A major focus of the laboratory involves studies of adult human hematopoietic stem cells. The cells are utilized primarily for studies of erythroid and iron biology using a variety of cellular and molecular techniques. Erythrocytes ultimately arise from hematopoietic stem cells in the peripheral blood of humans in response to erythropoietin. This process is called erythropoiesis. In many erythroid diseases including hemoglobinopathies, anemia and erythroleukemia, the process of erythropoiesis is abnormal or damaged by the underlying pathology. In order to better understand the molecular mechanisms responsible for these diseases, ex vivo model systems of erythropoiesis were developed using hematopoietic stem cells collected from the peripheral blood of humans. In this study, cells are harvested from peripheral blood and used for ex-vivo assays of erythropoiesis. In addition, collaborative efforts with the department of transfusion medicine are aimed toward improved methods for the collection of hematopoietic stem cells, and specialized methods are being developed for patients with erythroid diseases. Based upon interests in hemoglobin regulation, a genomics project was undertaken to determine differences between the fetal and adult reticulocyte transcriptome. As candidate genes are identified that may possess increased clinical relevance, the analyses are focused upon gene discovery and characterization. The laboratory has continued its efforts to update and expand the description of erythroid cell genetics in the context of post-genome biology. Previously, we reported the sequencing of human erythroblast libraries to generate an informatic database describing the erythroid transcriptome. Named Hembase (http://hembase.niddk.nih.gov/), the database is comprised of homology comparisons from our Expressed Sequence Tag (EST) collection with sequences contained within other publicly available databases. In addition, several disease-related search functions (Blood Groups, Cytoskeleton, Heme Synthesis, Hemoglobin, Hemolysis Related Enzymes) were integrated into Hembase to provide a genome-based organization of erythroid diseases. Based upon the recent discovery of functional RNA molecules that do not encode proteins, the database has been updated to include an undefined category of transcripts that do belong neither to clustered transcriptional foci within the genome nor protein-encoding regions. The database is presented on the worldwide web to disseminate information generated from the laboratory and to introduce the basic and clinical communities with a genome-organized view of erythroid disease. Among selected projects, the lab maintains a focus upon understanding ineffective erythropoiesis, iron regulation, and fetal hemoglobin production. Decreased expression of fetal hemoglobin around the time of birth represents the defining mechanism in the clinical manifestation of sickle-cell and beta-thalassemia syndromes. Therefore, prevention or reversal of that process represents a major clinical goal for the treatment of those diseases. An ongoing interest of the laboratory involves the study of development-related genes, signal transduction cascades, and growth manipulation for increasing levels of fetal hemoglobin in adult erythrocytes. These studies are useful for translational research aimed at malaria, hemoglobinopathies, anemia, leukemia, and iron pathologies. The projects have developed over several years with a primary focus upon the use of flow cytometry to identify, characterize and isolate the developmentally-staged cells. Flow cytometry has become sufficiently robust in recent years to provide analyses beyond the simple identification of the cells to include transcription, organelle development, and membrane specialization. Once purified, molecular biology applications have been developed for transcriptome analyses and quantitative analyses in single cells. Further, the approaches have been modified to include analyses of erythrocytes sampled directly patient volunteers. Finally, it has become possible to generate genetically manipulated erythrocytes in culture. As a result, the vast amount of information gained from the erythroid transcriptome is being exploited for translational research goals.

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de Vasconcellos, Jaira F; Lee, Y Terry; Byrnes, Colleen et al. (2016) HMGA2 Moderately Increases Fetal Hemoglobin Expression in Human Adult Erythroblasts. PLoS One 11:e0166928
Byrnes, Colleen; Lee, Y Terry; Meier, Emily R et al. (2016) Iron dose-dependent differentiation and enucleation of human erythroblasts in serum-free medium. J Tissue Eng Regen Med 10:E84-9
Lee, Y Terry; de Vasconcellos, Jaira F; Byrnes, Colleen et al. (2015) Erythroid-Specific Expression of LIN28A Is Sufficient for Robust Gamma-Globin Gene and Protein Expression in Adult Erythroblasts. PLoS One 10:e0144977
Glushakova, Svetlana; Balaban, Amanda; McQueen, Philip G et al. (2014) Hemoglobinopathic erythrocytes affect the intraerythrocytic multiplication of Plasmodium falciparum in vitro. J Infect Dis 210:1100-9
Meier, Emily Riehm; Wright, Elizabeth C; Miller, Jeffery L (2014) Reticulocytosis and anemia are associated with an increased risk of death and stroke in the newborn cohort of the Cooperative Study of Sickle Cell Disease. Am J Hematol 89:904-6
Lee, Y Terry; Kim, Ki Soon; Byrnes, Colleen et al. (2013) A synthetic model of human beta-thalassemia erythropoiesis using CD34+ cells from healthy adult donors. PLoS One 8:e68307
Miller, Jeffery L (2013) Iron deficiency anemia: a common and curable disease. Cold Spring Harb Perspect Med 3:
Meier, Emily Riehm; Byrnes, Colleen; Lee, Y Terry et al. (2013) Increased Reticulocytosis during Infancy Is Associated with Increased Hospitalizations in Sickle Cell Anemia Patients during the First Three Years of Life. PLoS One 8:e70794
Lee, Y Terry; de Vasconcellos, Jaira F; Yuan, Joan et al. (2013) LIN28B-mediated expression of fetal hemoglobin and production of fetal-like erythrocytes from adult human erythroblasts ex vivo. Blood 122:1034-41
Zafrani, Lara; Gerotziafas, Grigoris; Byrnes, Colleen et al. (2012) Calpastatin controls polymicrobial sepsis by limiting procoagulant microparticle release. Am J Respir Crit Care Med 185:744-55

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