Sotatercept, an Activin Receptor IIa Ligand Trap, Acts Through Bone Marrow Accessory Cells to Promote Late-Stage Erythropoiesis and a Rapid Induction of Red Blood Cell Number and Hemoglobin

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 372-372
Author(s):  
Soraya Carrancio ◽  
Jennifer A Markovics ◽  
Piu Wong ◽  
Jim Leisten ◽  
Matthew C Groza ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Blood Cell ◽  
Red Blood Cell ◽  
Erythroid Differentiation ◽  
Cell Number ◽  
Erythroid Progenitors ◽  
Activin Receptor ◽  
Accessory Cells ◽  
Tgfβ Superfamily

Abstract Abstract 372 The regulation of erythropoiesis requires stem cell factor and erythropoietin (EPO) for the proliferation and survival of erythroid progenitor and early precursor cells. While recombinant EPO is widely used for treating various types of anemia, it often lacks efficacy in cases of anemia due to ineffective erythropoiesis in which immature erythroid precursors undergo apoptosis. Thus, there is an need for new therapies to treat the later stages of erythropoiesis. Members of the transforming growth factor beta (TGFβ) superfamily have been studied as potential regulators of erythropoiesis, iron regulation and globin expression. Sotatercept (ACE-011), a recombinant fusion protein consisting of the extracellular domain of the human activin receptor IIA (ActRIIA) linked to the human immunoglobulin G1 (IgG1) Fc domain, is a ligand trap which binds a number of TGFβ superfamily ligands including activin A, activin B, growth differentiation factor-11 (GDF-11) and bone morphogenetic protein-10 (BMP-10). Administration of sotatercept led to substantial increases in red cell number and hemoglobin in human subjects, but the mechanism is not fully understood. We utilized both mouse in vivo and human in vitro models to investigate the mechanism of sotatercept in promoting erythropoiesis. In order to compare the effects of RAP-011 (the murine version of sotatercept) to EPO on red blood cell (RBC) parameters, C57/Bl mice were dosed with RAP-011, EPO or control vehicle. RAP-011-treated mice had a rapid and statistically significant increase in hematocrit, hemoglobin, and RBC number in less than 72-hours. As rapidly as 24 hours after treatment, RAP-011 induced a significant increase in RNA-negative, enucleated cells in the bone marrow (BM). RAP-011 also rapidly increased BM BFU-e and CFU-e erythroid progenitors, while EPO was more effective on spleen-derived progenitors. These data suggest that RAP-011 acts primarily on both bone marrow progenitor cells and late erythroblasts to promote erythropoiesis. In order to investigate the cellular mechanism by which RAP-011 increases red blood cell parameters, we conducted a series of in vitro experiments and found no evidence to support direct effects of RAP-011 on human CD34+ cells assessed in colony formation assays and in erythroid differentiation in liquid culture. As both clinical and pharmacological findings point to a clear role for RAP-011 in stimulating RBC parameters, we hypothesized that RAP-011 effects may be mediated by accessory cells in the BM microenvironment. Human CD36+ cells, which are highly enriched for erythroid progenitors, were co-cultured with long-term BM cultures and erythroid differentiation was assessed following 6 days of culture in EPO (2U/mL)-supplemented media. At day 6 the output of these cultures was predominantly characterized as EryA (∼basophilic erythroblast) but with the addition of RAP-011 (50μM), a significant fraction of CD36+ cells matured into EryB/C cells (polychromatic/orthochromatic erythroblasts), suggesting that factors produced by BM accessory cells mediate RAP-011 erythropoietic effects and that, in contrast to EPO, RAP-011 may play a role in the latter stages of erythroblast maturation. To identify cytokines that may mediate RAP-011 effects, CD36+ cells were treated with several activin receptor IIA ligands. GDF-11 treatment significantly decreased proliferation of GPA+ cells during the differentiation process and RAP-011 effectively reversed this effect, but had no consequence on untreated cells. These data suggest that GDF-11 may mediate the erythroipoietic stimulatory effects of RAP-011. In summary, RAP-011 induced a rapid increase in RBC parameters in mice (reflected in the number of enucleated cells found in the bone marrow), likely mediated by BM accessory cells. Our data also suggest that effects of sotatercept may be mediated at least partly by GDF-11, acting as a potential negative regulator of the terminal stages of erythropoiesis. The ability of sotatercept to reverse this inhibition would lead to a rapid release of terminal erythroid cells into the circulation. These data support the rationale to develop sotatercept for the treatment of anemia and ineffective erythropoiesis, especially in patients who may not respond to EPO. Disclosures: Carrancio: Celgene Corporation: Employment. Markovics:Celgene Corporation: Employment. Wong:Celgene Corporation: Employment. Leisten:Celgene Corporation: Employment. Groza:Celgene Corporation: Employment. Raymon:Celgene Corporation: Employment. Heise:Celgene Corporation: Employment. Chopra:Celgene Corp: Employment, Equity Ownership. Daniel:Celgene Corporation: Employment. Sung:Celgene Corporation: Employment.

