Deciphering the Molecular Mechanisms of Autonomic Nervous System Neuron Induction through Integrative Bioinformatics Analysis

Abstract

In vitro derivation of human neurons in the autonomic nervous system (ANS) is an important technology, given its regulatory roles in maintaining homeostasis in the human body. Although several induction protocols for autonomic lineages have been reported, the regulatory machinery remains largely undefined, primarily due to the absence of a comprehensive understanding of the molecular mechanism regulating human autonomic induction in vitro. In this study, our objective was to pinpoint key regulatory components using integrated bioinformatics analysis. A protein–protein interaction network construction for the proteins encoded by the differentially expressed genes from our RNA sequencing data, and conducting subsequent module analysis, we identified distinct gene clusters and hub genes involved in the induction of autonomic lineages. Moreover, we analyzed the impact of transcription factor (TF) activity on target gene expression, revealing enhanced autonomic TF activity that could lead to the induction of autonomic lineages. The accuracy of this bioinformatics analysis was corroborated by employing calcium imaging to observe specific responses to certain ANS agonists. This investigation offers novel insights into the regulatory machinery in the generation of neurons in the ANS, which would be valuable for further understanding and precise regulation of autonomic induction and differentiation.

Functional engineering of human iPSC‐derived parasympathetic neurons enhances responsiveness to gastrointestinal hormones

“…The correlation coefficients between receptor‐transduced iPSCs and wild‐type iPSCs (WT‐iPSCs) or MOCK‐iPSCs were > 0.98, indicating a high degree of similarity between the iPSC lines. However, the correlation coefficients between differentiated NCCs derived from WT‐iPSCs (day 13 of induction [ 21 , 23 ]) as a negative control and each type of iPSC line ranged 0.69–0.7, which were lower than the values obtained for different iPSC lines. Clustering analysis revealed that NCCs and iPSC lines clustered separately.…”

“…(OS‐762RC; Optime, Tokyo, Japan). Subsequently, iPSCs were differentiated into neural crest cells (NCCs), as previously reported [ 21 , 23 ]. Briefly, NCCs were induced from Ebs using knockout serum replacement (KSR) medium and N2 medium with graded addition of nerve growth factors and signaling factors.…”

“…Human‐derived cells inherently reflect our unique genetic and metabolic intricacies, ensuring a more accurate representation of human biology in experimental outcomes, potentially leading to more relevant therapeutic discoveries. Recent efforts have focused on utilizing in vitro approaches to generate human sensory [ 12 , 13 , 14 , 15 ], enteric [ 16 , 17 , 18 ], and autonomic nerves [ 19 , 20 , 21 , 22 , 23 ] from human embryonic stem cells (ESCs) and iPSCs. However, a clear limitation in these efforts is the absence of nerves responsive to GI hormones, which limits our ability to explore the intricate brain–gut axis and understand the etiology and potential interventions for metabolic or neurodegenerative diseases.…”

Computational study of the motor neuron protein KIF5A to identify nsSNPs, bioactive compounds, and its key regulators

“…We further analyze the PPIN of KIF5A using two local ranking methods degree (Pk) and maximum neighborhood component (MNC) and two global methods (bottleneck and closeness) with the help of the Cytoscape (v3.8.0) (Shannon et al, 2003) plugin CytoHubba (Chin et al, 2014). Using CytoHubba, we can identify the top 10 nodes which could play an important role in controlling the network and signal propagation (Chen et al, 2023;Takayama et al, 2023;Gu et al, 2023). We also performed the computational knockout experiments by eliminating the top significant node, resulting in network perturbations, a phenomenon known as the centrality-lethality rule (Jeong et al, 2001;Malik et al, 2019).…”