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

2020 ◽  
Author(s):  
Benoit P. Nicolet ◽  
Sjoert B.G. Jansen ◽  
Esther Heideveld ◽  
Willem H. Ouwehand ◽  
Emile van den Akker ◽  
...  
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Hematopoietic Cells ◽  
Blood Stream ◽  
Mass Spectrometry Analysis ◽  
Translation Regulation ◽  
Circular Rnas ◽  
Hematopoietic Stem ◽  
Erythroid Precursor ◽  
Spectrometry Analysis

ABSTRACTEach day, about 1012 erythrocytes and platelets are released into the blood stream. This substantial output from hematopoietic stem cells is tightly regulated by transcription factors and epigenetic modifications. Whether and how non-coding RNAs such as circular RNAs (circRNAs) contribute to the differentiation and/or identity of hematopoietic cells is to date not well understood. We recently reported a circRNA expression map of hematopoietic cells, revealing that erythrocytes and platelets contain the highest levels and numbers of circRNAs. Here, we provide the first detailed and comprehensive analysis of circRNA expression during red blood cell and megakaryocyte differentiation. CircRNA expression significantly increased during erythroid precursor differentiation into red blood cells and in differentiating megakaryocytes, in particular upon enucleation. To determine if circRNAs can modulate hematopoietic differentiation, we compared the expression levels of circRNAs and mRNAs with that of ribosomal foot printing read. This multi-omics approach revealed that only 20 out of 748 (2.6%) circRNAs associated with translation regulation of their mRNA counterparts. Furthermore, irrespective of the thousands of identified putative open reading frames in circRNAs, deep ribosome-footprinting sequencing and mass spectrometry analysis provided little evidence for translation of endogenously expressed circRNAs in erythroblasts, megakaryocytes and platelets. In conclusion, circRNAs in platelets and red blood cells are highly abundant and alter their expression profile during differentiation, yet their contribution to regulate cellular processes remains enigmatic.

scholarly journals Circular RNAs exhibit limited evidence for translation, or translation regulation of the mRNA-counterpart in terminal hematopoiesis

RNA ◽  
2021 ◽  
pp. rna.078754.121
Author(s):  
Benoit P Nicolet ◽  
Sjoert BG Jansen ◽  
Esther Heideveld ◽  
Willem H Ouwehand ◽  
Emile van den Akker ◽  
...  
Keyword(s):  
Hematopoietic Cells ◽  
Mrna Translation ◽  
Blood Stream ◽  
Open Reading Frames ◽  
Mass Spectrometry Analysis ◽  
Translation Regulation ◽  
Circular Rnas ◽  
Translation Efficiency ◽  
Hematopoietic Stem ◽  
Spectrometry Analysis

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.

scholarly journals Levels of carnitine and acylcarnitines in reconstituted red blood cell samples washed with different concentrations of saline solutions

2011 ◽  
pp. 344-348
Author(s):  
José Henry Osorio ◽  
Morteza Pourfarzam
Keyword(s):  
Red Blood Cells ◽  
Blood Cells ◽  
Saline Solution ◽  
Mass Spectrometry Analysis ◽  
Blood Samples ◽  
Spectrometry Analysis ◽  
Tandem Mass Spectrometry Analysis ◽  
Human Blood Samples ◽  
Saline Solutions ◽  
Long Chain

Objective: To evaluate the percentage of carnitine and acylcarnitines remaining in red blood cells after washing them with different concentrations of saline solution. Materials and methods: Human blood samples were centrifuged and the blood cells were washed with different saline solutions. The final pellet was resuspended in PBS for card preparation and tandem mass spectrometry analysis. Results: It was found that carnitine, as well as short-chain, medium-chain, and long-chain acylcarnitines remain in red blood cells at average percentages of 19.3; 34; 34; and 32%, respectively. Significant differences were found for carnitine and acylcarnitine levels in blood washed with an isotonic solution compared to their levels using several hypotonic solutions (p<0.05). Conclusion: Because carnitine and acylcarnitines remained associated with the blood cells, we recommend using whole blood to measure these metabolites.

scholarly journals Mesenchymal stromal cells can be applied to red blood cells storage as a kind of cellular additive

