Yolk sac erythromyeloid progenitors expressing gain of function PTPN11 have functional features of JMML but are not sufficient to cause disease in mice.

BACKGROUND: Accumulating evidence suggests the origin of juvenile myelomonocytic leukemia (JMML) is closely associated with fetal development. Nevertheless, the contribution of embryonic progenitors to JMML pathogenesis remains unexplored. We hypothesized that expression of JMML-initiating PTPN11 mutations in HSC-independent yolk sac erythromyeloid progenitors (YS EMPs) would result in a mouse model of pediatric myeloproliferative neoplasm (MPN). RESULTS: E9.5 YS EMPs from VavCre+;PTPN11D61Y embryos demonstrated growth hypersensitivity to granulocyte-macrophage colony-stimulating factor (GM-CSF) and hyperactive RAS-ERK signaling. Mutant EMPs engrafted the spleens of neonatal recipients, but did not cause disease. To assess MPN development during unperturbed hematopoiesis we generated CSF1R-MCM+;PTPN11E76K ;ROSAYFP mice in which oncogene expression was restricted to EMPs. Yellow fluorescent protein-positive progeny of mutant EMPs persisted in tissues one year after birth and demonstrated hyperactive RAS-ERK signaling. Nevertheless, these mice had normal survival and did not demonstrate features of MPN. CONCLUSIONS: YS EMPs expressing mutant PTPN11 demonstrate functional and molecular features of JMML but do not cause disease following transplantation nor following unperturbed development. Developmental Dynamics 246:1001-1014, 2017. © 2017 Wiley Periodicals, Inc.

A somatic mutation in erythro-myeloid progenitors causes neurodegenerative disease.

The pathophysiology of neurodegenerative diseases is poorly understood and there are few therapeutic options. Neurodegenerative diseases are characterized by progressive neuronal dysfunction and loss, and chronic glial activation. Whether microglial activation, which is generally viewed as a secondary process, is harmful or protective in neurodegeneration remains unclear. Late-onset neurodegenerative disease observed in patients with histiocytoses, which are clonal myeloid diseases associated with somatic mutations in the RAS-MEK-ERK pathway such as BRAF(V600E), suggests a possible role of somatic mutations in myeloid cells in neurodegeneration. Yet the expression of BRAF(V600E) in the haematopoietic stem cell lineage causes leukaemic and tumoural diseases but not neurodegenerative disease. Microglia belong to a lineage of adult tissue-resident myeloid cells that develop during organogenesis from yolk-sac erythro-myeloid progenitors (EMPs) distinct from haematopoietic stem cells. We therefore hypothesized that a somatic BRAF(V600E) mutation in the EMP lineage may cause neurodegeneration. Here we show that mosaic expression of BRAF(V600E) in mouse EMPs results in clonal expansion of tissue-resident macrophages and a severe late-onset neurodegenerative disorder. This is associated with accumulation of ERK-activated amoeboid microglia in mice, and is also observed in human patients with histiocytoses. In the mouse model, neurobehavioural signs, astrogliosis, deposition of amyloid precursor protein, synaptic loss and neuronal death were driven by ERK-activated microglia and were preventable by BRAF inhibition. These results identify the fetal precursors of tissue-resident macrophages as a potential cell-of-origin for histiocytoses and demonstrate that a somatic mutation in the EMP lineage in mice can drive late-onset neurodegeneration. Moreover, these data identify activation of the MAP kinase pathway in microglia as a cause of neurodegeneration and this offers opportunities for therapeutic intervention aimed at the prevention of neuronal death in neurodegenerative diseases.

Caspase-3 is involved in the signalling in erythroid differentiation by targeting late progenitors.

