Ngn2-Induced Differentiation of the NG108-15 Cell Line Enhances Motor Neuronal Differentiation and Neuromuscular Junction Formation.

in Biomolecules by Madeline Meli, Kristy Swiderski, Jinchao Gu, Ben Rollo, Ben Bartlett, Marissa K Caldow, Gordon S Lynch, Patrick Kwan, Huseyin Sumer, Brett Cromer

TLDR

  • The study showed that conditionally expressing the neurogenin-2 transcription factor in NG108-15 cells led to more rapid, efficient, and functional differentiation into cholinergic neurons.

Abstract

The neuronal progenitor NG108-15 neuroblastoma x glioma cell line proliferates indefinitely in vitro and is capable of directed differentiation into cholinergic neurons. The cell line is a robust model for investigating neuronal differentiation and function in vitro. The lineage-specific transcription factor-mediated differentiation of pluripotent stem cell lines (PSCs) leads to more rapid, efficient, and functional neurons. In this study, we tested the hypothesis that transcription factors could also drive the fate of an immortalised cell line. We first established a stable NG108-15 cell line, by piggyBac (pBac) transposition, that conditionally expresses neurogenin-2 (Ngn2), a common transcription factor for specifying neuronal fate. Following doxycycline-induction of Ngn2, we observed more rapid and efficient differentiation, and improved neurite outgrowth and viability compared with the WT cell line. Moreover, when co-cultured with C2C12 mouse myotubes, the modified NG108-15 cells resulted in significantly larger acetylcholine receptor (AChR) aggregates, suggesting enhanced neuromuscular junction (NMJ) formation. These findings describe a novel methodology for differentiating NG108-15 cells more efficiently, to enhance the usefulness of the cell line as a motor neuron model.

Overview

  • The study tested the hypothesis that transcription factors could drive the fate of an immortalized cell line, specifically the NG108-15 cell line, to more efficiently differentiate into cholinergic neurons.
  • The researchers established a stable NG108-15 cell line with conditionally expressed neurogenin-2 (Ngn2), a transcription factor that specifies neuronal fate, using piggyBac transposition.
  • The primary objective of the study was to enhance the usefulness of the NG108-15 cell line as a model for motor neuron differentiation and function by utilizing Ngn2-mediated differentiation.

Comparative Analysis & Findings

  • The study found that Ngn2-induced differentiation of NG108-15 cells led to more rapid and efficient differentiation compared to the wild-type cell line.
  • The modified cells exhibited improved neurite outgrowth and viability after differentiation induction.
  • Co-culture of the modified NG108-15 cells with C2C12 mouse myotubes resulted in significantly larger acetylcholine receptor (AChR) aggregates, suggesting enhanced neuromuscular junction (NMJ) formation.

Implications and Future Directions

  • The findings demonstrate a novel methodology for differentiating NG108-15 cells more efficiently, which can enhance the usefulness of the cell line as a model for motor neuron studies.
  • Future studies can focus on optimizing the Ngn2-mediated differentiation protocol to further enhance the quality and quantity of differentiated neurons.
  • The modified NG108-15 cell line may be useful for investigating motor neuron development, function, and disease modeling, paving the way for the discovery of new therapeutic strategies.
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