.

    Article  CAS  PubMed  Google Scholar 

  20. Zhou T, Benda C, Duzinger S, Huang Y, Li X, Li Y, et al. Generation of induced pluripotent stem cells from urine. J Am Soc Nephrol. 2011;22:1221–8.

    Article  PubMed  PubMed Central  Google Scholar 

  21. Si-Tayeb K, Idriss S, Champon B, Caillaud A, Pichelin M, Arnaud L, et al. Urine-sample-derived human induced pluripotent stem cells as a model to study PCSK9-mediated autosomal dominant hypercholesterolemia. DMM Dis Models Mech [Internet]. 2016;9:81–90.

    CAS  Google Scholar 

  22. Bharadwaj S, Liu G, Shi Y, Wu R, Yang B, He T, et al. Multipotential differentiation of human urine-derived stem cells: potential for therapeutic applications in urology. Stem Cells. 2013;31:1840–56.

    Article  CAS  PubMed  Google Scholar 

  23. Cheng L, Lei Q, Yin C, Wang HY, Jin K, Xiang M. Generation of urine cell-derived non-integrative human iPSCs and iNSCs: a step-by-step optimized protocol. Front Mol Neurosci. 2017;10:1–8.

    Article  PubMed  PubMed Central  Google Scholar 

  24. Wezel F, Pearson J, Kirkwood LA, Southgate J. Differential expression of oct4 variants and pseudogenes in normal urothelium and urothelial cancer. Am J Pathol. 2013;183:1128–36.

    Article  CAS  PubMed  Google Scholar 

  25. Atlasi Y, Mowla SJ, Ziaee SAM, Gokhale PJ, Andrews PW. OCT4 spliced variants are differentially expressed in human pluripotent and nonpluripotent cells. Stem Cells. 2008;26:3068–74.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgements

The authors thank Dra Anny Robert and Dra Bruna Marcon of the Platform of Microscopy of Fiocruz PR for their help in image acquisition and training. We also thank MSc Crisciele Kuligovski of Cell Culture Lab for the mycoplasma testing. Lastly, we thank Wagner Nagib for the abstract graphical design.

Funding

This research was funded by Conselho Nacional de Desenvolvimento Científico e Tecnológico—CNPq/Instituto Carlos Chagas No. 15/2019—PROEP/ICC—442324/2019-7. PS received a scholarship from Conselho Nacional de Desenvolvimento Científico e Tecnológico—CNPq. The funders had no role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.

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Authors and Affiliations

  1. Stem Cell Basic Biology Laboratory, Instituto Carlos Chagas, Fiocruz PR, Curitiba, PR, 81310-020, Brazil

    Isabelle Leticia Zaboroski Silva, Rubens Gomes-Júnior, Evelin Brandão da Silva & Patrícia Shigunov

  2. Core for Cell Technology, School of Medicine, Pontifícia Universidade Católica do Paraná, Curitiba, PR, 80215-901, Brazil

    Isadora May Vaz & Valderez Ravaglio Jamur

  3. Gonçalo Moniz Institute, Oswaldo Cruz Foundation (FIOCRUZ), Salvador, 40296-710, Brazil

    Bruno Solano de Freitas Souza

  4. D’Or Institute for Research and Education (IDOR), Salvador, 41253-190, Brazil

    Bruno Solano de Freitas Souza

Authors
  1. Isabelle Leticia Zaboroski Silva
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  2. Rubens Gomes-Júnior
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  3. Evelin Brandão da Silva
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  4. Isadora May Vaz
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  5. Valderez Ravaglio Jamur
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  6. Bruno Solano de Freitas Souza
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  7. Patrícia Shigunov
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Contributions

ILZS designed and conducted most of the experimental work and was a major contributor to writing the manuscript. RG-J and EBS conducted experimental work. IMV and VRJ conducted the karyotyping acquisition and analysis. BSFS contributed to STR acquisition and analysis. PS contributed to study conception, supervision, and project administration.

Corresponding author

Correspondence to Patrícia Shigunov.

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The authors declare that they have no conflict of interest.

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The study has approval by the Research Ethics Committee (CEP) of Fiocruz, and by the Brazilian National Research Ethics Committee (CONEP), under opinion number 3.856.868, CAAE: 16918819.7.0000.5248.

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Silva, I.L.Z., Gomes-Júnior, R., da Silva, E.B. et al. Generation of an induced pluripotent stem cell line from a patient with epileptic encephalopathy caused by the CYFIP2 R87C variant. Human Cell 36, 2237–2246 (2023). https://doi.org/10.1007/s13577-023-00978-4

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