Technologies to study and control patterning and differentiation in multicellular development, a case study for multilayered arterial wall of controlled thickness

How do cells in mammalian organisms integrate various inputs to generate elaborate spatial arrangement of differentiated cell types as seen in tissues and organs? Naturally evolved genetic networks based on ligands and receptors are essential for embryonic development and maintaining adult tissues. Alterations in genes, effector proteins, and cellular environments can disrupt normal development, leading to congenital disorders and adult diseases such as cancer and degeneration. Recently, synthetic genetic networks based on synthetic receptors have been developed and used in research settings to perturb and reconstruct complex multicellular networks (e.g., synNotch receptors that we developed). In our lab, we have two main goals: to develop new technologies to manipulate cell differentiation in space and time with synthetic signaling systems, and to apply these technologies to reconstruct specific examples of complex arrangement of cells, for example here a multilayered arterial vessel. In the Research Strategy section of this proposal, we outline two research Tracks, and their respective goals: 1. Development of New Technologies for 3D Differentiation Control: We will develop and integrate two technologies: (i) control of differentiation in three dimensions (3D) layers of defined thickness around spherical or thread-like scaffolds to model organs with radial symmetry around a nucleus like liver, branched epithelia, skin, and blood vessels among others; (ii) autonomously patterning genetic circuits of the reaction-diffusion family to obtain 2D and 3D gene expression domains like spots, stripes and labyrinth, known as Turing-like patterns. 2. Study and control of cell-cell communication among differentiating endothelial and vascular support cells: we will utilize patterned Syn-Notch signaling to build a perfusable vasculature comprised of an endothelial intima and a smooth muscle media layer of controlled thickness. We will generate and perturb these constructs where human induced pluripotent stem cells are differentiating to endothelial cells and vascular smooth muscle cells in geometrically controlled fashion in 2D and in 3D. We will identify and use the signals and communication network that support construction of perfusable functional tissues. These studies aim to enhance synthetic and developmental biology by deepening our understanding of cell signaling mechanisms in multicellular communities. Ultimately, these insights could advance cell-based therapies and improve disease treatment strategies. Project Number: 1R35GM162013-01 | Fiscal Year: 2026 | NIH Institute/Center: National Institute of General Medical Sciences (NIGMS) | Principal Investigator: Leonardo Morsut | Institution: UNIVERSITY OF SOUTHERN CALIFORNIA, Los Angeles, CA | Award Amount: $656,500 | Activity Code: R35 | Study Section: Special Emphasis Panel[ZRG1 MCST-Q (55)] View on NIH RePORTER: https://reporter.nih.gov/project-details/11263492