The Presence of Novel Amino Acids in the Cytoplasmic Domain of Stem Cell Factor Results in Hematopoietic Defects inSteel17H Mice

Blood ◽  
1999 ◽  
Vol 94 (6) ◽  
pp. 1915-1925 ◽  
Author(s):  
Reuben Kapur ◽  
Ryan Cooper ◽  
Xingli Xiao ◽  
Mitchell J. Weiss ◽  
Peter Donovan ◽  
...  
Keyword(s):  
Amino Acids ◽  
Bone Marrow ◽  
Biological Activity ◽  
Stem Cell ◽  
Blood Cell ◽  
Red Blood Cell ◽  
Stem Cell Factor ◽  
Cytoplasmic Domain

Abstract Stem cell factor (SCF) is expressed as an integral membrane growth factor that may be differentially processed to produce predominantly soluble (S) (SCF248) or membrane-associated (MA) (SCF220) protein. A critical role for membrane presentation of SCF in the hematopoietic microenvironment (HM) has been suggested from the phenotype of the Steel-dickie(Sld) mice, which lack MA SCF, and by studies performed in our laboratory (and by others) using long-term bone marrow cultures and transgenic mice expressing different SCF isoforms.Steel17H (Sl17H) is an SCF mutant that demonstrates melanocyte defects and sterility in males but not in females. The Sl17H allele contains a intronic mutation resulting in the substitution of 36 amino acids (aa’s) in the SCF cytoplasmic domain with 28 novel aa’s. This mutation, which affects virtually the entire cytoplasmic domain of SCF, could be expected to alter membrane SCF presentation. To investigate this possibility, we examined the biochemical and biologic properties of the Sl17H-encoded protein and its impact in vivo and in vitro on hematopoiesis and on c-Kit signaling. We demonstrate that compound heterozygous Sl/Sl17H mice manifest multiple hematopoietic abnormalities in vivo, including red blood cell deficiency, bone marrow hypoplasia, and defective thymopoiesis. In vitro, both S and MA Sl17H isoforms of SCF exhibit reduced cell surface expression on stromal cells and diminished biological activity in comparison to wild-type (wt) SCF isoforms. These alterations in presentation and biological activity are associated with a significant reduction in the proliferation of an SCF-responsive erythroid progenitor cell line and in the activation of phosphatidylinositol 3-Kinase/Akt and mitogen-activated protein-Kinase signaling pathways. In vivo, transgene expression of the membrane-restricted (MR) (SCFX9/D3) SCF in Sl/Sl17H mutants results in a significant improvement in peripheral red blood cell counts in comparison toSl/Sl17H mice.

scholarly journals Retrovirus-induced feline pure red blood cell aplasia: pathogenesis and response to suramin

Blood ◽  
1991 ◽  
Vol 77 (7) ◽  
pp. 1442-1451
Author(s):  
JL Abkowitz
Keyword(s):  
Blood Cell ◽  
Red Blood Cell ◽  
Mononuclear Cells ◽  
Leukemia Virus ◽  
Transcriptase Inhibitor ◽  
Feline Leukemia Virus ◽  
Erythroid Progenitors ◽  
Marrow Cultures

