Biology subjects

Heideveld, E.

Publications and source records attributed to Heideveld, E..

3 recordsLinked to original sources

bioRxiv2020

Abundant circular RNA expression in terminal myeloid differentiation does not correlate with frequent circRNA translation, or with circRNA-mediated translation regulation

Each day, about 1012 erythrocytes and platelets are released into the blood stream. This substantial output from hematopoietic stem cells is tightly regulated by transcriptional and epigenetic factors. Whether and how circular RNAs (circRNAs) contribute to the differentiation and/or identity of hematopoietic cells is to date not known. We recently reported that erythrocytes and platelets contain the highest levels and numbers of circRNAs amongst hematopoietic cells. Here, we provide the first detailed analysis of circRNA expression during erythroid and megakaryoid differentiation. CircRNA expression not only significantly increased upon enucleation, but also had limited overlap between progenitor cells and mature cells, suggesting that circRNA expression stems from regulated processes rather than resulting from mere accumulation. To study circRNA function in hematopoiesis, we first compared the expression levels of circRNAs with the translation efficiency of their mRNA-counterpart. We found that only 1 out of 2531 (0.04%) circRNAs associated with mRNA-translation regulation. Furthermore, irrespective of 1000s of identified putative open reading frames, deep ribosome-footprinting sequencing and mass spectrometry analysis provided little evidence for translation of endogenously expressed circRNAs. In conclusion, circRNAs alter their expression profile during terminal hematopoietic differentiation, yet their contribution to regulate cellular processes remains enigmatic.

molecular biology↗

bioRxiv2020

Mouse Classical and Non-Classical Monocytes Express Comparable Levels of Chemokine Receptor CX3CR1

Human and mouse monocytes are divided into two subpopulations, classical (C-Mo) and non-classical (NC-Mo) monocytes. CC-chemokine receptor 2 (CCR2) and CX3C-chemokine receptor 1 (CX3CR1) are common features of monocyte subsets between humans and mice, i.e., C-Mo and NC-Mo are characterized as CCR2highCX3CR1low and CCR2lowCX3CR1high. Since many studies utilize mouse models to investigate roles of monocytes in human diseases, it is important to understand the similarities and differences between human and mouse monocytic subsets. In this study, we show that the expression of Cx3cr1 mRNA and CX3CR1 cell surface protein are different between circulating monocytic subsets in human but not in mice. We analyzed monocyte subsets in the blood using wild type C57BL/6 and Cx3cr1-GFP knock-in (Cx3cr1GFP/+) reporter mice. We observed higher Cx3cr1 promoter activity indicated by GFP expression in NC-Mo compared to C-Mo. However, there were no differences between the subsets in CX3CR1 mRNA nor surface protein expression determined by anti-CX3CR1 antibody or binding of fluorophore-conjugated ligand. However in the bone marrow of Cx3cr1GFP/+ mice, CX3CR1 expression was higher in NC-Mo compared to C-Mo, suggesting that mouse NC-Mo express higher level of CX3CR1 than C-Mo in the bone but this difference disappears in the blood. In contrast, human NC-Mo differentially expressed CX3CR1 compared to C-Mo in both blood and bone marrow. Given these findings, the discrepancy between promoter activity and protein levels should be considered when the roles of CX3CR1 are investigated in mouse models of human diseases. Summary sentenceIn this study, we show that the expression of Cx3cr1 mRNA is different between circulating monocytic subsets in human but not in mice.

immunology↗

bioRxiv2019

Large-scale in-vitro production of red blood cells from human peripheral blood mononuclear cells

Transfusion of donor-derived red blood cells is the most common form of cellular therapy. Donor availability and the potential risk of alloimmunization and other transfusion-related complications may, however, limit the availability of transfusion units especially for chronically transfused patients. In-vitro cultured, customizable red blood cells would negate these concerns and introduce precision medicine. Large-scale, cost effective production depends on optimization of culture conditions. We developed a defined medium and adapted our protocols to GMP culture requirements, which reproducibly provided pure erythroid cultures from peripheral blood mononuclear cells without prior CD34+ isolation, and a 3x107-fold increase in erythroblasts in 25 days. Expanded erythroblast cultures could be differentiated to CD71dimCD235a+CD44+CD117-DRAQ5- red blood cells in 12 days. More than 90% of the cells enucleated and expressed adult hemoglobin as well as the correct blood group antigens. Deformability and oxygen binding capacity of cultured red blood cells was comparable to in-vivo reticulocytes. Daily RNA sampling during differentiation followed by RNA-seq provided a high-resolution map/resource of changes occurring during terminal erythropoiesis. The culture process was compatible with upscaling using a G-Rex bioreactor with a capacity of 1L per reactor, allowing transition towards clinical studies and small-scale applications.

cell biology↗