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Proceedings Paper

Third harmonic generation imaging of intact human cerebral organoids to assess key components of early neurogenesis in Rett Syndrome (Conference Presentation)
Author(s): Murat Yildirim; Danielle Feldman; Tianyu Wang; Dimitre G. Ouzounov; Stephanie Chou; Justin Swaney; Kwanghun Chung; Chris Xu; Peter T. C. So; Mriganka Sur
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Paper Abstract

Rett Syndrome (RTT) is a pervasive, X-linked neurodevelopmental disorder that predominantly affects girls. It is mostly caused by a sporadic mutation in the gene encoding methyl CpG-binding protein 2 (MeCP2).The clinical features of RTT are most commonly reported to emerge between the ages of 6-18 months and implicating RTT as a disorder of postnatal development. However, a variety of recent evidence from our lab and others demonstrates that RTT phenotypes are present at the earliest stages of brain development including neurogenesis, migration, and patterning in addition to stages of synaptic and circuit development and plasticity. We have used RTT patient-derived induced pluripotent stem cells to generate 3D human cerebral organoids that can serve as a model for human neurogenesis in vitro. We aim to expand on our existing findings in order to determine aberrancies at individual stages of neurogenesis by performing structural and immunocytochemical staining in isogenic control and MeCP2-deficient organoids. In addition, we aim to use Third Harmonic Generation (THG) microscopy as a label-free, nondestructive 3D tissue visualization method in order to gain a complete understanding of the structural complexity that underlies human neurogenesis. As a proof of concept, we have performed THG imaging in healthy intact human cerebral organoids cleared with SWITCH. We acquired an intrinsic THG signal with the following laser configurations: 400 kHz repetition rate, 65 fs pulse width laser at 1350 nm wavelength. In these THG images, nuclei are clearly delineated and cross sections demonstrate the depth penetration capacity (< 1mm) that extends throughout the organoid. Imaging control and MeCP2-deficient human cerebral organoids in 2D sections reveals structural and protein expression-based alterations that we expect will be clearly elucidated via both THG and three-photon fluorescence microscopy.

Paper Details

Date Published: 24 April 2017
PDF: 1 pages
Proc. SPIE 10069, Multiphoton Microscopy in the Biomedical Sciences XVII, 100690A (24 April 2017); doi: 10.1117/12.2256182
Show Author Affiliations
Murat Yildirim, Massachusetts Institute of Technology (United States)
Danielle Feldman, Massachusetts Institute of Technology (United States)
Tianyu Wang, Cornell Univ. (United States)
Dimitre G. Ouzounov, Cornell Univ. (United States)
Stephanie Chou, Massachusetts Institute of Technology (United States)
Justin Swaney, Massachusetts Institute of Technology (United States)
Kwanghun Chung, Massachusetts Institute of Technology (United States)
Chris Xu, Cornell Univ. (United States)
Peter T. C. So, Massachusetts Institute of Technology (United States)
Mriganka Sur, Massachusetts Institute of Technology (United States)


Published in SPIE Proceedings Vol. 10069:
Multiphoton Microscopy in the Biomedical Sciences XVII
Ammasi Periasamy; Peter T. C. So; Karsten König; Xiaoliang S. Xie, Editor(s)

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