J Genet Med.  2015 Jun;12(1):25-30. 10.5734/JGM.2015.12.1.25.

Development of cell models for high-throughput screening system of Charcot-Marie-Tooth disease type 1

Affiliations
  • 1Department of Neurology, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea. bochoi77@hanmail.net, youngbinhong@gmail.com
  • 2Department of Biochemistry, Ewha Womans University School of Medicine, Seoul, Korea.
  • 3Stem Cell & Regenerative Medicine Center, Samsung Medical Center, Sungkyunkwan University School of Medicine, Seoul, Korea.

Abstract

PURPOSE
Charcot-Marie-Tooth disease (CMT) is a peripheral neuropathy mainly divided into CMT type 1 (CMT1) and CMT2 according to the phenotype and genotype. Although molecular pathologies for each genetic causative have not been revealed in CMT2, the correlation between cell death and accumulation of misfolded proteins in the endoplasmic reticulum (ER) of Schwann cells is well documented in CMT1. Establishment of in vitro models of ER stress-mediated Schwann cell death might be useful in developing drug-screening systems for the treatment of CMT1.
MATERIALS AND METHODS
To develop high-throughput screening (HTS) systems for CMT1, we generated cell models using transient expression of mutant proteins and chemical induction.
RESULTS
Overexpression of wild type and mutant peripheral myelin protein 22 (PMP22) induced ER stress. Similar results were obtained from mutant myelin protein zero (MPZ) proteins. Protein localization revealed that expressed mutant PMP22 and MPZ proteins accumulated in the ER of Schwann cells. Overexpression of wild type and L16P mutant PMP22 also reduced cell viability, implying protein accumulation-mediated ER stress causes cell death. To develop more stable screening systems, we mimicked the ER stress-mediated cell death in Schwann cells using ER stress inducing chemicals. Thapsigargin treatment caused cell death via ER stress in a dose dependent manner, which was measured by expression of ER stress markers.
CONCLUSION
We have developed genetically and chemically induced ER stress models using Schwann cells. Application of these models to HTS systems might facilitate the elucidation of molecular pathology and development of therapeutic options for CMT1.

Keyword

Charcot-Marie-Tooth disease; Endoplasmic reticulum stress; Human PMP22 protein; Myelin P0 protein; Thapsigargin
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