A role for caspase activation in erythroid differentiation has been established, yet its precise mode of action remains elusive. A drawback of all previous investigations on caspase activation in ex vivo erythroid differentiation is the lack of an in vitro model producing full enucleation of erythroid cells. Using a culture system which renders nearly 100% enucleated red cells from human CD34(+) cells, we investigated the role of active caspase-3 in erythropoiesis. Profound effects of caspase-3 inhibition were found on erythroid cell growth and differentiation when inhibitors were added to CD34(+) cells at the start of the culture and showed dose-response to the concentration of inhibitor employed. Enucleation was only reduced as a function of the reduced maturity of the culture and the increased cell death of mature cells while the majority of cells retained their ability to extrude their nuclei. Cell cycle analysis after caspase-3 inhibition showed caspase-3 to play a critical role in cell proliferation and highlighted a novel function of this protease in erythroid differentiation, i.e. its contribution to cell cycle regulation at the mitotic phase. While the effect of caspase-3 inhibitor treatment on CD34(+) derived cells was not specific to the erythroid lineage, showing a similar reduction of cell expansion in myeloid cultures, the mechanism of action in both lineages appeared to be distinct with a strong induction of apoptosis causing the decreased yield of myeloid cells. Using a series of colony-forming assays we were able to pinpoint the stage at which cells were most sensitive to caspase-3 inhibition and found activated caspase-3 to play a signalling role in erythroid differentiation by targeting mature BFU-E and CFU-E but not early BFU-E.

JAK2-V617F-mediated signalling is dependent on lipid rafts and statins inhibit JAK2-V617F-dependent cell growth.

Aberrant JAK2 signalling plays an important role in the aetiology of myeloproliferative neoplasms (MPNs). JAK2 inhibitors, however, do not readily eliminate neoplastic MPN cells and thus do not induce patient remission. Further understanding JAK2 signalling in MPNs may uncover novel avenues for therapeutic intervention. Recent work has suggested a potential role for cellular cholesterol in the activation of JAK2 by the erythropoietin receptor and in the development of an MPN-like disorder in mice. Our study demonstrates for the first time that the MPN-associated JAK2-V617F kinase localizes to lipid rafts and that JAK2-V617F-dependent signalling is inhibited by lipid raft disrupting agents, which target membrane cholesterol, a critical component of rafts. We also show for the first time that statins, 3-hydroxy-3-methyl-glutaryl coenzyme A (HMG-CoA) reductase inhibitors, widely used to treat hypercholesterolaemia, induce apoptosis and inhibit JAK2-V617F-dependent cell growth. These cells are more sensitive to statin treatment than non-JAK2-V617F-dependent cells. Importantly, statin treatment inhibited erythropoietin-independent erythroid colony formation of primary cells from MPN patients, but had no effect on erythroid colony formation from healthy individuals. Our study is the first to demonstrate that JAK2-V617F signalling is dependent on lipid rafts and that statins may be effective in a potential therapeutic approach for MPNs.

SOD2 deficiency in hematopoietic cells in mice results in reduced red blood cell deformability and increased heme degradation.

Among the three types of super oxide dismutases (SODs) known, SOD2 deficiency is lethal in neonatal mice owing to cardiomyopathy caused by severe oxidative damage. SOD2 is found in red blood cell (RBC) precursors, but not in mature RBCs. To investigate the potential damage to mature RBCs resulting from SOD2 deficiency in precursor cells, we studied RBCs from mice in which fetal liver stem cells deficient in SOD2 were capable of efficiently rescuing lethally irradiated host animals. These transplanted animals lack SOD2 only in hematopoietically generated cells and live longer than SOD2 knockouts. In these mice, approximately 2.8% of their total RBCs in circulation are iron-laden reticulocytes, with numerous siderocytic granules and increased protein oxidation similar to that seen in sideroblastic anemia. We have studied the RBC deformability and oxidative stress in these animals and the control group by measuring them with a microfluidic ektacytometer and assaying fluorescent heme degradation products with a fluorimeter, respectively. In addition, the rate of hemoglobin oxidation in RBCs from these mice and the control group were measured spectrophotometrically. The results show that RBCs from these SOD2-deficient mice have reduced deformability, increased heme degradation products, and an increased rate of hemoglobin oxidation compared with control animals, indicative of increased RBC oxidative stress.

