ABSTRACT
[This corrects the article DOI: 10.1371/journal.pone.0237030.].
ABSTRACT
Curative therapeutic options for a number of immunological disorders remain to be established, and approaches for identifying drug candidates are relatively limited. Furthermore, phenotypic screening methods using induced pluripotent stem cell (iPSC)-derived immune cells or hematopoietic cells need improvement. In the present study, using immortalized monocytic cell lines derived from iPSCs, we developed a high-throughput screening (HTS) system to detect compounds that inhibit IL-1ß secretion and NLRP3 inflammasome activation from activated macrophages. The iPSCs were generated from a patient with neonatal onset multisystem inflammatory disease (NOMID) as a model of a constitutively activated NLRP3 inflammasome. HTS of 4,825 compounds including FDA-approved drugs and compounds with known bioactivity identified 7 compounds as predominantly IL-1ß inhibitors. Since these compounds are known inflammasome inhibitors or derivatives of, these results prove the validity of our HTS system, which can be a versatile platform for identifying drug candidates for immunological disorders associated with monocytic lineage cells.
Subject(s)
High-Throughput Screening Assays/methods , Inflammasomes/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/metabolism , Carrier Proteins/metabolism , Caspase 1/metabolism , Cell Line , Cells, Cultured , Cryopyrin-Associated Periodic Syndromes/immunology , Cryopyrin-Associated Periodic Syndromes/physiopathology , Humans , Induced Pluripotent Stem Cells/metabolism , Inflammasomes/physiology , Inflammation/metabolism , Interleukin-1beta , Macrophages/metabolism , Monocytes/metabolism , NLR Family, Pyrin Domain-Containing 3 Protein/physiologyABSTRACT
Various systems for differentiating hematopoietic cells from human pluripotent stem cells (PSCs) have been developed, although none have been fully optimized. In this report, we describe the development of a novel three-dimensional system for differentiating hematopoietic cells from PSCs using collagen sponges (CSs) reinforced with poly(ethylene terephthalate) fibers as a scaffold. PSCs seeded onto CSs were differentiated in a stepwise manner with appropriate cytokines under serum-free and feeder-free conditions. This process yielded several lineages of floating hematopoietic cells repeatedly for more than 1 month. On immunohistochemical staining, we detected CD34+ cells and CD45+ cells in the surface and cavities of the CS. Taking advantage of the portability of this system, we were able to culture multiple CSs together floating in medium, making it possible to harvest large numbers of hematopoietic cells repeatedly. Given these findings, we suggest that this novel three-dimensional culture system may be useful in the large-scale culture of PSC-derived hematopoietic cells.
