Supplementary MaterialsFigure S1: Image of floating hematopoietic cells derived from iPS cells Phase contrast image of floating hematopoietic cells derived from iPS-201B7 at day 21 (step 4 4). Characteristics of primary monocytes and macrophages. (A) Phase contrast image and (B) flow cytometric evaluation of macrophages produced from major monocytes. (C) The degrees of IL-6 and TNF- in supernatants of major monocyte tradition moderate 4 hours after LPS excitement. (D) The degrees of IL-1 had been assessed 4 hours after LPS excitement with/without yet another 30-minute ATP excitement.(PDF) pone.0059243.s003.pdf (98K) GUID:?F71EBA2C-F9EB-4AF5-988C-B1DA6427DFA6 Shape S4: Features and functional assays of dendritic cells produced from primary monocytes. (A) Movement cytometric evaluation of immature/mature DCs produced BAPTA/AM from major monocytes. (B) The degrees of IL-10 and TNF- in supernatants of tradition moderate with primary-DCs a day after LPS excitement. (C) The proliferation of allogeneic na?ve T cells (1105 cells per very well) co-cultured with 40 Gy-irradiated stimulator cells for 3 times was examined. The proliferation of na?ve T cells within the last 16 hours was measured by 3H-thymidine uptake.(PDF) pone.0059243.s004.pdf (176K) GUID:?ADCB416C-D20E-412C-B46F-09DFF72EA2Advertisement Figure S5: Features and functional assays of M1/M2 macrophages produced from major monocytes. (A) Flow cytometric analysis of M1/M2 macrophages derived from primary monocytes. (B) The levels of IL-12p70 and IL-10 in supernatants of culture medium with M1/M2 macrophages derived from primary monocytes 24 hours after LPS stimulation.(PDF) pone.0059243.s005.pdf (100K) GUID:?5E3D6F4D-634C-4957-A529-C481BAAA1158 Figure S6: Replication assays for 3 additional pluripotent stem cell lines. (A) Phase contrast image (left) and May-Giemsa staining BAPTA/AM (right) of mature DCs derived from iPSC clones. (B) Phase contrast image of macrophages derived from iPSC clones. Mouse monoclonal to Galectin3. Galectin 3 is one of the more extensively studied members of this family and is a 30 kDa protein. Due to a Cterminal carbohydrate binding site, Galectin 3 is capable of binding IgE and mammalian cell surfaces only when homodimerized or homooligomerized. Galectin 3 is normally distributed in epithelia of many organs, in various inflammatory cells, including macrophages, as well as dendritic cells and Kupffer cells. The expression of this lectin is upregulated during inflammation, cell proliferation, cell differentiation and through transactivation by viral proteins. (C) Flow cytometric analysis of immature/mature DCs and macrophages derived BAPTA/AM from iPSC clones.(PDF) pone.0059243.s006.pdf (268K) GUID:?7019D992-3C4F-46C2-AE50-CF8E4A66C5FC Table S1: Primers for RT-PCR. (PDF) pone.0059243.s007.pdf (9.7K) GUID:?A61751DF-95BE-4AF4-BDE0-400FFC7F551B Abstract Monocytic lineage cells (monocytes, macrophages and dendritic cells) play important roles in immune responses and are involved in various pathological conditions. The development of monocytic cells from human embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) is of particular interest because it provides an unlimited cell source for clinical application and basic research on disease pathology. Although the methods for monocytic cell differentiation from ESCs/iPSCs using embryonic body or feeder co-culture systems have already been established, these methods depend on the use of xenogeneic materials and, therefore, have a relatively poor-reproducibility. Here, we established a robust and highly-efficient method to differentiate functional monocytic cells from ESCs/iPSCs under serum- and feeder cell-free conditions. This method produced 1.31060.3106 floating monocytes from approximately 30 clusters of ESCs/iPSCs 5C6 times per course of differentiation. Such monocytes could be differentiated into functional macrophages and dendritic cells. This method should be useful for regenerative medicine, disease-specific iPSC studies and drug discovery. Introduction Monocytic lineage cells, such as monocytes, macrophages and dendritic cells (DCs), are central to immune responses and play key roles in various pathological conditions. C Monocytes are the myeloid progeny of hematopoietic stem/progenitor cells ; they are a type of mononuclear cell circulating in the bloodstream and act as gatekeepers in innate immunity. While they replenish macrophages and DCs, monocytes themselves respond to various inflammatory stimuli by migrating into inflamed tissues, phagocytosing pathological small particles and producing proinflammatory cytokines and chemokines. Therefore, monocytes not only contribute to host defense against pathogenic microorganisms, but are from the pathogenesis of chronic sterile irritation carefully.  Macrophages have a home in tissue and robustly phagocytose microorganisms and mobile debris. Among the essential BAPTA/AM hallmarks of monocytic lineage cells is certainly their useful plasticity. In response BAPTA/AM to cytokines and microbial items, macrophages polarize into distinct M1 and M2 cells functionally.  Classically turned on M1 macrophages are induced by interferon- (IFN), while activated M2 macrophages could be alternatively.