Protein kinase D-HDAC5 signaling regulates erythropoiesis and contributes to erythropoietin cross-talk with GATA1.

In red cell development, the differentiation program directed by the transcriptional regulator GATA1 requires signaling by the cytokine erythropoietin, but the mechanistic basis for this signaling requirement has remained unknown. Here we show that erythropoietin regulates GATA1 through protein kinase D activation, promoting histone deacetylase 5 (HDAC5) dissociation from GATA1, and subsequent GATA1 acetylation. Mice deficient for HDAC5 show resistance to anemic challenge and altered marrow responsiveness to erythropoietin injections. In ex vivo studies, HDAC5(-/-) progenitors display enhanced entry into and passage through the erythroid lineage, as well as evidence of erythropoietin-independent differentiation. These results reveal a molecular pathway that contributes to cytokine regulation of hematopoietic differentiation and offer a potential mechanism for fine tuning of lineage-restricted transcription factors by lineage-specific cytokines.

JAK-STAT and AKT pathway-coupled genes in erythroid progenitor cells through ontogeny.

BACKGROUND: It has been reported that the phosphatidylinositol 3-kinase (PI3K)-AKT signaling pathway regulates erythropoietin (EPO)-induced survival, proliferation, and maturation of early erythroid progenitors. Erythroid cell proliferation and survival have also been related to activation of the JAK-STAT pathway. The goal of this study was to observe the function of EPO activation of JAK-STAT and PI3K/AKT pathways in the development of erythroid progenitors from hematopoietic CD34+ progenitor cells, as well as to distinguish early EPO target genes in human erythroid progenitors during ontogeny. METHODS: Hematopoietic CD34+ progenitor cells, isolated from fetal and adult hematopoietic tissues, were differentiated into erythroid progenitor cells. We have used microarray analysis to examine JAK-STAT and PI3K/AKT related genes, as well as broad gene expression modulation in these human erythroid progenitor cells. RESULTS: In microarray studies, a total of 1755 genes were expressed in fetal liver, 3844 in cord blood, 1770 in adult bone marrow, and 1325 genes in peripheral blood-derived erythroid progenitor cells. The erythroid progenitor cells shared 1011 common genes. Using the Ingenuity Pathways Analysis software, we evaluated the network pathways of genes linked to hematological system development, cellular growth and proliferation. The KITLG, EPO, GATA1, PIM1 and STAT3 genes represent the major connection points in the hematological system development linked genes. Some JAK-STAT signaling pathway-linked genes were steadily upregulated throughout ontogeny (PIM1, SOCS2, MYC, PTPN11), while others were downregulated (PTPN6, PIAS, SPRED2). In addition, some JAK-STAT pathway related genes are differentially expressed only in some stages of ontogeny (STATs, GRB2, CREBB). Beside the continuously upregulated (AKT1, PPP2CA, CHUK, NFKB1) and downregulated (FOXO1, PDPK1, PIK3CG) genes in the PI3K-AKT signaling pathway, we also observed intermittently regulated gene expression (NFKBIA, YWHAH). CONCLUSIONS: This broad overview of gene expression in erythropoiesis revealed transcription factors differentially expressed in some stages of ontogenesis. Finally, our results show that EPO-mediated proliferation and survival of erythroid progenitors occurs mainly through modulation of JAK-STAT pathway associated STATs, GRB2 and PIK3 genes, as well as AKT pathway-coupled NFKBIA and YWHAH genes.

Differential biological activity of disease-associated JAK2 mutants.

