Neural Modeling Based on iPSCs Reveals New Target NPTX2 for Neurodegenerative Diseases

TAR DNA binding protein-43 (TDP-43) is a multifunctional DNA- and RNA-binding protein, which can be involved in a number of functions under normal physiological conditions, including the regulation of RNA metabolism, protein quality control system and mitochondrial quality control system. In recent years, TDP-43 has been implicated in neurodegenerative diseases such as amyotrophic lateral sclerosis (ALS), frontotemporal lobar degeneration (FTLD), and Alzheimer's disease (AD), but the pathogenic mechanisms behind these diseases are not clearly linked to TDP-43.

In response, a group of researchers from the University of Zurich and other institutions have used a neural cell culture model to reveal the neurodegenerative mechanisms involved in the aforementioned neurodegenerative diseases. In this article, Creative Biolabs describes the study, the results of which may pave the way for drug discovery for ALS and FTLD. For scientists, researchers and biopharmaceutical companies in this field, we can provide the following related services.

Neural Model iNet Derived from iPSC

In this study, the researchers created a novel neural cell culture model, iNet, derived from human iPSCs, which they first used to generate self-renewing human neural stem cell lines (iCoMoNSC). As the maturation of iCoMoNSC-derived neurons increased in culture, the differentiated iCoMoNSC formed a self-organizing multicellular system, which was named iNet.

• Based on scRNA-seq data, this neural model contained neurons and glial cells that were transcriptionally mature and similar to 3D brain organoids. Despite differences in the source cells and the absence of 3D organoid organization, neuronal and glial gene expression in the iNet is similar to that of cells in cortical organoids.
• The iNet model cultures last up to a year and are easily replicated, properties that allow them to be used to study the mechanisms involved in neuronal decline over time.
• The robustness of the aging iNet allows researchers to perform many experiments that would otherwise be impossible. And the model's flexibility makes it suitable for a wide range of experimental approaches.
• The iNet is easier to manipulate, clone and image than organoids, and the approach combines the organoid features of the brain with the flexibility of iPSC-derived monocultures.

Functional Evaluation of Neural Model iNet

The iPSC-derived iCoMoNSC formed a self-organized multicellular system. In the study, the researchers used the following methods for functional characterization.

These functional metrics suggest that the maturation of iCoMoNSC-derived neurons in culture increases as the functional neuronal network develops. The researchers used it to simulate TDP-43 pathology in cells from FTLD patients. It has been shown that only about 2% of cortical neurons in postmortem samples from FTLD patients exhibit signs of TDP-43-related pathology. The researchers transduced young iNet cultures with low-titer lentiviral vectors carrying labeled wild-type TDP-43. Analysis of iNet lysates showed that labeled TDP-43 progressively aggregated over time, similar to what occurs in TDP-43 protein-associated diseases.

Association Study of TDP-43 with NPTX2 in Neurodegenerative Diseases

NPTX2 accumulates consistently in neurons of patients with FTLD and ALS with TDP-43 pathology. So how does TDP-43 excess correlate with NPTX2 transcriptional upregulation? The researchers found that under physiological conditions, TDP-43 is able to bind directly to NPTX2 mRNA. In the brains of FTLD patients, this TDP-43-NPTX2 interaction was dramatically reduced.

Fig. 1 Immunofluorescence of TDP-43–HA in iNet neurons.¹

Therefore, the researchers directly linked TDP-43 dysregulation and NPTX2 accumulation, and found the following conclusions through the study.

• From the RNA sequencing data, the translation of NPTX2 is not controlled because TDP-43 binds in insoluble clusters.
• When NPTX2 was overexpressed in iNet, it exhibited neurotoxicity, and neurons induced to degeneration by TDP-43 could be partially rescued by correcting NPTX2 dysregulation.

Based on this, the team conducted an additional experiment using the iNet model to test whether NPTX2 could be used as a drug design target for FTLD and ALS. The team designed a shRNA targeting NPTX2 to facilitate its testing in an iNet model overexpressing the TDP-43 protein.

The results showed that reducing NPTX2 levels partially ameliorated the neurodegenerative lesions that had already occurred in iNet neurons. It is hypothesized that drugs used to target and reduce NPTX2 proteins may be able to halt neurodegenerative lesions in patients with ALS and FTLD.

Reference

1. Hruska-Plochan, Marian, et al. "A model of human neural networks reveals NPTX2 pathology in ALS and FTLD." Nature 626.8001 (2024): 1073-1083. Distributed under Open Access license CC BY 4.0. The image was modified by extracting and using only Part A-H of the original image.

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