Feline leukemia virus, subgroup C/Sarma (FeLV-C/Sarma) induces pure red blood cell aplasia in cats. Although erythroid (BFU-E and CFU-E) and granulocyte/macrophage (CFU-GM) progenitors are infected with this virus, only erythropoiesis is impaired. Two to 3 weeks before the onset of anemia, CFU-E become undetectable in marrow cultures while earlier erythroid progenitors (BFU-E) persist, suggesting that FeLV-C/Sarma (presumably via its envelope glycoprotein gp70) inhibits the differentiation of BFU-E to CFU-E in vivo. To correlate in vitro observations with the progression of disease, prospective studies were performed in six cats. These studies showed that at the time that the frequencies of CFU-E decreased in marrow cultures, BFU-E no longer responded to hematopoietic growth factor(s), although the responses of CFU-GM were unchanged. In further studies, anemic cats received suramin, a reverse-transcriptase inhibitor with other diverse effects. Within 4 to 14 days, erythropoiesis improved and up to 1,616 CFU-E were detected per 10(5) marrow mononuclear cells. However, progenitor cells remained infected, suggesting that suramin modulated erythroid differentiation without inhibiting progenitor infection. These observations led to the hypothesis that the gp70 of FeLV-C/Sarma impairs BFU-E differentiation by interference with ligand/receptor interactions or signal transduction pathways unique to erythroid cells. Understanding this mechanism should provide insights into the interactions controlling early erythropoiesis.

scholarly journals Interleukin-11 stimulates multiple phases of erythropoiesis in vitro

Blood ◽  
1992 ◽  
Vol 80 (5) ◽  
pp. 1218-1223
Author(s):  
VF Quesniaux ◽  
SC Clark ◽  
K Turner ◽  
B Fagg
Keyword(s):  
Bone Marrow ◽  
Erythroid Differentiation ◽  
Erythroid Progenitors ◽  
Murine Bone Marrow ◽  
Kit Ligand ◽  
B Cell Maturation ◽  
Stromal Cell Line ◽  
Interleukin 11

Interleukin-11 (IL-11), a pleiotropic cytokine originally isolated from a primate bone marrow stromal cell line, has been shown to stimulate T- cell-dependent B-cell maturation, megakaryopoiesis, and various stages of myeloid differentiation, but to inhibit adipogenesis. Because stromal cells are essential for the maintenance of early hematopoietic progenitor cells in long-term culture, we investigated the effects of IL-11 on multipotent and erythroid precursors from murine bone marrow in vitro in suspension and semisolid cultures. Our results show that in the presence of IL-3 or c-kit ligand (KL), IL-11 has profound stimulatory effects on primitive multilineage hematopoietic progenitors, pre-CFC(multi), as well as on precursors representing various stages of erythroid differentiation observable in vitro, including CFC(multi), BFU-E, and CFU-E. In addition, the combination of KL with IL-11 also stimulated highly proliferative erythroid progenitors that yield remarkable macroscopic erythroblast colonies in culture. These results indicate that IL-11 is likely to play a pivotal role in early hematopoiesis and at multiple stages of erythropoiesis.

scholarly journals Red Blood Cell Lysis Procedure Improves the Performances of ELN Dyserythropoiesis Score Compared to Nuclear Dye Use without Lysis

Blood ◽  
2018 ◽  
Vol 132 (Supplement 1) ◽  
pp. 3108-3108
Author(s):  
Hugo Germain ◽  
Sihem Tarfi ◽  
Nicolas Freynet ◽  
Andrea Toma ◽  
Bouchra Badaoui ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Flow Cytometry ◽  
Blood Cell ◽  
Red Blood Cell ◽  
Cell Lysis ◽  
Similar Amount ◽  
Erythroid Cells ◽  
Erythroid Progenitors ◽  
Specificity And Sensitivity ◽  
Control Samples

