Background Blood transfusion is an essential part of medicine. However, many countries have been facing a national blood crisis. To address this ongoing blood shortage issue, there have been efforts to generate red blood cells (RBCs) in vitro, especially from human-induced pluripotent stem cells (hiPSCs). However, the best source of hiPSCs for this purpose is yet to be determined. Methods In this study, hiPSCs were established from three different hematopoietic stem cell sources—peripheral blood (PB), cord blood (CB) and bone marrow (BM) aspirates (n = 3 for each source)—using episomal reprogramming vectors and differentiated into functional RBCs. Various time-course studies including immunofluorescence assay, quantitative real-time PCR, flow cytometry, karyotyping, morphological analysis, oxygen binding capacity analysis, and RNA sequencing were performed to examine and compare the characteristics of hiPSCs and hiPSC-differentiated erythroid cells. Results hiPSC lines were established from each of the three sources and were found to be pluripotent and have comparable characteristics. All hiPSCs differentiated into erythroid cells, but there were discrepancies in differentiation and maturation efficiencies: CB-derived hiPSCs matured into erythroid cells the fastest while PB-derived hiPSCs required a longer time for maturation but showed the highest degree of reproducibility. BM-derived hiPSCs gave rise to diverse types of cells and exhibited poor differentiation efficiency. Nonetheless, erythroid cells differentiated from all hiPSC lines mainly expressed fetal and/or embryonic hemoglobin, indicating that primitive erythropoiesis occurred. Their oxygen equilibrium curves were all left-shifted. Conclusions Collectively, both PB- and CB-derived hiPSCs were favorably reliable sources for the clinical production of RBCs in vitro, despite several challenges that need to be overcome. However, owing to the limited availability and the large amount of CB required to produce hiPSCs, and the results of this study, the advantages of using PB-derived hiPSCs for RBC production in vitro may outweigh those of using CB-derived hiPSCs. We believe that our findings will facilitate the selection of optimal hiPSC lines for RBC production in vitro in the near future.
Inborn errors of immunity (IEI) include a variety of heterogeneous genetic disorders in which defects in the immune system lead to an increased susceptibility to infections and other complications. Accurate, prompt diagnosis of IEI is crucial for treatment plan and prognostication. In this study, clinical utility of clinical exome sequencing (CES) for diagnosis of IEI was evaluated. For 37 Korean patients with suspected symptoms, signs, or laboratory abnormalities associated with IEI, CES that covers 4,894 genes including genes related to IEI was performed. Their clinical diagnosis, clinical characteristics, family history of infection, and laboratory results, as well as detected variants, were reviewed. With CES, genetic diagnosis of IEI was made in 15 out of 37 patients (40.5%). Seventeen pathogenic variants were detected from IEI-related genes, BTK, UNC13D, STAT3, IL2RG, IL10RA, NRAS, SH2D1A, GATA2, TET2, PRF1, and UBA1, of which four variants were previously unreported. Among them, somatic causative variants were identified from GATA2, TET2, and UBA1. In addition, we identified two patients incidentally diagnosed IEI by CES, which was performed to diagnose other diseases of patients with unrecognized IEI. Taken together, these results demonstrate the utility of CES for the diagnosis of IEI, which contributes to accurate diagnosis and proper treatments.
Background: Accurate blood typing is essential for blood transfusions, and requires the constant evaluation and maintenance of ABO and D blood grouping reagents. In the present study, we developed cryopreserved red blood cell (RBC) panels and evaluated their feasibility as a standard reference material to verify the quality of ABO and D blood grouping reagents in Korea. Methods: RBC units obtained from healthy donors were cryopreserved using a high-glycerol method. A total of 400 sets of RBC panels were prepared, composed of blood group A (N=5), B (N=5), O (N=10), AB (N=4), Rh D-positive (N=4), Rh D-negative (N=5), and weak-D (N=1), and 200 sets of RBC subgroup panels composed of A2, A2B, A2B3, A1B3, and B3, and A2, A2B, A2B3, A1B3, and A3B (N=1, each). Quality assessment of the cryopreserved RBC panels before and after cryopreservation was performed by measuring their sensitivity, specificity, avidity, and potency titers. Results: Our cryopreserved ABO and D RBC panels had a sensitivity and specificity of 100% to existing monoclonal blood grouping reagents, regardless of blood type and cryopreservation time. There were no significant differences in the avidity time and potency titers of the cryopreserved RBCs before and after 6 or 12 months of cryopreservation. Conclusions: The quality parameters measured here suggest that our newly developed cryopreserved RBC panels were reliable for use as a standard reference material for the performance evaluation of anti-A, -B, and -D blood grouping reagents.
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