2017 ◽  
Vol 37 (5) ◽  
Author(s):  
Yaozhen Chen ◽  
Jing Zhang ◽  
Shunli Gu ◽  
Dandan Yin ◽  
Qunxing An ◽  
...  
Keyword(s):  
Red Blood Cells ◽  
Mesenchymal Stromal Cells ◽  
Stromal Cells ◽  
Blood Cells ◽  
Storage Duration ◽  
Hematopoietic Stem ◽  
Diminished Capacity ◽  
Blood Banks ◽  
Citrate Phosphate

During storage in blood banks, red blood cells (RBCs) undergo the mechanical and metabolic damage, which may lead to the diminished capacity to deliver oxygen. At high altitude regions, the above-mentioned damage may get worse. Thus, more attention should be paid to preserve RBCs when these components need transfer from plain to plateau regions. Recently, we found that mesenchymal stromal cells (MSCs) could rescue from anemia, and MSCs have been demonstrated in hematopoietic stem cells (HSCs) transplantation to reconstitute hematopoiesis in vivo by us. Considering the functions and advantages of MSCs mentioned above, we are trying to find out whether they are helpful to RBCs in storage duration at high altitudes. In the present study, we first found that mice MSCs could be preserved in citrate phosphate dextrose adenine-1 (CPDA-1) at 4 ± 2°C for 14 days, and still maintained great viability, even at plateau region. Thus, we attempted to use MSCs as an available supplement to decrease RBCs lesion during storage. We found that MSCs were helpful to support RBCs to maintain biochemical parameters and kept RBCs function well on relieving anemia in an acute hemolytic murine model. Therefore, our investigation developed a method to get a better storage of RBCs through adding MSCs, which may be applied in RBCs storage as a kind of cellular additive into preservation solution.

scholarly journals Merocyanine 540-sensitized photoinactivation of human erythrocytes parasitized by Plasmodium falciparum

Blood ◽  
1992 ◽  
Vol 80 (1) ◽  
pp. 21-24 ◽  
Author(s):  
OM Smith ◽  
SA Dolan ◽  
JA Dvorak ◽  
TE Wellems ◽  
F Sieber
Keyword(s):  
Stem Cells ◽  
Plasmodium Falciparum ◽  
Hematopoietic Stem Cells ◽  
Red Blood Cells ◽  
Human Blood ◽  
Blood Cells ◽  
Potential Usefulness ◽  
Hematopoietic Stem ◽  
Infected Blood ◽  
Merocyanine 540

The purpose of this study was to evaluate the photosensitizing dye merocyanine 540 (MC540) as a means for extracorporeal purging of Plasmodium falciparum-infected erythrocytes from human blood. Parasitized red blood cells bound more dye than nonparasitized cells, and exposure to MC540 and light under conditions that are relatively well tolerated by normal erythrocytes and normal pluripotent hematopoietic stem cells reduced the concentration of parasitized cells by as much as 1,000-fold. Cells parasitized by the chloroquine- sensitive HB3 clone and the chloroquine-resistant Dd2 clone of P falciparum were equally susceptible to MC540-sensitized photolysis. These data suggest the potential usefulness of MC540 in the purging of P falciparum-infected blood.

scholarly journals Peripheral Red Blood Cell Split Chimerism as a Consequence of Intramedullary Selective Apoptosis of Recipient Red Blood Cells in a Case of Sickle Cell Disease

2014 ◽  
Vol 6 (1) ◽  
pp. e2014066 ◽  
Author(s):  
Marco Marziali ◽  
Antonella Isgrò ◽  
Pietro Sodani ◽  
Javid Gaziev ◽  
Daniela Fraboni ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Red Blood Cells ◽  
Sickle Cell ◽  
Sickle Cell Anemia ◽  
Blood Cells ◽  
Marrow Transplant ◽  
Mixed Chimerism ◽  
Radical Cure ◽  
Hematopoietic Stem ◽  
Hematopoietic Chimerism

Allogeneic cellular gene therapy through hematopoietic stem cell transplantation is the only radical cure for congenital hemoglobinopathies like thalassemia and sickle cell anemia. Persistent mixed hematopoietic chimerism (PMC) has been described in thalassemia and sickle cell anemia. Here, we describe the clinical course of a 6-year-old girl who had received bone marrow transplant for sickle cell anemia. After the transplant, the patient showed 36% donor hematopoietic stem cells in the bone marrow, whereas in the peripheral blood there was evidence of 80%  circulating donor red blood cells (RBC). The analysis of apoptosis at the Bone Marrow  level suggests that Fas might contribute to the cell death of host erythroid precursors. The increase in NK cells and the regulatory T cell population observed in this patient suggests that these cells might contribute to the condition of mixed chimerism.

