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  3. Protocol for a Wnt reporter assay to measure its activity in human neural stem cells derived from induced pluripotent stem cells

Protocol for a Wnt reporter assay to measure its activity in human neural stem cells derived from induced pluripotent stem cells

  • Curr Res Neurobiol. 2023 Jun 17:5:100095. doi: 10.1016/j.crneur.2023.100095.
Cristine Marie Yde Ohki 1 2,  Natalie Monet Walter 1,  Michelle Rickli 1,  José Maria Salazar Campos 1,  Anna Maria Werling 1,  Christian Döring 1,  Susanne Walitza 1 3 4,  Edna Grünblatt 1 3 4
Affiliations

Affiliations

  • 1 Department of Child and Adolescent Psychiatry and Psychotherapy, Translational Molecular Psychiatry, Psychiatric University Hospital Zurich, University of Zurich, Wagistrasse 12, 8952, Schlieren, Switzerland.
  • 2 Biomedicine PhD Program, University of Zurich, Winterthurerstrasse 11, 8057, Zurich, Switzerland.
  • 3 Neuroscience Center Zurich, University of Zurich and the ETH Zurich, Winterthurerstrasse 11, 8057, Zurich, Switzerland.
  • 4 Zurich Center for Integrative Human Physiology, University of Zurich, Winterthurerstrasse 11, 8057, Zurich, Switzerland.
PMID: 37426743 DOI: 10.1016/j.crneur.2023.100095
Abstract

The canonical Wnt signaling is an essential pathway that regulates cellular proliferation, maturation, and differentiation during neurodevelopment and maintenance of adult tissue homeostasis. This pathway has been implicated with the pathophysiology of neuropsychiatric disorders and was associated with cognitive processes, such as learning and memory. However, the molecular investigation of the Wnt signaling in functional human neural cell lines might be challenging since brain biopsies are not possible and animal models may not represent the polygenic profile of some neurological and neurodevelopmental disorders. In this context, using induced pluripotent stem cells (iPSCs) has become a powerful tool to model disorders that affect the Central Nervous System (CNS) in vitro, by maintaining patients' genetic backgrounds. In this method paper, we report the development of a virus-free Wnt reporter assay in neural stem cells (NSCs) derived from human iPSCs from two healthy individuals, by using a vector containing a reporter gene (luc2P) under the control of a TCF/LEF (T-cell factor/lymphoid enhancer factor) responsive element. Dose-response curve analysis from this luciferase-based method might be useful when testing the activity of the Wnt signaling pathway after agonists (e.g. Wnt3a) or antagonists (e.g. DKK1) administration, comparing activity between cases and controls in distinct disorders. Using such a reporter assay method may help to elucidate whether neurological or neurodevelopmental mental disorders show alterations in this pathway, and testing whether targeted treatment may reverse these. Therefore, our established assay aims to help researchers on the functional and molecular investigation of the Wnt pathway in patient-specific cell types comprising several neuropsychiatric disorders.

Keywords

Human stem cells; Induced pluripotent stem cells; Neural stem cells; Neuropsychiatry; Reporter assay; Wnt signaling.

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