Cell cycle.independent roles of p19INK4d in human terminal erythropoiesis

Normally, cyclin interacts with cyclin-dependent kinase (CDK) to form a cyclin-CDK complex, which promotes cell cycle progression, whereas cyclin-dependent kinase inhibitor (CDKI) molecules inhibit the formation of cyclin- CDK complex, arresting cell cycle. Terminal erythropoiesis is closely coordinated with cell cycle exit, which is regulated by cyclins, CDKs, and CDKIs [1]. In the global transcriptome of human terminal erythropoiesis [2], p19INK4d is expressed highly, and its expression is significantly up-regulated during human terminal erythropoiesis. However, the roles of p19INK4d in terminal erythropoiesis are still unknown.

Erythropoiesis Stimulating Agents (ESAs) in the Treatment of Cardio-Renal Syndrome Anaemia

Coexistence of chronic kidney disease (CKD) and chronic heart failure (CHF) define a recently recognized clinical entity known as cardio-renal syndrome. Sufficient evidence suggests that the two pathological conditions share common pathogenic etiology which is not yet fully defined. Superimposed anaemia is a common finding among patients suffering from cardio-renal syndrome. The combination of CKD, CHF and anaemia increase the probability of death by 6 times compared to normal individuals. Early attempts to restore anaemia either by iron supplementation, erythropoiesis stimulating agents (ESAs) or combination of the two have reported to improve quality of life, morbidity and mortality especially among patients treated by cardiologists. Recent publications of well controlled epidemiological studies failed to prove convincing beneficial effect of the above mentioned therapy moreover skepticism has raised concerning the safety of restoring anaemia among patients with cardio-renal syndrome as well as used medications. There are still unresolved problems concerning the definition of anaemia, by means of hemoglobin level among these patients, the target hemoglobin level and the therapeutic regimen of ESAs administration and iron supplementation. We need much more evidence in order to define an effective and safe treatment strategy correcting anaemia among patients with cardio-renal syndrome.

Iron-Restricted Erythropoiesis in Anaemic Patients with Giant Cell Arteritis and Polymyalgia Rheumatica

The aim of this observational study was to biochemically characterize the anaemia in GCA (giant cell arteritis) and PMR (polymyalgia rheumatica) patients. Values for mean corpuscular volume, mean corpuscular hemoglobin and soluble transferrin receptor were normal, whereas serum iron and total iron binding capacity (TIBC) were subnormal, and mean ferritin was above the upper reference limit. Iron-restricted erythropoiesis (IRE), defined as a bone marrow smear staining positive for iron in combination with transferrin saturation less than 20%, was present in all patients. All patients exhibited clinical and biochemical signs of active inflammation with elevated C-reactive protein and an increased erythrocyte sedimentation rate.

Comprehensive Characterization and Global Transcriptome Analysis of Human Fetal Liver Terminal Erythropoiesis

The fetal liver(FL)is the key erythropoietic organ during fetal development,but knowledge on human FL erythropoiesis is very limited.In this study,we sorted primary erythroblasts from FL cells and performed RNA sequencing(RNA-seq)analyses.We found that temporal gene expression patterns reflected changes in function during primary human FL terminal erythropoiesis.Notably,the expression of genes enriched in proteolysis and autophagy was up-regulated in orthochromatic erythroblasts(OrthoEs),suggesting the involvement of these pathways in enucleation.We also performed RNA-seq of in vitro cultured erythroblasts derived from FL CD34+cells.Comparison of transcriptomes between the primary and cultured erythroblasts revealed significant differences,indicating impacts of the culture system on gene expression.Notably,the expression of lipid metabolism-related genes was increased in cultured erythroblasts.We further immortalized erythroid cell lines from FL and cord blood(CB)CD34+cells(FL-iEry and CB-iEry,respectively).FL-iEry and CB-iEry were immortalized at the proerythroblast stage and can be induced to differentiate into OrthoEs,but their enucleation ability was very low.Comparison of the transcriptomes between OrthoEs with and without enucleation capability revealed the down-regulation of pathways involved in chromatin organization and mitophagy in OrthoEs without enucleation capacity,indicating that defects in chromatin organization and mitophagy contribute to the inability of OrthoEs to enucleate.Additionally,the expression of HBE1,HBZ,and HBG2 was up-regulated in FL-iEry compared with CB-iEry,and such up-regulation was accompanied by down-regulated expression of BCL11A and up-regulated expression of LIN28B and IGF2BP1.Our study provides new insights into human FL erythropoiesis and rich resources for future studies.