Differential Requirements for Survivin in Hematopoietic Cell Development.

Blood ◽  
2004 ◽  
Vol 104 (11) ◽  
pp. 1257-1257
Author(s):  
Yanfei Xu ◽  
Sandeep Gurbuxani ◽  
Ganesan Keerthivasan ◽  
Amittha Wickrema ◽  
John D. Crispino
Keyword(s):  
Cell Cycle ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Caspase 3 ◽  
Terminal Differentiation ◽  
Dependent Manner ◽  
Erythroid Cells ◽  
Hematopoietic Stem ◽  
Stage Of Development ◽  
Primary Mouse

Abstract The development of the complete repertoire of blood cells from a common progenitor, the hematopoietic stem cell, is a tightly controlled process that is regulated, in part, by the activity of lineage specific transcription factors. Despite our knowledge of these factors, the mechanisms that regulate the formation and growth of distinct, but closely related lineages, such as erythroid cells and megakaryocytes, remain largely uncharacterized. Here we show that Survivin, a member of the inhibitor of apoptosis (IAP) family that also plays an essential role in cytokinesis, is differentially expressed during erythroid versus megakaryocyte development. Erythroid cells express Survivin throughout their maturation, up to the terminal stage of differentiation (orthochromatic), even after the cells exit the cell cycle. This is surprising because Survivin is generally expressed in a cell cycle dependent manner and not thought to be expressed in terminally differentiated cells. In contrast, purified murine megakaryocytes express nearly 5-fold lower levels of Survivin mRNA compared to erythroid cells. To investigate whether Survivin is involved in the differentiation and/or survival of hematopoietic progenitors, we infected primary mouse bone marrow cells with retroviruses harboring either the human Survivin cDNA or a mouse Survivin shRNA, and then induced erythroid and megakaryocyte differentiation in both liquid culture and colony-forming assays. These studies revealed that overexpression of Survivin promoted the terminal differentiation of red blood cells, while its reduction, by RNA interference, inhibited their differentiation. In contrast, downregulation of Survivin facilitated the expansion of megakaryocytes, and its overexpression antagonized megakaryocyte formation. In addition, consistent with a role for survivin in erythropoiesis, downregulation of Survivin expression in MEL cells led to a block in terminal differentiation. Finally, since caspase activity is known to be required for erythroid maturation, we investigated whether survivin associated with cleaved caspase-3 in erythroid cells. Immunofluorescence revealed that Survivin and cleaved caspase-3 co-localized to discrete foci within the cytoplasm of erythroid cells at the orthochromatic stage of development. Based on these findings, we hypothesize that Survivin cooperates with cleaved caspase-3 in terminal maturation of red blood cells. Together, our findings demonstrate that Survivin plays multiple, distinct roles in hematopoiesis.

Arginine Methylation Of RBM15 By PRMT1 Controls Alternative Splicing Of Genes Involved In Megakaryocyte Differentiation

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 2443-2443
Author(s):  
Xinyang Zhao ◽  
Li Zhang ◽  
Rui Wang ◽  
Ngoc Tung Trans ◽  
Hairui Su ◽  
...  
Keyword(s):  
Alternative Splicing ◽  
Rna Binding ◽  
Conflicts Of Interest ◽  
Chromosome Translocation ◽  
Arginine Methylation ◽  
Mass Spectrometry Analysis ◽  
Dependent Manner ◽  
Hematopoietic Stem ◽  
Spectrometry Analysis ◽  
Megakaryocyte Differentiation

