Morphological Changes in Neural Progenitors Derived from Human Induced Pluripotent Stem Cells and Transplanted into the Striatum of a Parkinson's Disease Rat Model

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Abstract

Introduction. Development of cell therapy for Parkinson's disease (PD) requires protocols based on transplantation of neurons derived from human induced pluripotent stem cells (hiPSCs) into the damaged area of the brain.

Objective: to characterize neurons transplanted into a rat brain and evaluate neural transplantation efficacy using a PD animal model.

Materials and methods. Neurons derived from hiPSCs (IPSRG4S line) were transplanted into the striatum of rats after intranigral injection of 6-hydroxydopamine (6-OHDA). Immunostaining was performed to identify expression of glial and neuronal markers in the transplanted cells within 2–24 weeks posttransplant.

Results. 4 weeks posttransplant we observed increased expression of mature neuron markers, decreased expression of neural progenitor markers, and primary pro-inflammatory response of glial cells in the graft. Differentiation and maturation of neuronal cells in the graft lasted over 3 months. At 3 and 6 months we detected 2 graft zones: one mainly contained the transplanted neurons and the other — human astrocytes. We detected human neurites in the corpus callosum and surrounding striatal tissue and large human tyrosine hydroxylase-expressing neurons in the graft.

Conclusion. With graft's morphological characteristics identified at different periods we can better understand pathophysiology and temporal patterns of new dopaminergic neurons integration and striatal reinnervation in a rat PD model in the long-term postoperative period.

About the authors

Dmitry N. Voronkov

Research Center of Neurology

Email: annaly-nevrologii@neurology.ru
ORCID iD: 0000-0001-5222-5322

Cand. Sci. (Biol.), senior researcher, Laboratory of neuromorphology

Russian Federation, Moscow

Alla V. Stavrovskaya

Research Center of Neurology

Author for correspondence.
Email: alla_stav@mail.ru
ORCID iD: 0000-0002-8689-0934

Cand. Sci. (Biol.), leading researcher, Laboratory of experimental pathology of the nervous system

Russian Federation, Moscow

Olga S. Lebedeva

Lopukhin Federal Research and Clinical Center of Physical and Chemical Medicine

Email: annaly-nevrologii@neurology.ru
ORCID iD: 0000-0003-0767-5265

senior researcher, Laboratory of cell biology

Russian Federation, Moscow

Wen Li

Health Sciences Institute, China Medical University

Email: annaly-nevrologii@neurology.ru
ORCID iD: 0000-0002-0383-0240

PhD, Professor, Institute of Health Sciences

Taiwan, Province of China, Shenyang

Artem S. Olshansky

Research Center of Neurology

Email: annaly-nevrologii@neurology.ru
ORCID iD: 0000-0002-5696-8032

Cand. Sci. (Biol.), senior researcher, Laboratory of experimental pathology of the nervous system