Post-transcriptional regulation of erythropoiesis

Erythropoiesis is a complex,precise,and lifelong process that is essential for maintaining normal body functions.Its strict regulation is necessary to prevent a variety of blood diseases.Normal erythropoiesis is precisely regulated by an intricate network that involves transcription levels,signal transduction,and various epigenetic modifications.In recent years,research on posttranscriptional levels in erythropoiesis has expanded significantly.The dynamic regulation of splicing transitions is responsible for changes in protein isoform expression that add new functions beneficial for erythropoiesis.RNA-binding proteins adapt the translation of transcripts to the protein requirements of the cell,yielding mRNA with dynamic translation efficiency.Noncoding RNAs,such as microRNAs and lncRNAs,are indispensable for changing the translational efficiency and/or stability of targeted mRNAs to maintain the normal expression of genes related to erythropoiesis.N6-methyladenosine-dependent regulation of mRNA translation plays an important role in maintaining the expression programs of erythroid-related genes and promoting erythroid lineage determination.This review aims to describe our current understanding of the role of post-transcriptional regulation in erythropoiesis and erythroid-associated diseases,and to shed light on the physiological and pathological implications of the post-transcriptional regulation machinery in erythropoiesis.These may help to further enrich our understanding of the regulatory network of erythropoiesis and provide new strategies for the diagnosis and treatment of erythroid-related diseases.

Long non-coding RNAs during normal erythropoiesis

The transcription of essentially the entire eukaryotic genome produces a huge amount of non-coding RNAs.Among them,long noncoding RNAs(lncRNAs)consist of a significant portion that widely exists across mammal genome,generating from high-throughput transcriptomic studies in the last decade.Although the functions of most lncRNAs remain to be further investigated,many of them have already been shown to play critical roles during normal development and disease conditions.Increasing evidence indicates that lncRNAs involve in versatile biological processes during erythroid proliferation and differentiation,including erythroid cell survival,heme metabolism,globin switching and regulation,erythroid enucleation,etc,via cis-or trans-mediated molecular mechanisms.In this review,we focus on recent advances regarding the functions and mechanisms of lncRNAs in normal erythropoiesis.

Crosstalk between catecholamines and erythropoiesis

BACKGROUND： Erythropoiesis is regulated by a range of intrinsic and extrinsic factors, including different cytokines. Recently, the role of catecholamines has been highlighted in the development of erythroid cell lineages. OBJECTIVE： This study focuses on the biological links interconnecting erythroid development and the sympathetic nervous system. The emerging evidence that underscores the role of catecholamines in the regulation of erythropoietin and other erythropoiesis cytokines are thoroughly reviewed, in addition to elements such as iron and the leptin hormone that are involved in erythropoiesis. METHODS： Relevant English-language studies were identified and retrieved from the PubMed search engine （1981-2017） using the following keywords： ＂Erythropoiesis＂, ＂Catecholamines＂, ＂Nervous system＂, and ＂Cytokines.＂ RESULTS： Chronic social stress alters and suppresses erythroid development. However, the physiological release of catecholamines is an additional stimulator of erythropoiesis in the setting of anemia. Therefore, the severity and timing of catecholamine secretion might distinctly regulate erythroid homeostasis. CONCLUSION： Understanding the relationship of catecholamines with different elements of the erythroid islands will be essential to find the tightly regulated production of red blood cells （RBCs） in both chronic and physiological catecholamine activation.