Abstract More than 90% of under one year old infants with acute megakaryoblastic leukemia (AMKL) have chromosome translocation t(1;22)(p13;q13) with RBM15 fused to MKL1. RBM15 encodes an RNA binding protein important for hematopoietic stem cell self-renewal and differentiation. In agreement with its roles in AMKL, RBM15 is required for normal megakaryocyte differentiation. We found that higher expression of PRMT1 (Protein Arginine Methyltransferase) is commonly seen in M7 leukemia patient samples than other types of myeloid leukemia and that RBM15 is a bona fide methylation target for PRMT1. Using mass spectrometry analysis, we mapped the PRMT1 mediated mono-methylated site. The enzymatic activity of the PRMT1 V2 isoform is required for RBM15 degradation, as both shRNA molecules knocking down PRMT1 and small chemical PRMT1 inhibitors stabilize the RBM15 protein. Mutation of the methylation site to lysine blocks the ubiquitylation mediated degradation. Thus the degradation is a methylation dependent process. We identified the E3 ligase responsible for the degradation. Down-regulation of the RBM15 protein changes the isoform ratio of genes including GATA1 critical megakaryocyte differentiation. We found that RBM15 regulates its interaction with SF3B1A in methylation dependent manner during alternative splicing of GATA1 pre-mRNA. Thus, via methylation triggered RBM15 degradation, the megakaryocyte progenitor cells maintain a delicate balance between differentiation and proliferation by keeping the proper ratio of GATA1s and GATA1-full length mRNA. SF3B1A has been shown to be mutated in myeloid dysplasia syndrome and in several different types of leukemia. Methylation by PRMT1 links the two types of leukemic genes into a single pathway. Our results imply that targeting PRMT1/RBM15 pathway might be a potential therapy for AMKL and other blood malignancies. Disclosures: No relevant conflicts of interest to declare.

Ex vivo generation of fully mature human red blood cells from hematopoietic stem cells

2005 ◽  
Vol 23 (1) ◽  
pp. 69-74 ◽  
Author(s):  
Marie-Catherine Giarratana ◽  
Ladan Kobari ◽  
Hélène Lapillonne ◽  
David Chalmers ◽  
Laurent Kiger ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Red Blood Cells ◽  
Blood Cells ◽  
Ex Vivo ◽  
Hematopoietic Stem ◽  
Human Red Blood Cells

scholarly journals In VitroLarge Scale Production of Human Mature Red Blood Cells from Hematopoietic Stem Cells by Coculturing with Human Fetal Liver Stromal Cells

2013 ◽  
Vol 2013 ◽  
pp. 1-12 ◽  
Author(s):  
Jiafei Xi ◽  
Yanhua Li ◽  
Ruoyong Wang ◽  
Yunfang Wang ◽  
Xue Nan ◽  
...  
Keyword(s):  
Stem Cells ◽  
Hematopoietic Stem Cells ◽  
Cord Blood ◽  
Red Blood Cells ◽  
Stromal Cells ◽  
Blood Cells ◽  
Fetal Liver ◽  
Erythroid Differentiation ◽  
Erythroid Cells ◽  
Hematopoietic Stem

In vitromodels of human erythropoiesis are useful in studying the mechanisms of erythroid differentiation in normal and pathological conditions. Here we describe an erythroid liquid culture system starting from cord blood derived hematopoietic stem cells (HSCs). HSCs were cultured for more than 50 days in erythroid differentiation conditions and resulted in a more than 109-fold expansion within 50 days under optimal conditions. Homogeneous erythroid cells were characterized by cell morphology, flow cytometry, and hematopoietic colony assays. Furthermore, terminal erythroid maturation was improved by cosculturing with human fetal liver stromal cells. Cocultured erythroid cells underwent multiple maturation events, including decrease in size, increase in glycophorin A expression, and nuclear condensation. This process resulted in extrusion of the pycnotic nuclei in up to 80% of the cells. Importantly, they possessed the capacity to express the adult definitiveβ-globin chain upon further maturation. We also show that the oxygen equilibrium curves of the cord blood-differentiated red blood cells (RBCs) are comparable to normal RBCs. The large number and purity of erythroid cells and RBCs produced from cord blood make this method useful for fundamental research in erythroid development, and they also provide a basis for future production of available RBCs for transfusion.

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