The JAK2V617F mutation has been identified in most patients with myeloproliferative neoplasms (MPNs), including polycythemia vera, essential thrombocythemia and primary myelofibrosis. Although JAK2V617F is the predominant allele associated with MPNs, other activating Janus kinase 2 (JAK2) alleles (such as K539L, T875N) also have been identified in distinct MPNs. The basis for the differences in the in vivo effects of different JAK2 alleles remains unclear. We have characterized three different classes of disease-associated JAK2 mutants (JAK2V617F, JAK2K539L and JAK2T875N) and found significant differences in biochemical, signaling and transforming properties among these different classes of JAK2 mutants.

Essential role for Mnk kinases in type II interferon (IFNgamma) signaling and its suppressive effects on normal hematopoiesis.

IFNγ exhibits potent antitumor effects and plays important roles in the innate immunity against cancer. However, the mechanisms accounting for the antiproliferative effects of IFNγ still remain to be elucidated. We examined the role of Mnk1 (MAPK-interacting protein kinase 1) in IFNγ signaling. Our data demonstrate that IFNγ treatment of sensitive cells results in engagement of Mnk1, activation of its kinase domain, and downstream phosphorylation of the cap-binding protein eIF4E on Ser-209. Such engagement of Mnk1 plays an important role in IFNγ-induced IRF-1 (IFN regulatory factor 1) gene mRNA translation/protein expression and is essential for generation of antiproliferative responses. In studies aimed to determine the role of Mnk1 in the induction of the suppressive effects of IFNs on primitive hematopoietic progenitors, we found that siRNA-mediated Mnk1/2 knockdown results in partial reversal of the suppressive effects of IFNγ on human CD34+-derived myeloid (CFU-GM) and erythroid (BFU-E) progenitors. These findings establish a key role for the Mnk/eIF4E pathway in the regulatory effects of IFNγ on normal hematopoiesis and identify Mnk kinases as important elements in the control of IFNγ-inducible ISG mRNA translation.

Interferon-gamma-mediated pathways are induced in human CD34(+) haematopoietic stem cells.

Pro-inflammatory cytokines like interferon-gamma (IFN-gamma) are considered to be important in the development of anaemia of chronic disease (ACD). Both, inhibitory and stimulatory activities of IFN-gamma on erythropoiesis have been observed in vitro earlier. IFN-gamma induces several biochemical pathways in human monocytes, among them neopterin formation by GTP-cyclohydrolase I (GTP-CH I) and tryptophan degradation by the enzyme indoleamine 2,3-dioxygenase (IDO). IDO-mediated tryptophan deprivation efficiently inhibits the growth of proliferating cells and microbes, thus we wanted to examine whether enhanced tryptophan degradation by monocytic precursor cells also suppresses erythropoiesis. Therefore, IFN-gamma-mediated pathways were investigated in human CD34(+) progenitor cells, and effects of IFN-gamma on the proliferative activity of different progenitor subpopulations were studied. Cells were either cultivated in agar-conditioned medium (ACM) or in medium containing erythroid growth factors interleukin-3 (IL-3) and stem cell factor (SCF; EGFCM). Stimulation of CD34(+) cells with IFN-gamma in different doses (either 5000U/ml once or 200 and 400U/ml every other day) induced tryptophan degradation and in parallel also neopterin formation. Unstimulated cells cultured with ACM produced higher amounts of neopterin and kynurenine (all p<0.05). IFN-gamma stimulated higher kynurenine and neopterin formation in cells cultivated in EGFCM, stimulation with 400U IFN-gamma every other day was most effective. IFN-gamma stimulated the growth and proliferation of CFU-E and BFU-E (3-8) in both media. In conclusion, stimulation of haematopoietic stem cells with IFN-gamma activates IDO and neopterin formation, and it also exerts an influence on the proliferation of various stem cell populations.

Theoretical and experimental analysis links isoform-specific ERK signalling to cell fate decisions.