MicroRNAs in erythropoiesis and red blood cell disorders

MicroRNAs （miRNAs） are 19-24 nucleotide non-coding ribonucleic acids binding DNA or RNA and controlling gene expression via mRNA degradation or its transcription inhibition. Erythropoies is a multi step differentiation process of erythroid progenitors to nucleate red blood cells. Maturation, proliferation and differentiation of red blood cells is affected by erythroid factors, signaling pathways in niche of hematopoietic cells, transcription factors as well as miRNAs. Expression of different types of miRNAs during erythroid development provides a background for the study of these molecules to control erythroid differentiation and maturation as well as their use as diagnostic and prognostic markers to treat erythroid disorders like thalassemia, sickle cell disease and erythrocyte enzyme deficiencies. In this paper, with reference to biosynthesis of miRNAs, their function in normal and anemic erythropoiesis has been investigated. The target molecule of each of these miRNAs has been cited in an attempt to elucidate their role in erythropoiesis.

Zebrafish ELL-associated factors Eaf1/2 modulate erythropoiesis via regulating gata1a expression and WNT signaling to facilitate hypoxia tolerance

EAF1 and EAF2,the eleven-nineteen lysine-rich leukemia(ELL)-associated factors which can assemble to the super elongation complex(AFF1/4,AF9/ENL,ELL,and P-TEFb),are reported to participate in RNA polymeraseⅡto actively regulate a variety of biological processes,including leukemia and embryogenesis,but whether and how EAF1/2 function in hematopoietic system related hypoxia tolerance during embryogenesis remains unclear.Here,we unveiled that deletion of EAF1/2(eaf1^(-/-)and eaf2^(-/-))caused reduction in hypoxia tolerance in zebrafish,leading to reduced erythropoiesis during hematopoietic processes.Meanwhile,eaf1^(-/-)and eaf2^(-/-)mutants showed significant reduc-tion in the expression of key transcriptional regulators scl,lmo2,and gata1a in erythropoiesis at both 24 h post fertilization(hpf)and 72 hpf,with gata1a downregulated while scl and lmo2 upregulated at 14 hpf.Mechanistically,eaf1^(-/-)and eaf2^(-/-)mutants exhibited significant changes in the expression of epigenetic modified histones,with a significant increase in the binding enrichment of modified histone H3K27me3 in gata1a promoter rather than scl and lmo2 promoters.Additionally,eaf1^(-/-)and eaf2^(-/-)mutants exhibited a dynamic expression of canonical WNT/β-catenin signaling during erythropoiesis,with significant reduction in p-β-Catenin level and in the binding enrichment of both scl and lmo2 promoters with the WNT transcriptional factor TCF4 at 24 hpf.These findings demonstrate an important role of Eaf1/2 in erythropoiesis in zebrafish and may have shed some light on regeneration medicine for anemia and related diseases and on molecular basis for fish economic or productive traits,such as growth,disease resistance,hypoxia tolerance,and so on.