Russian Federation, Moscow

Anastasia S. Gushchina

Research Center of Neurology

Email: annaly-nevrologii@neurology.ru
ORCID iD: 0000-0003-3026-0279

researcher, Laboratory of experimental pathology of the nervous system

Russian Federation, Moscow

Marina R. Kapkaeva

Research Center of Neurology

Email: annaly-nevrologii@neurology.ru
ORCID iD: 0000-0002-2833-2897

junior researcher, Laboratory of neurobiology and tissue engineering

Russian Federation, Moscow

Alexandra N. Bogomazova

Lopukhin Federal Research and Clinical Center of Physical and Chemical Medicine

Email: annaly-nevrologii@neurology.ru
ORCID iD: 0000-0003-1549-1984

Head, Laboratory of cell biology

Russian Federation, Moscow

Maria A. Lagarkova

Lopukhin Federal Research and Clinical Center of Physical and Chemical Medicine

Email: annaly-nevrologii@neurology.ru
ORCID iD: 0000-0001-9594-1134

D. Sci. (Biol.), Corresponding Member of the Russian Academy of Sciences, Director

Russian Federation, Moscow

Sergey N. Illarioshkin

Research Center of Neurology

Email: annaly-nevrologii@neurology.ru
ORCID iD: 0000-0002-2704-6282

D. Sci. (Med.), Prof., Academician of the Russian Academy of Sciences, Deputy Director, Head, Brain Research Institute

Russian Federation, Moscow

References

  1. Lebedeva O.S., Lagarkova M.A. Pluripotent stem cells for modelling and cell therapy of Parkinson’s disease. Biochemistry (Moscow). 2018;83(9):1046–1056. doi: 10.1134/S0006297918090067
  2. Penney J., Ralvenius W.T., Tsai L.-H. Modeling Alzheimer’s disease with iPSC-derived brain cells. Mol. Psychiatry. 2020;25(1):148–167. doi: 10.1038/s41380-019-0468-3
  3. Schweitzer J.S., Song B., Herrington T.M. et al. Personalized iPSC-derived dopamine progenitor cells for Parkinson’s disease. N. Engl. J. Med. 2020;382(20):1926–1932. doi: 10.1056/NEJMoa1915872
  4. Wu R., Luo S., Yang H., Transplantation of neural progenitor cells generated from human urine epithelial cell-derived induced pluripotent stem cells improves neurological functions in rats with stroke. Dis. Med. 2020;29(156):53–64.
  5. Ghosh B., Zhang C., Ziemba K.S. et al. Partial reconstruction of the nigrostriatal circuit along a preformed molecular guidance pathway. Mol. Ther. Methods Clin. Dev. 2019; 14:217–227. doi: 10.1016/j.omtm.2019.06.008
  6. Kriks S., Shim J.-W., Piao J. et al. Dopamine neurons derived from human ES cells efficiently engraft in animal models of Parkinson’s disease. Nature. 2011;480(7378):547–551. doi: 10.1038/nature10648
  7. Arenas E., Denham M., Villaescusa J.C. How to make a midbrain dopaminergic neuron. Development. 2015;142(11):1918–1936. doi: 10.1242/dev.097394
  8. Engel M., Do-Ha D., Muñoz S.S., Ooi L. Common pitfalls of stem cell differentiation: a guide to improving protocols for neurodegenerative disease models and research. Cell. Mol. Life Sci. 2016;73(19):3693–3709. doi: 10.1007/s00018-016-2265-3
  9. Antonov S.A., Novosadova E.V., Kobylyansky A.G. et al. Expression and functional properties of NMDA and GABAA receptors during differentiation of human induced pluripotent stem cells into ventral mesencephalic neurons. Biochemistry (Moscow). 2019;84(3):310–320. doi: 10.1134/S0006297919030131
  10. Sefiani A., Geoffroy C.G. The potential role of inflammation in modulating endogenous hippocampal neurogenesis after spinal cord injury. Front. Neurosci. 2021;15:682259. doi: 10.3389/fnins.2021.682259
  11. Tomov N., Surchev L., Wiedenmann C. et al. Astrogliosis has different dynamics after cell transplantation and mechanical impact in the rodent model of Parkinson’s disease. Balkan Med. J. 2018;35(2):141–147. doi: 10.4274/balkanmedj.2016.1911
  12. Llorens-Bobadilla E., Zhao S., Baser A. et al. Single-cell transcriptomics reveals a population of dormant neural stem cells that become activated upon brain injury. Cell Stem Cell. 2015;17(3):329–340. doi: 10.1016/j.stem.2015.07.002
  13. Johann V., Schiefer J., Sass C. et al. Time of transplantation and cell preparation determine neural stem cell survival in a mouse model of Huntington’s disease. Exp. Brain Res. 2007;177(4):458–470. doi: 10.1007/s00221-006-0689-y
  14. Tom C.M., Younesi S., Meer E. et al. Survival of iPSC-derived grafts within the striatum of immunodeficient mice: Importance of developmental stage of both transplant and host recipient. Exp. Neurol. 2017;297:118–128. doi: 10.1016/j.expneurol.2017.07.018
  15. Kopach O. Monitoring maturation of neural stem cell grafts within a host microenvironment. World J. Stem Cells. 2019;11(11):982–989. doi: 10.4252/wjsc.v11.i11.982
  16. Holmqvist S., Lehtonen Š., Chumarina M. et al. Creation of a library of induced pluripotent stem cells from Parkinsonian patients. NPJ Parkinson Dis. 2016;2(1):16009. doi: 10.1038/npjparkd.2016.9
  17. Voronkov D.N., Stavrovskaya A.V., Guschina A.S. et al. Morphological characterization of astrocytes in a xenograft of human iPSC-derived neural precursor cells. Acta Naturae. 2022;14(3):100–108. doi: 10.32607/actanaturae.11710
  18. Paxinos G., Watson C. The Rat Brain in Stereotaxic Coordinates. 6th еd. San Diego; 2007.
  19. Krishnasamy S., Weng Y.C., Thammisetty S.S. et al. Molecular imaging of nestin in neuroinflammatory conditions reveals marked signal induction in activated microglia. J. Neuroinflammation. 2017;14(1):45. doi: 10.1186/s12974-017-0816-7
  20. Verwer R.W., Sluiter A.A., Balesar R.A. et al. Mature astrocytes in the adult human neocortex express the early neuronal marker doublecortin. Brain. 2007;130(12):3321-3335. doi: 10.1093/brain/awm264
  21. Sun W., Cornwell A., Li J. et al. SOX9 is an astrocyte-specific nuclear marker in the adult brain outside the neurogenic regions. J. Neurosci. 2017;37(17):4493–4507. doi: 10.1523/JNEUROSCI.3199-16.2017
  22. Sensenbrenner M., Lucas M., Deloulme J.-C. Expression of two neuronal markers, growth-associated protein 43 and neuron-specific enolase, in rat glial cells. J. Mol. Med. 1997;75(9):653–663. doi: 10.1007/s001090050149
  23. Harrill J.A., Chen H., Streifel K.M. et al. Ontogeny of biochemical, morphological and functional parameters of synaptogenesis in primary cultures of rat hippocampal and cortical neurons. Mol. Brain. 2015;8(1):10. doi: 10.1186/s13041-015-0099-9
  24. White R.B., Thomas M.G. Moving beyond tyrosine hydroxylase to define dopaminergic neurons for use in cell replacement therapies for Parkinson’s disease. CNS Neurol. Disord. Drug Targets. 2012;11(4):340–349. doi: 10.2174/187152712800792758

