Liora Altman-Sagan, BS, MS
Tufts University
A 3D Skin-Like Model to Study Progression of Organ Fibrosis and Screen Drugs for Targeted Therapies
Understanding mechanisms driving fibrosis has been limited by existing in vitro monolayer culture systems that do not mimic the cellular complexity and structure of fibrosis in organs. We have developed and studied 3D skin-like tissue models of fibrosis that incorporate patient-derived cells (macrophages, fibroblasts, T cells, keratinocytes). To increase the complexity of our tissue model, we incorporated Scleroderma (Ssc) patient-derived CD4+ T-cells and CD14+ macrophages to elucidate the role of cytokine-mediated, pro-inflammatory crosstalk between these cells and fibroblasts. When incorporating Ssc patient cells, typical 3D tissue architecture and differentiation was maintained. 3D tissues were analyzed by immunofluorescence staining, ELISA analysis, and single cell RNA sequencing. Immune cells produced cytokines (IL-6, IL-13) known to stimulate fibroblast activation and drive fibrosis in skin and other organs. Single cell analysis identified five distinct T cell subpopulations (CD8 T cells, proliferating CD4 T cells, activated CD4 T cells, naive CD4 T cells, Th17 CD4 T cells). This suggests that T cells and macrophages may communicate through cytokine cross talk to contribute to more closely model profibrotic phenotypes in 3D tissues. We are currently using this tissue model to test its utility as a drug screening platform for multiple fibrotic skin diseases mediated by JAK/STAT signaling as a step towards a more generalizable 3D tissue model of fibrosis in other organs.
Understanding mechanisms driving fibrosis has been limited by existing in vitro monolayer culture systems that do not mimic the cellular complexity and structure of fibrosis in organs. We have developed and studied 3D skin-like tissue models of fibrosis that incorporate patient-derived cells (macrophages, fibroblasts, T cells, keratinocytes). To increase the complexity of our tissue model, we incorporated Scleroderma (Ssc) patient-derived CD4+ T-cells and CD14+ macrophages to elucidate the role of cytokine-mediated, pro-inflammatory crosstalk between these cells and fibroblasts. When incorporating Ssc patient cells, typical 3D tissue architecture and differentiation was maintained. 3D tissues were analyzed by immunofluorescence staining, ELISA analysis, and single cell RNA sequencing. Immune cells produced cytokines (IL-6, IL-13) known to stimulate fibroblast activation and drive fibrosis in skin and other organs. Single cell analysis identified five distinct T cell subpopulations (CD8 T cells, proliferating CD4 T cells, activated CD4 T cells, naive CD4 T cells, Th17 CD4 T cells). This suggests that T cells and macrophages may communicate through cytokine cross talk to contribute to more closely model profibrotic phenotypes in 3D tissues. We are currently using this tissue model to test its utility as a drug screening platform for multiple fibrotic skin diseases mediated by JAK/STAT signaling as a step towards a more generalizable 3D tissue model of fibrosis in other organs.