Stem cell autotomy and niche interaction in different systems

The best known cases of cell autotomy are the formation of erythrocytes and thrombocytes(platelets) from progenitor cells that reside in special niches. Recently, autotomy of stem cells and its enigmatic interaction with the niche has been reported from male germline stem cells(GSCs) in several insect species. First described in lepidopterans, the silkmoth, followed by the gipsy moth and consecutively in hemipterans, foremost the milkweed bug. In both, moths and the milkweed bug, GSCs form finger-like projections toward the niche, the apical cells(homologs of the hub cells in Drosophila). Whereas in the milkweed bug the projection terminals remain at the surface of the niche cells, in the gipsy moth they protrude deeply into the singular niche cell. In both cases, the projections undergo serial retrograde fragmentation with progressing signs of autophagy. In the gipsy moth, the autotomized vesicles are phagocytized and digested by the niche cell. In the milkweed bug the autotomized vesicles accumulate at the niche surface and disintegrate. Autotomy and sprouting of new projections appears to occur continuously. The significance of the GSC-niche interactions, however, remains enigmatic. Our concept on the signaling relationship between stem cell-niche in general and GSC and niche(hub cells and cyst stem cells) in particular has been greatly shaped by Drosophila melanogaster. In comparing the interactions of GSCs with their niche in Drosophila with those in species exhibiting GSC autotomy it is obvious that additional or alternative modes of stem cell-niche communication exist. Thus, essential signaling pathways, including niche-stem cell adhesion(E-cadherin) and the direction of asymmetrical GSC division- as they were found in Drosophila- can hardly be translated into the systems where GSC autotomywas reported. It is shown here that the serial autotomy of GSC projections shows remarkable similarities with Wallerian axonal destruction, developmental axon pruning and dying-back degeneration in neurodegenerative diseases. Especially the hypothesis of an existing evolutionary conserved "autodestruction program" in axons that might also be active in GSC projections appears attractive. Investigations on the underlying signaling pathways have to be carried out. There are two other well known cases of programmed cell autotomy: the enucleation of erythroblasts in the process of erythrocyte maturation and the segregation of thousands of thrombocytes(platelets) from one megakaryocyte. Both progenitor cell types- erythroblasts and megakaryocytes- are associated with a niche in the bone marrow, erythroblasts with a macrophage, which they surround, and the megakaryocytes with the endothelial cells of sinusoids and their extracellular matrix. Although the regulatory mechanisms may be specific in each case, there is one aspect that connects all described processes of programmed cell autotomy and neuronal autodestruction: apoptotic pathways play always a prominent role. Studies on the role of male GSC autotomy in stem cell-niche interaction have just started but are expected to reveal hitherto unknown ways of signal exchange. Spermatogenesis in mammals advance our understanding of insect spermatogenesis. Mammal and insect spermatogenesis share some broad principles, but a comparison of the signaling pathways is difficult. We have intimate knowledge from Drosophila, but of almost no other insect, and we have only limited knowledge from mammals. The discovery of stem cell autotomy as part of the interaction with the niche promises new general insights into the complicated stem cell-niche interdependence.

Does Carica papaya leaf-extract increase the platelet count? An experimental study in a murine model

Objective:To investigate the potential role of fresh Carica papaya(C.papaya)leaf extract on haematological and biochemical parameters and toxicological changes in a murine model.Methods:In total 36 mice were used for the trial.Fresh C.papaya leaf extract[0.2 mL(2 g)/mouse]was given only to the test group(18 mice).General behavior,clinical signs and feeding patterns were recorded.Blood and tissue samples were collected at intervals.Haematological parameters including platelet,red blood cell(RBC),white blood cell<(WBC),packed cell volume(PCV),serum biochemistry including serum creatinine,serum glutamic-oxaloacetic transaminase(SCOT)and serum glutamic-pyruvic transaminase(SGPT)were determined.Organs for possible histopathological changes were examined.Results:Neither group exhibited alteration of behavior or reduction in food and water intake.Similarly,no significant changes in SCOT,SGPT and serum creatinine levels were delected in the test group.Histopathological organ changes were not observed in either group of mice except in three liver samples of the test group which had a mild focal necrosis.The platelet count(11.33±0.35)×10~5VμL(P=0.00004)and the RBC count(7.97±0.61)×10~/μL(P=0.00003)were significandy increased in the test group compared to that of the controls.However,WBC count and PCV(%)values were not changed significantly in the test group.The platelet count in the test group started to increase significantly from Day 3(3.4±0.18×10~5/μL),reaching almost a fourfold higher at Day 21(11.3×10~5/μL),while it was 3.8X10^5/μL and 5.5×10~5/μL at Day 3 and Day 21 respectively in the control.Likewise,the RBC count in the test group increased from 6×10~6/μL to 9×10~6/μL at Day 21 while it remained near constant in the control group(6×10~6μL).Conclusions:Fresh C.papaya leaf extract significandy increased the platelet and RBC counts in the test group as compared to controls.Therefore,it is very important to identify those chemicals of C.papaya leaves as it can be recommended to be used as a medication to boost thrombopoiesis and erythropoiesis in humans and in animals in which these cell lineages have been compromised.

