d erivation of glutamatergic neurons from human pluripotent stem cells as a therapeutic intervention...
TRANSCRIPT
DERIVATION OF GLUTAMATERGIC NEURONS
FROM HUMAN PLURIPOTENT STEM CELLS AS A THERAPEUTIC
INTERVENTION FOR ALZHEIMER’S DISEASE
Joshue Leyva
REǀNOUS
Winter Meeting 1
01/27/14
“Efficient derivation of cortical glutamatergic neurons from human
pluripotent stem cells: A model system to study neurotoxicity in
Alzheimer's disease”
Neurobiology of Disease (2014): 62-72
Tandis Vazin, K. Aurelia Ball, Hui Luc, Hyungju Park, Yasaman Ataeijannati, Teresa Head-Gordon, Mu-ming Poo, David V. Schaffer
OUTLINE
Stem cells general reviewHuman embryonic stem cells (hESCs)Human induced pluripotent stem cells (hiPSCs)
Creating neuronal progenitor cellsGlutamatergic neuronsGABAergic neurons
Glutamatergic neurons as a therapeutic model
Differential neurotoxic selectivity on neuronal populations
• Adult (Ex. muscle stem cells, hematopoietic stem cells)
– Limited potential (multipotent)
– Rare
– Some are difficult to isolate
– Do not form teratomas
– In active clinical use
Stem cell types
• Embryonic (Ex. hESC or hiPSC)
– Can form all cell types (pluripotent)
– immortal in culture
– Can form tumors (teratomas)
– Methods to control differentiation poorly understood
Types of Pluripotent Stem Cells
Donovan and Gearhart Nature 2001
DERIVATION OF NEURAL PROGENITOR CELLS
Glutamatergic neurons1. Induce hPSCs or hESCs into neuronal lineage
by inhibiting SMAD signaling pathway2. Inhibit Shh signaling pathway to form the
dorsal telencephalon phenotype GABAergic neurons
1. Induce hPSCs or hESCsinto neuronal lineage by inhibiting SMAD signaling pathway
2. Proceed with Shh signaling pathway to form the ventral telencephalic neuronal phenotype
DERIVATION OF NEURAL PROGENITOR CELLS
74% Glutamatergic; 20% GABA 18% Glut; 62% GABAergic
GLUTAMATERGIC NEURONS AS A THERAPEUTIC MODEL
• Create Aβ globulomers (stable, soluble from of amyloid-beta)
• Treat hESC-derived glutamatergic neurons with 2 μM globulomers, get neuronal cell death after 72 h
GLUTAMATERGIC NEURONS AS A THERAPEUTIC MODEL
Progressive increase in glutamatergic neuron death with increasing Aβ globulomer concentration
GLUTAMATERGIC NEURONS AS A THERAPEUTIC MODEL
Upon adding the pre-fibrillar from of Aβ to an increased cell culture maturation time, Aβ binding to cell membrane and dendritic spines significantly increased
25 days 58 days
NEURONAL PHENOTYPE SENSITIVITIES TO AΒ
Method:
give hESC-derived cultures (primarily comprised of glutamatergic or GABAergic neurons) Aβ globulomer and assess globulomer binding and neuronal apoptosis
NEURONAL PHENOTYPE SENSITIVITIES TO AΒ
Glutamatergic neurons GABAergic neurons
Glutamatergic neurons exhibit progressively higher cell death with increasing concentrations of the globulomeric form of Aβ
NEURONAL PHENOTYPE SENSITIVITIES TO AΒ
Glutamatergic neurons GABAergic neurons
Quantitative analysis demonstrate a concentration dependent onset of apoptosis in glutamatergic neurons treated with Aβ globulomers (as seen by caspase-3)
NEURONAL PHENOTYPE SENSITIVITIES TO AΒ(QUANTITATIVE DATA)
CONCLUSIONS
Active manipulation with a Shh antagonist can leads to dorsal telencephalic NPCs, yielding glutamate expressing neurons
In the absence of Shh inhibition, NPCs adapted a ventral phenotype giving rise to GABAergic neurons
Glutamatergic neurons are more susceptible to Aβ toxicity and decrease in numbers with increasing Aβ concentrations