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2. Fig. 1. Assessment of rats' locomotor activity in an open field test following intranigral injection of 6-OHDA (A) and 3 weeks and 3 months posttransplant (В). *p < 0.05 compared to the control group.

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3. Fig. 2. Detection of key protein markers at different posttransplant periods, ×20. А — detection of Nes and Sox9, early neural progenitor markers, in the graft (at 2 weeks); B — activation of microglia (IBA1+) surrounding the graft cells (at 2 weeks); C — astrocytes (GFAP+) forming a glial scar around the graft (at 2 weeks); D — detection of SYP and DCX, a neural progenitor marker (at 4 weeks); E — detection of mature neurons (NeuN+) in the graft (at 12 weeks); F — growth of transplanted cells' projections (NSE+) into a rat's striatum (at 12 weeks).

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4. Fig. 3. Quantitative analysis of morphological changes in the graft 2–12 weeks posttransplant (changes in staining intensity of key protein mar- kers). Results are expressed as log2 of the fold change (y-axis' 1 equals a 2-fold increase). *p < 0.05 compared to 2 weeks posttransplant.

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5. Fig. 4. Graft 6 months posttransplant. А — mature neurons (PGP+/MTC+) in the graft's center (1) and human glial cells (PGP–/MTC+) in the peripheral area (2), ×10; В — human tyrosine hydroxylase-containing neurons in the graft (TH+/HNA+), ×40; С — mature human astroglia in the graft (ALDH+/HNA+), ×20.

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Copyright (c) 2023 Voronkov D.N., Stavrovskaya A.V., Lebedeva O.S., Li W., Olshansky A.S., Gushchina A.S., Kapkaeva M.R., Bogomazova A.N., Lagarkova M.A., Illarioshkin S.N.

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This work is licensed under a Creative Commons Attribution 4.0 International License.

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