Pharmacodynamics of CNTO 530 and Darbepoetin-α in Human TNF-α Transgenic Mice, a Murine Model of Anemia of Chronic Disease

CNTO 530 and darbepoetin-a are long lived erythropoietin receptor agonists (ERAs). Clinically, anemia of chronic disease (ACD) is associated with increased expression of tumor necrosis factor-a (TNF-a) and mice transgenic for human TNF-a develop ACD. The purpose of this investigation was to compare the effects of these agents in a murine model of ACD. Human TNF-a expressing (Tg 197) mice were administered a single subcutaneous dose of CNTO 530 or darbepoetin-a and the pharmacodynamic response in bone marrow spleen and peripheral blood evaluated. RBC life span and reticulocyte age distribution were also evaluated. CNTO 530 induced a dose responsive increase in reticulocytes, RBCs and Hgb in both wild type and Tg197 mice. Although the reticulocyte response was similar to wild types, the RBC and Hgb response to CNTO 530 in Tg197 mice was blunted. There was no statistically significant difference in RBC life span with either compound. Darbepoetin-α caused a greater peak in % dead Pro/basophilic erythroblasts, greater peak EMH in the spleen and a greater increase in reticulocyte maturation time. In contrast, despite a similar peak increase, CNTO 530 caused a more sustained response of reticulocyte, EMH, RBC and Hgb, consistent with increased exposure. In conclusion, CNTO 530 and darbepoetin-a increased RBC and hemoglobin in a murine model of ACD. Compared to darbepoetin-a, CNTO 530 had a more sustained effect, consistent with increased exposure.

Pharmacological Adjuvants to Limit Erythropoietin Stimulating Agents Exposure

Anemia in chronic kidney disease (CKD) is common, causing morbidity and mortality, and is primarily due to reduced erythropoietin (EPO) release and, to a lesser degree, shortened red cell survival. Erythropoietin Stimulating Agents like epoetin Alfa and darbepoetin alpha are used commonly to treat this form of anemia. Recent evidence suggests increased morbidity and mortality associated with higher hemoglobin in the setting of these agents use. Whether these complications are due to higher dose of erythropoietin or its resistance (i.e. inflammation), or achieving a higher hemoglobin remains unclear. Tightening restrictions on these agents has led to increase interest in the use of non-ESA adjuvants to improve erythropoiesis. This review will highlight the most promising of these agents.

Mapping Human Pluripotent Stem Cell-derived Erythroid Differentiation by Single-cell Transcriptome Analysis

There is an imbalance between the supply and demand of functional red blood cells(RBCs)in clinical applications.This imbalance can be addressed by regenerating RBCs using several in vitro methods.Induced pluripotent stem cells(iPSCs)can handle the low supply of cord blood and the ethical issues in embryonic stem cell research,and provide a promising strategy to eliminate immune rejection.However,no complete single-cell level differentiation pathway exists for the iPSC-derived erythroid differentiation system.In this study,we used iPSC line BC1 to establish a RBC regeneration system.The 10X Genomics single-cell transcriptome platform was used to map the cell lineage and differentiation trajectory on day 14 of the regeneration system.We observed that iPSC differentiation was not synchronized during embryoid body(EB)culture.The cells(on day 14)mainly consisted of mesodermal and various blood cells,similar to the yolk sac hematopoiesis.We identified six cell classifications and characterized the regulatory transcription factor(TF)networks and cell-cell contacts underlying the system.iPSCs undergo two transformations during the differentiation trajectory,accompanied by the dynamic expression of cell adhesion molecules and estrogen-responsive genes.We identified erythroid cells at different stages,such as burst-forming unit erythroid(BFU-E)and orthochromatic erythroblast(ortho-E)cells,and found that the regulation of TFs(e.g.,TFDP1 and FOXO3)is erythroid-stage specific.Immune erythroid cells were identified in our system.This study provides systematic theoretical guidance for optimizing the iPSC-derived erythroid differentiation system,and this system is a useful model for simulating in vivo hematopoietic development and differentiation.