Cell fate decisions are regulated by the coordinated activation of signalling pathways such as the extracellular signal-regulated kinase (ERK) cascade, but contributions of individual kinase isoforms are mostly unknown. By combining quantitative data from erythropoietin-induced pathway activation in primary erythroid progenitor (colony-forming unit erythroid stage, CFU-E) cells with mathematical modelling, we predicted and experimentally confirmed a distributive ERK phosphorylation mechanism in CFU-E cells. Model analysis showed bow-tie-shaped signal processing and inherently transient signalling for cytokine-induced ERK signalling. Sensitivity analysis predicted that, through a feedback-mediated process, increasing one ERK isoform reduces activation of the other isoform, which was verified by protein over-expression. We calculated ERK activation for biochemically not addressable but physiologically relevant ligand concentrations showing that double-phosphorylated ERK1 attenuates proliferation beyond a certain activation level, whereas activated ERK2 enhances proliferation with saturation kinetics. Thus, we provide a quantitative link between earlier unobservable signalling dynamics and cell fate decisions.

Hemozoin (malarial pigment) directly promotes apoptosis of erythroid precursors.

Severe malarial anemia is the most common syndrome of severe malaria in endemic areas. The pathophysiology of chronic malaria is characterised by a striking degree of abnormal development of erythroid precursors (dyserythropoiesis) and an inadequate erythropoietic response in spite of elevated levels of erythropoietin. The cause of dyserythropoiesis is unclear although it has been suggested that bone-marrow macrophages release cytokines, chemokines or lipo-peroxides after exposure to hemozoin, a crystalloid form of undigested heme moieties from malarial infected erythrocytes, and so inhibit erythropoiesis. However, we have previously shown that hemozoin may directly inhibit erythroid development in vitro and the levels of hemozoin in plasma from patients with malarial anemia and hemozoin within the bone marrow was associated with reduced reticulocyte response. We hypothesized that macrophages may reduce, not enhance, the inhibitory effect of hemozoin on erythropoiesis. In an in vitro model of erythropoiesis, we now show that inhibition of erythroid cell development by hemozoin isolated from P. falciparum is characterised by delayed expression of the erythroid markers and increased apoptosis of progenitor cells. Crucially, macrophages appear to protect erythroid cells from hemozoin, consistent with a direct contribution of hemozoin to the depression of reticulocyte output from the bone marrow in children with malarial anemia. Moreover, hemozoin isolated from P. falciparum in vitro inhibits erythroid development independently of inflammatory mediators by inducing apoptotic pathways that not only involve activation of caspase 8 and cleavage of caspase 3 but also loss of mitochondrial potential. Taken together these data are consistent with a direct effect of hemozoin in inducing apoptosis in developing erythroid cells in malarial anemia. Accumulation of hemozoin in the bone marrow could therefore result in inadequate reticulocytosis in children that have adequate levels of circulating erythropoietin.

Structure of the membrane proximal oxidoreductase domain of human Steap3, the dominant ferrireductase of the erythroid transferrin cycle.

The daily production of 200 billion erythrocytes requires 20 mg of iron, accounting for nearly 80% of the iron demand in humans. Thus, erythroid precursor cells possess an efficient mechanism for iron uptake in which iron loaded transferrin (Tf) binds to the transferrin receptor (TfR) at the cell surface. The Tf:TfR complex then enters the endosome via receptor-mediated endocytosis. Upon endosomal acidification, iron is released from Tf, reduced to Fe(2+) by Steap3, and transported across the endosomal membrane by divalent metal iron transporter 1. Steap3, the major ferrireductase in erythrocyte endosomes, is a member of a unique family of reductases. Steap3 is comprised of an N-terminal cytosolic oxidoreductase domain and a C-terminal heme-containing transmembrane domain. Cytosolic NADPH and a flavin are predicted cofactors, but the NADPH/flavin binding domain differs significantly from those in other eukaryotic reductases. Instead, Steap3 shows remarkable, although limited homology to FNO, an archaeal oxidoreductase. We have determined the crystal structure of the human Steap3 oxidoreductase domain in the absence and presence of NADPH. The structure reveals an FNO-like domain with an unexpected dimer interface and substrate binding sites that are well positioned to direct electron transfer from the cytosol to a heme moiety predicted to be fixed within the transmembrane domain. Here, we discuss possible gating mechanisms for electron transfer across the endosomal membrane.

