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Pizza club - October 2016 - Lisa

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Midbrain-like organoids from Human Pluripotent Stem Cells contain functional dopaminergic and neuromelanin-producing neurons

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Pizza club - October 2016 - Lisa

  1. 1. Journal Club - Lisa Smits 25th October 2016
  2. 2. Why 3D cell culture? What are organoids? • Multiple organ-specific cell types • Spatial organisation • Physiological functions Utility in research: • Disease mechanisms • Regenerative medicine • Drug testing Yin et al., 2016
  3. 3. Development of mDA neurons Image adapted from Lumsden et al., 1995, Gale et al., 2008, Aguila et al., 2012 midbrain identity: OTX2, FOXA2, LMX1A mDA neuron progenitor: FOXA2, LMX1A, NURR1 mDA neurons: TH, DAT, FOXA2, LMX1A
  4. 4. Method - Derivation of hMLOs
  5. 5. Characterisation of hMLOs Ki67, EdU: proliferation MAP2, MASH1: neuronal OTX2: midbrain NURR1: mDA progenitor • transitioned from progenitors to mature neurons • developing neuroectoderm toward floor plate
  6. 6. Identification of mDA neurons in hMLOs FOXA2, OTX2, LMX1A: midbrain TH, DAT: dopamine • time-dependent induction of floor plate precursors • differentiation into mDA neurons within developing hMLOs
  7. 7. Transcriptional characterisation of hMLOs in vivo in vitro • gene expression profiles of hMLOs reflects prenatal midbrain identity • contain cells that are not present in 2D Oligodendrocytes Glial cells
  8. 8. Identification of neuromelanin in hMLOs • NM granules secreted from mDA neurons within hMLOs • role of NM unclear
  9. 9. Functional maturation of DA neurons in hMLOs • mDA neurons exhibited mature neuronal properties • synapse formation with other neurons • DA production within hMLO Biocytin/THM
  10. 10. • apico-basal polarity of midbrain specific cells • identification of neuromelanin in mDA neurons • electrophysiological activity and suppression of mDA firing • hMLO transcriptome profile closer to prenatal midbrain compared to 2D mDA neuronal cultures • detection of dopamine using HPLC Summary Thanks for your attention!

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