Sundberg M, Tochitsky I, Buchholz DE, Winden K, Kujala V,
Kapur K, Cataltepe D, Turner D, Han MJ, Woolf CJ, Hatten ME, Sahin M. Purkinje
cells derived from TSC patients display hypoexcitability and synaptic deficits
associated with reduced FMRP levels and reversed by rapamycin. Mol
Psychiatry. 2018 Feb 15. doi:10.1038/s41380-018-0018-4. [Epub ahead of print]
Abstract
Accumulating evidence suggests that cerebellar dysfunction
early in life is associated with autism spectrum disorder (ASD), but the
molecular mechanisms underlying the cerebellar deficits at the cellular level
are unclear. Tuberous sclerosis complex (TSC) is a neurocutaneous disorder that
often presents with ASD. Here, we developed a cerebellar Purkinje cell (PC)
model of TSC with patient-derived human induced pluripotent stem cells (hiPSCs)
to characterize the molecular mechanisms underlying cerebellar abnormalities in
ASD and TSC. Our results show that hiPSC-derived PCs from patients with
pathogenic TSC2 mutations displayed mTORC1 pathway hyperactivation, defects in
neuronal differentiation and RNA regulation, hypoexcitability and reduced
synaptic activity when compared with those derived from controls. Our gene
expression analyses revealed downregulation of several components of fragile X
mental retardation protein (FMRP) targets in TSC2-deficient hiPSC-PCs. We
detected decreased expression of FMRP, glutamate receptor δ2 (GRID2), and pre-
and post-synaptic markers such as synaptophysin and PSD95 in the TSC2-deficient
hiPSC-PCs. The mTOR inhibitor rapamycin rescued the deficits in
differentiation, synaptic dysfunction, and hypoexcitability of TSC2 mutant
hiPSC-PCs in vitro. Our findings suggest that these gene expression changes and
cellular abnormalities contribute to aberrant PC function during development in
TSC affected individuals.
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Increasing evidence has linked autism spectrum disorder
(ASD) with dysfunction of the brain's cerebellum, but the details have been
unclear. In a new study, researchers at Boston Children's Hospital used stem
cell technology to create cerebellar cells known as Purkinje cells from
patients with tuberous sclerosis complex (TSC), a genetic syndrome that often
includes ASD-like features. In the lab, the cells showed several
characteristics that may help explain how ASD develops at the molecular level…
TSC, a rare condition in which benign tumors grow in
multiple organs of the body, is associated with ASD in about half of all cases.
Previous brain autopsies have shown that patients with TSC, as well as patients
with ASD, have reduced numbers of Purkinje cells, the main type of neuron that
communicates out of the cerebellum. In a 2012 mouse study, Sahin and colleagues
knocked out a TSC gene (Tsc1) in Purkinje cells and found social deficits and
repetitive behaviors in the mice, together with abnormalities in the cells.
In the new paper, Sahin and colleagues took their
observations to humans, studying Purkinje cells derived from three patients
with TSC (two also had ASD symptoms, and all three also had epilepsy).
To make the cells, Sundberg first created induced
pluripotent stem cells from patients' blood cells or skin cells, then
differentiated these into neural progenitor cells and finally Purkinje cells.
The team then compared them with Purkinje cells derived from unaffected people
(parents or gender-matched controls) and with cells whose TSC mutation was
corrected using CRISPR-Cas9 gene editing.
"We saw changes," says Sahin. "The cells are
bigger and fire less than control cells - exactly what we see in the mouse
model."
Purkinje cells with the TSC genetic defect were harder to
differentiate from neural progenitor cells, suggesting that TSC may impair the
early development of cerebellar tissue. On examination, the patient-derived
Purkinje cells showed structural abnormalities in dendrites (the projections
neurons use to take in signals) and signs of impaired development of synapses
(junctions with other neurons)…
The study was the first to create human Purkinje cells using
TSC patients' own stem cells. In future studies, Sahin and colleagues hope to
generate larger numbers of patient-derived cells to investigate differences
between patients with TSC alone and those who also have ASD. Additionally, they
hope to use the Purkinje cell platform to study other ASD-related genetic
disorders, including Fragile X and SHANK3 mutation, and to test potential
drugs. Sundberg also plans to create other types of neurons for modeling ASD.
http://www.sciencenewsline.com/news/2018021706520005.html
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