Selective inhibition of JAK2-driven erythroid differentiation of polycythemia vera progenitors.

Polycythemia Vera (PV) is a myeloproliferative disorder (MPD) that is commonly characterized by mutant JAK2 (JAK2V617F) signaling, erythrocyte overproduction, and a propensity for thrombosis, progression to myelofibrosis, or acute leukemia. In this study, JAK2V617F expression by human hematopoietic progenitors promoted erythroid colony formation and erythroid engraftment in a bioluminescent xenogeneic immunocompromised mouse transplantation model. A selective JAK2 inhibitor, TG101348 (300 nM), significantly inhibited JAK2V617F+ progenitor-derived colony formation as well as engraftment (120 mg/kg) in xenogeneic transplantation studies. TG101348 treatment decreased GATA-1 expression, which is associated with erythroid-skewing of JAK2V617F+ progenitor differentiation, and inhibited STAT5 as well as GATA S310 phosphorylation. Thus, TG101348 may be an effective inhibitor of JAK2V617F+ MPDs in clinical trials.

Epigenetic analysis of the human alpha- and beta-globin gene clusters.

K562 erythroleukemia cells have been widely used as a model for the study of globin gene regulation. A number of agents have been shown to activate or suppress globin gene expression in these cells. However, the molecular effects of these agents on the epigenetic configuration of the alpha- and gamma-globin genes that encode HbF are not known. In this report, we investigated the relationship between globin expression and histone acetylation of the human alpha- and beta-globin clusters in the fetal erythroid environment of K562 cells. Our studies suggest that acetylation of histone H3 may be important in regulating developmental stage-specific expression of the different beta-like globin genes while acetylation of both histones H3 and H4 may be important for the regulation of tissue-specific expression of these genes. In contrast, acetylation of both histones H3 and H4 at the alpha-like globin promoters appears to be important for both developmental stage- and tissue-specific expression. Interestingly, butyrate-induced activation of alpha-globin gene expression in K562 cells is associated with significant increase in histone acetylation levels while TPA-induced inhibition is associated with decreased histone acetylation at its promoters. In contrast, changes in histone acetylation and DNA methylation do not appear to be important in the regulation of gamma-globin gene expression by the same agents. These data suggest that the butyrate-mediated induction of the fetal gamma-globin genes in K562 cells is not a direct result of its histone deacetylase inhibitor activity of butyrate on the chromatin of the gamma-globin promoters, while the induction of the alpha-globin genes could be a result of a direct effect of butyrate on chromatin at its promoters. This is another example of the important differences in the molecular mechanisms of regulation of the genes of the alpha- and beta-like globin clusters.

A role for caspases in the differentiation of erythroid cells and macrophages.

Several cysteine proteases of the caspase family play a central role in many forms of cell death by apoptosis. Other enzymes of the family are involved in cytokine maturation along inflammatory response. In recent years, several caspases involved in cell death were shown to play a role in other cellular processes such as proliferation and differentiation. In the present review, we summarize the current knowledge of the role of caspases in the differentiation of erythroid cells and macrophages. Based on these two examples, we show that the nature of involved enzymes, the pathways leading to their activation in response to specific growth factors, and the specificity of the target proteins that are cleaved by the activated enzymes strongly differ from one cell type to another. Deregulation of these pathways is thought to play a role in the pathophysiology of low-grade myelodysplastic syndromes, characterized by excessive activation of caspases and erythroid precursor apoptosis, and that of chronic myelomonocytic leukemia, characterized by a defective activation of caspases in monocytes exposed to M-CSF, which blocks their differentiation.

[Adaptogenic effect of the vitamin D3 containing supplement "videchol" on glucose-6-phosphate dehydrogenase activity in erythrone of irradiated rats].

