Modeling Dysregulation of Neurogenesis and Neurodevelopment in Tuberous Sclerosis Complex with Human Brain Organoids Open Access

Russell, Tiffany (Summer 2021)

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Tuberous sclerosis complex (TSC) is a genetic disorder in humans that manifests in the growth of benign tumors primarily in the brain as well as in other body systems. The neurological pathologies of TSC include cortical tubers, subependymal nodules (SENs), and subependymal giant cell astrocytomas (SEGAs), all of which are abnormal growths. These pathologies can result in symptoms such as epilepsy, autism spectrum disorder, attention deficit hyperactivity disorder, and other cognitive and learning disabilities in patients with TSC. TSC is caused by a mutation in one or both tumor suppressor genes TSC1 and TSC2; specifically, the mutation that results in an individual having TSC is a loss-of-function mutation in either or both genes. The TSC1 gene codes for a protein called hamartin (TSC1) and the TSC2 gene codes for a protein called tuberin (TSC2). Both hamartin and tuberin are responsible for inhibiting the mechanistic target of rapamycin complex 1 (mTORC1) protein complex, which is crucial in the regulation of cell growth through several mechanisms. When TSC1 and TSC2 are mutated and hamartin and tuberin are respectively not produced correctly, mTORC1 is upregulated, which allows for the growth of tumors observed in individuals with TSC. Brain organoids generated from iPSCs serve as small-scale models of the brain and are useful in studying morphological differences in neurogenesis and neurodevelopment between individuals with TSC and individuals without TSC. This study investigated the impact that TSC has on the developing brain in regards to neural progenitor cell proliferation and soma size using human brain organoids.  The hypothesis was that the loss of TSC1 and TSC2 function will result in abnormal hyperproliferation and soma overgrowth in neural progenitor cells (NPCs) in TSC cortical organoids. This hypothesis was supported by the data collected through fluorescence microscope analysis; it was observed that (1) a significantly higher proportion of NPCs were proliferating in the TSC organoids than in the control organoids and (2) NPC soma sizes were significantly larger in TSC organoids in comparison to control organoids. 

Table of Contents

Abstract: 1


Introduction and Background: 3

What Is Tuberous Sclerosis Complex?

Genetic Basis and Neurological Impacts

Using Induced Pluripotent Stem Cells to Investigate TSC



Methods: 9

Organoid Preparation

Immunostaining and Fluorescence Microscope Imaging

Cell Counting and Measurement

Statistical Analysis


Results: 12

TSC Cortical Organoids Demonstrated Significantly Higher Proportions of Proliferating Neural Progenitor Cells Than Control Cortical Organoids

TSC Cortical Organoids Demonstrated Significantly Larger Neural Progenitor Cell Soma Sizes Than Control Cortical Organoids


Figures: 14

Figure 1: Difference in Proportion of Proliferating Neural Progenitor Cells Out of All Neural Progenitor Cells Between Control and TSC Cortical Organoids

Figure 2: Difference in Size of Neural Progenitor Cells Somas Between Control and TSC Cortical Organoids

Discussion: 18

Hyperproliferation of Neural Progenitor Cells in TSC Cortical Organoids

Enlarged Neural Progenitor Cell Somas in TSC Cortical Organoids


Future Directions: 21


References: 22

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