UCSF

The gastrointestinal tract is a complex organ system whose function depends on coordinated activity of epithelial, mesenchymal, and neural compartments. During embryonic development, these compartments undergo orchestrated morphogenetic events to establish the architectural and functional features of the mature intestine. This thesis integrates developmental biology, stem cell biology, and human genetics to elucidate mechanisms governing intestinal morphogenesis and enteric nervous system (ENS) development, and to identify genetic and cellular pathways underlying disorders of gut-brain interaction (DGBIs).Chapter 1 provides an introduction to intestinal villus morphogenesis, ENS development and patterning, motor neuron specification, and the genetic susceptibility to DGBIs, establishing the context for the experimental work presented in subsequent chapters.Chapter 2 investigates the role of extracellular matrix (ECM) adhesion and remodeling in intestinal villus morphogenesis. Using mouse explant cultures, pharmacological inhibition of matrix metalloproteinases, and single-cell RNA sequencing, this chapter demonstrates that integrin-mediated cell-ECM interactions and MMP-mediated matrix degradation are essential for PDGFRα+ mesenchymal condensation and epithelial folding. These findings establish ECM remodeling as a central mechanism underlying villus formation.Chapter 3 investigates the mechanisms of ENS patterning within the regionally specialized gut environment. Using multiplexed single-cell RNA sequencing of the developing mouse small intestine (E13.5–E18.5), this chapter demonstrates that while the epithelium and mesenchyme maintain strong anterior–posterior (A-P) graded gene expression, the ENS lacks overt intrinsic regionalization. Instead, the ENS follows a predominantly temporal maturation trajectory defined by distinct glial and neuronal differentiation states. Integrative ligand-receptor analysis identifies the PTN/MDK-PTPRZ1 signaling axis as a critical, spatially graded microenvironmental input derived from the mesenchyme. Functional perturbation of this pathway in human pluripotent stem cell-derived ENS cultures confirms that PTPRZ1 signaling fine-tunes progenitor proliferation, neurogenesis, and neurotransmitter specification. Collectively, these findings support a model in which ENS maturation relies on a uniform core program that is secondarily refined by extrinsic, region-specific mesenchymal cues.Chapter 4 focuses on enteric motor neuron specification and functional characterization, particularly nitrergic inhibitory neurons. Using hPSC-derived enteric ganglioid models and high-throughput screening, this chapter identifies signaling pathways that promote nitrergic neuron differentiation and drugs that modulate nitrergic neuron activity.Chapter 5 leverages genome-wide association studies using the All of Us Research Program database to identify genetic variants and candidate genes associated with DGBIs, including irritable bowel syndrome, functional diarrhea, functional dyspepsia, and functional fecal incontinence. The chapter reveals enrichment of genes involved in neuronal excitability, synaptic transmission, and ion channel regulation, supporting the hypothesis that genetic variants affecting ENS development or function contribute to DGBI pathophysiology.Collectively, these studies demonstrate the power of integrating mouse developmental genetics, human pluripotent stem cell and ex-vivo models, transcriptomics, and human genetics to address fundamental questions in gut biology and disease.