Two fast migrating, major, multiple molecular forms (MMF) of glucose-6-phosphate dehydrogenase [EC:1.1.1.49]: G-6-PDH-1 and G-6-PDH-2, and two minor forms: G-6-PDH-3 and G-6-PDH-4 were revealed in the electrophoregrams of both erythrocytes haemolisates as well in the homogenates of bone marrow cellular lines of rats at control conditions. Daily 1 cGy irradiation of rats up to a cumulative dose of 20 cGy led to a drop of G-6-PDH total activity and it caused a redistribution of the MMF of the enzyme in bone marrow cellular populations. However, G-6-PDH activity in erythrocytes exceeded the control means in all the experimental terms. The calculation of the local redistribution coefficient (l(G-6-FDH-i)) showed that these changes are mainly determined by the increase of the activity of the isoform G-6-PDH-3. Vitamin D3 administration to rats generated a correction of G-6-PDH activity in all studied cellular populations. Meanwhile, the MMF profiles were characterized by multidirectional rearrangements in the bone marrow erythroid and granulocyte-monocyte cells and in erythrocytes. The specificity of changes in the distribution of the MMF of G-6-PDH in the three studied cellular populations depends on the particularities of their energetic metabolism at irradiation conditions and on the modifying action of the natural adaptogen 1,25-dihydroxicholecalciferol.

Cdc42 critically regulates the balance between myelopoiesis and erythropoiesis.

The Rho GTPase Cdc42 regulates adhesion, migration, and homing, as well as cell cycle progression, of hematopoietic stem cells, but its role in multilineage blood development remains unclear. We report here that inducible deletion of cdc42 in cdc42-floxed mouse bone marrow by the interferon-responsive, Mx1-Cre-mediated excision led to myeloid and erythroid developmental defects. Cdc42 deletion affected the number of early myeloid progenitors while suppressing erythroid differentiation. Cdc42-deficient mice developed a fatal myeloproliferative disorder manifested by significant leukocytosis with neutrophilia, myeloid hyperproliferation, and myeloid cell infiltration into distal organs. Concurrently, Cdc42 deficiency caused anemia and splenomegaly accompanied with decreased bone marrow erythroid burst-forming units (BFU-Es) and colony-forming units-erythroid (CFU-Es) activities and reduced immature erythroid progenitors, suggesting that Cdc42 deficiency causes a block in the early stage of erythropoiesis. Cdc42 activity is responsive to stimulation by SCF, IL3, SDF-1alpha, and fibronectin. The increased myelopoiesis and decreased erythropoiesis of the knockout mice are associated with an altered gene transcription program in hematopoietic progenitors, including up-regulation of promyeloid genes such as PU.1, C/EBP1alpha, and Gfi-1 in the common myeloid progenitors and granulocyte-macrophage progenitors and down-regulation of proerythroid gene such as GATA-2 in the megakaryocyte-erythroid progenitors. Thus, Cdc42 is an essential regulator of the balance between myelopoiesis and erythropoiesis.

[The influence of some retarding agents NOS of dihydrothiazine-thiazoline rank on postradiational of recovery endogenous CFU-S-8 of mice].

In this work the attempt to estimate a nitric oxide (NO*) role in regulation of the number of pool haemopoietic stem cells at the irradiated mice was made. With this purpose the number of new compounds from dihydrothiazine-thiazoline line was synthesized, their NO-inhibiting activity was investigated in vivo by the method of ESR-spectroscopy of spin trap and their influence on an output endogenous spleen colonies (CFU-S-8) after the total sublethal y-irradiation of mice in a doze of 6 Gy was also investigated. Was shown, that the tested compounds reduced the contents of NO* in a liver tissue of mice which have received an injection of nitric oxide synthesis inductor - lipopolysaccharide, and also increased an output CFU-S-8 forming endogenous colonies in the spleen of the irradiated mice. Received data testify to perceptivity of search radioprotective agents among NO* synthesis inhibitors.