Remediating fibrosis for salivary gland regeneration

ABSTRACT Although extracellular matrix (ECM) remodeling is required for organ development and is a natural response in tissue repair, excessive ECM deposition in response to continual stress, injury, or aging is known as fibrosis, which impairs organ function and the potential for regenerative responses. Fibrosis is a significant cause of morbidity and mortality and contributes to up to 45% of all deaths in the US. 1-3 Fibrosis of the salivary gland occurs following radiation therapy for head and neck cancers and in patients with Sjögren’s Disease; however, the mechanisms driving reversibility of salivary gland fibrosis are not understood and are critical to understand for improved therapeutic development. While fibroblasts and TGFß signaling are known to be primary drivers of fibrosis, the inability to target cell-specific effects of TGFß limits the usefulness of TGFß therapeutics due to its pleiotropic effects on immune function. In prior work, we used single-cell RNA sequencing to identify a Pdgfra+, Pdgfrß+ co-positive subpopulation as the primary fibrogenic cell type that overproduces extracellular matrix in response to reversible salivary gland injury. As reduction or reversal of fibrosis could restore function, the objectives of this proposal are to define the contribution of TGFß modulation of PDGFRa+ stromal fibroblasts to matrisome composition and organization that drives reversible and irreversible salivary gland fibrosis and to test scaffold-mediated transplantation of mesenchymal stromal cells for improved ability to remediate a fibrotic response. We will address these outstanding questions in the field to better inform future therapeutic approaches to remediate or reverse fibrosis: 1) How does the TGFß-driven matrisome change in reversible vs irreversible fibrosis? 2) Can cell-specific manipulation of TGFß signaling in fibroblasts control fibrosis? 3) Can scaffold mediated delivery of mesenchymal stromal cells and their exosomes remediate fibrosis and promote gland regeneration? We will apply transcriptomic, proteomic, and quantitative histological approaches to reversible and irreversible models of salivary gland fibrosis and organoid models for human cells to define the cell-type specific responses of fibroblasts to create matrisome changes during fibrosis progression and recovery. These studies will significantly increase our understanding of the TGFß-dependent mechanisms that drive salivary gland fibrosis to inform improved regenerative medicine approaches and establish a foundation for testing novel therapeutics in organoids and mouse models to modulate TGFß and other effectors and matrisomal proteins in fibrotic diseases.

ABSTRACT Although extracellular matrix (ECM) remodeling is required for organ development and is a natural response in tissue repair, excessive ECM deposition in response to continual stress, injury, or aging is known as fibrosis, which impairs organ function and the potential for regenerative responses. Fibrosis is a significant cause of morbidity and mortality and contributes to up to 45% of all deaths in the US. 1-3 Fibrosis of the salivary gland occurs following radiation therapy for head and neck cancers and in patients with Sjögren’s Disease; however, the mechanisms driving reversibility of salivary gland fibrosis are not understood and are critical to understand for improved therapeutic development. While fibroblasts and TGFß signaling are known to be primary drivers of fibrosis, the inability to target cell-specific effects of TGFß limits the usefulness of TGFß therapeutics due to its pleiotropic effects on immune function. In prior work, we used single-cell RNA sequencing to identify a Pdgfra+, Pdgfrß+ co-positive subpopulation as the primary fibrogenic cell type that overproduces extracellular matrix in response to reversible salivary gland injury. As reduction or reversal of fibrosis could restore function, the objectives of this proposal are to define the contribution of TGFß modulation of PDGFRa+ stromal fibroblasts to matrisome composition and organization that drives reversible and irreversible salivary gland fibrosis and to test scaffold-mediated transplantation of mesenchymal stromal cells for improved ability to remediate a fibrotic response. We will address these outstanding questions in the field to better inform future therapeutic approaches to remediate or reverse fibrosis: 1) How does the TGFß-driven matrisome change in reversible vs irreversible fibrosis? 2) Can cell-specific manipulation of TGFß signaling in fibroblasts control fibrosis? 3) Can scaffold mediated delivery of mesenchymal stromal cells and their exosomes remediate fibrosis and promote gland regeneration? We will apply transcriptomic, proteomic, and quantitative histological approaches to reversible and irreversible models of salivary gland fibrosis and organoid models for human cells to define the cell-type specific responses of fibroblasts to create matrisome changes during fibrosis progression and recovery. These studies will significantly increase our understanding of the TGFß-dependent mechanisms that drive salivary gland fibrosis to inform improved regenerative medicine approaches and establish a foundation for testing novel therapeutics in organoids and mouse models to modulate TGFß and other effectors and matrisomal proteins in fibrotic diseases.