Erythroblast island macrophages: recent discovery and future perspectives

Erythroblastic island(EBI),composed of a central macrophage surrounded by developing erythroid cells,is a structure found in hematopoietic tissues such as fetal liver and bone marrow.It is the first described hematopoietic niche that predominantly supports erythropoiesis.Although it is well accepted that EBIs and EBI macrophage play important roles during erythropoiesis,the mechanisms by which they support erythropoiesis remain largely unclear due to our inability to identify and isolate EBI macrophages.Earlier efforts to identify surface markers for EBI macrophages have focused on the adhesion molecules which are involved in macrophage’s interaction with erythroblasts.These include EMP,Vcam1,CD169,CD163,and aV integrin.Findings from these earlier studies suggested that combination of Vcam1,CD169,and mouse macrophage surface marker F4/80 can be used to define mouse EBI macrophage.We found that not all F4/80+Vcam1+CD169+macrophages are EBI macrophages.Instead,we discovered that EBI macrophages are characterized by the expression of Epor in both mouse and man.RNA-seq analyses of the newly identified EBI macrophages revealed that EBI macrophages have involved specialized function in supporting erythropoiesis.Our findings provide foundation for future studies.Here we will review current knowledge of EBI macrophages and discuss future perspectives.

Kzp Regulates the Transcription of gata2 and pu.1 during Primitive Hematopoiesis in Zebrafish Embryos

Kaiso zinc finger-containing protein（Kzp）,a maternally-derived transcription factor,controls dorsoventral patterning during zebrafish gastrulation.Here,we uncovered a new function for Kzp in zebrafish embryonic primitive hematopoiesis.The depletion of kzp led to defects in primitive hematopoiesis including the development of the erythroid and myeloid lineages.On the other hand,overexpression of kzp caused the ectopic expression of gata1,gata2,and pu.1.Chromosome immunoprecipitation assays revealed that Kzp protein directly binds to gata1,gata2,and pu.1 promoters.Interestingly,the ectopic expression of gatal was able to rescue the erythroid,but not the myeloid lineage in kzp-depleted zebrafish embryos.gata1 expression controlled by Kzp was dependent on gatal during primitive erythropoiesis.Our results indicate that Kzp is a critical transcriptional factor for the expression of gata2 and pu.1 to modulate primitive hematopoiesis.

Recombinant Human Prolactin Protects against Irradiation-Induced Myelosuppression

Prolactin is a multifunctional hormone that exerts many separate functions and acts as an important connection between the endocrine and immune systems. There are increasing researches implicating the role of prolactin in hematopoiesis. Enhanced erythropoiesis in pregnant women and direct erythropoietic effects in vitro of plasma either from pregnant or lactating mice have been reported. Furthermore, regression of erythroblastic leukemia has been observed in a significant number of rats after hypophysectomy. In this study, the effects of recombinant human prolactin （rhPRL） on hematopoiesis were assessed in irradiated mice. Mice were treated with rhPRL for five consecutive days after exposure to a lethal dose or a sub-dose irradiation. Prolonged survival rate and increased erythropoiesis were observed in the irradiation-induced myelosuppressive mice. It was concluded that rhPRL might act on erythropoiesis and could be a potential candidate for the treatment of irradiation-induced myelosuppresion in clinic.