A proof-of-concept screen for identification of novel natural product modulators involved in mouse Treg/Th17 differentiation by using high-throughput Flow Cytometry (124.2)

Abstract The recent discovery of two novel subsets of CD4+ T lymphocytes (Th17 and Treg) has led to a paradigm shift in the understanding of how autoimmune responses are mediated and regulated. Interestingly, both Th17 and Treg cells can develop from the same naïve CD4+ T cell precursors and in some circumstances they can convert to each other. Several recent publications highlight the role of the aryl hydrocarbon receptor (Ahr) in regulating Treg and Th17 cell differentiation and possible application to human diseases. Depending on the ligands, Ahr activation can control naïve CD4+ T cell differentiation towards either a Treg (TCDD) or a Th17 (FICZ) phenotype. Identifying novel agents that promote or inhibit differentiation of naïve CD4+ T cells to Treg or Th17 could be beneficial in the area of Oncology and autoimmune diseases. Here we describe an assay developed for identification of new modulators based on T cell differentiation by using high-throughput flow cytometry. With TGF-β or TGF-β/IL-6 stimulation isolated mouse naïve CD4 cells can be differentiated efficiently into Treg or Th17 lineages. Treg or Th17-specific expression was detected by intracellular staining of FoxP3 or IL-17, respectively. A proof-of-concept screening against 1600 pure natural products (NPs) was performed and seventeen Th17 specific hits were identified