Abstract Introduction: The IMDSFlow working group reported recently the usefulness of immunophenotypic analysis of erythroid dysplasia in myelodysplastic syndromes (MDS) (Westers, 2017). This multicenter study revealed that analysis of CD36 and CD71 expression on nucleated erythroid cells as coefficient of variation (CV), in combination with CD71 fluorescence intensity and the percentage of CD117+ erythroid progenitors provided the best discrimination between MDS and non-clonal cytopenias. These four parameters were analyzed on bone marrow samples after red blood cell lysis (RBC) procedure. Since this latter could also remove some nucleated erythroid cells, it was proposed to use a nuclear dye in a whole no-lysis bone marrow strategy for analysis of CD36 and CD71 expression (Mathis, 2013). So far, no study has compared the four ELN dyserythropoiesis parameters after or without lysis procedure. Objective:We aimed to evaluate whether no-lysis procedure can lead to a better assessment of dyserythropoiesis by flow cytometry compared to lysis procedure by preserving erythroblast cells. Methods: One hundred patients referred to our laboratory for bone marrow investigations between November 2017 and June 2018 were included in this prospective study. Nineteen patients were used as control samples without cytopenia whereas 81 presented at least one cytopenia. Complete blood count parameters as well as morphological and cytogenetic analyses were used to classify these patients as 25 MDS or 56 non-MDS, referred thereafter as pathological controls. Bone marrow specimens were collected on EDTA and were processed within 4h following aspiration. A similar amount of each sample was stained in parallel with the same antibody panel (CD36, CD71, CD117 and CD45), yet according to two different protocols, one with CyTrak orange without lysis procedure and one with RBC lysis using VersaLyse (Beckman-Coulter). Data were acquired using a Navios cytometer and analyzed using the Kaluza software (BC). Geometrical means of fluorescence (GMFI) of CD71 as well as CD36 and CD71 coefficients of variation and the percentage of CD117+ erythroid progenitors were collected in order to calculate the ELN dyserythropoiesis score as previously described (see Figure 1 A-B for erythroblast selection). Results: Firstly, we compared the percentages of erythroblasts obtained with the no-lysis and the RBC lysis strategies as reported to the total nucleated cells analyzed (corresponding to the CD45 positive cells in addition to erythroblasts selected as CD71 and CD36 positive cells). Surprisingly, we found that the lysis protocol led to a higher percentage of erythroblasts than the no-lysis protocol (19.0±11.8% vs 15.4±10.8%; p<0.0001), yet both flow cytometry approaches gave lower percentages than morphology (30.4±12.5%; p<0.0001) as expected. Samples stained according to the two different protocols were analyzed with the same calibrated cytometer and the same settings for all markers (especially CD71 and CD36), allowing comparison of the raw data. The four parameters of the ELN score were significantly different between the two protocols for the 100 patients analyzed: both CV of CD36 and CD71 and % CD117 progenitors were higher with no-lysis protocol while GMFI CD71 was lower (Figure 1 C-F). We then calculated the ELN score with the recommended weighted manner: 4 points for increase in CD36 CV, 3 points for increase in CD71 CV, 2 points for decreased CD71 MFI and 2 points in the case of decreased or increased percentage of CD117+ erythroid cells. For each of the four parameters, cut-offs for the identification of MDS-associated aberrancies were defined against the 10th and/or 90th percentiles of the values obtained either from control samples (Figure 1 G) or from pathological controls (Figure 1 H) for both lysis and no-lysis protocols. With cut-offs defined against control samples, both specificity and sensitivity were better for lysis vs no-lysis protocol (93.3% vs 88.0% and 20.0% vs 12.0%, respectively). With cut-offs defined against pathological controls, again both specificity and sensitivity were higher for lysis vs no-lysis protocol (94.7% vs 88.0% and 20.0% vs 8.0%, respectively). Conclusion: Our results demonstrate that the lysis protocol leads to better specificity and sensitivity of the ELN dyserythropoiesis score than the no-lysis protocol with cut-offs defined against either control samples or pathological controls. Figure 1. Figure 1. Disclosures No relevant conflicts of interest to declare.

The Presence of Novel Amino Acids in the Cytoplasmic Domain of Stem Cell Factor Results in Hematopoietic Defects inSteel17H Mice

Blood ◽  
1999 ◽  
Vol 94 (6) ◽  
pp. 1915-1925 ◽  
Author(s):  
Reuben Kapur ◽  
Ryan Cooper ◽  
Xingli Xiao ◽  
Mitchell J. Weiss ◽  
Peter Donovan ◽  
...  
Keyword(s):  
Amino Acids ◽  
Bone Marrow ◽  
Biological Activity ◽  
Stem Cell ◽  
Blood Cell ◽  
Red Blood Cell ◽  
Stem Cell Factor ◽  
Cytoplasmic Domain

Stem cell factor (SCF) is expressed as an integral membrane growth factor that may be differentially processed to produce predominantly soluble (S) (SCF248) or membrane-associated (MA) (SCF220) protein. A critical role for membrane presentation of SCF in the hematopoietic microenvironment (HM) has been suggested from the phenotype of the Steel-dickie(Sld) mice, which lack MA SCF, and by studies performed in our laboratory (and by others) using long-term bone marrow cultures and transgenic mice expressing different SCF isoforms.Steel17H (Sl17H) is an SCF mutant that demonstrates melanocyte defects and sterility in males but not in females. The Sl17H allele contains a intronic mutation resulting in the substitution of 36 amino acids (aa’s) in the SCF cytoplasmic domain with 28 novel aa’s. This mutation, which affects virtually the entire cytoplasmic domain of SCF, could be expected to alter membrane SCF presentation. To investigate this possibility, we examined the biochemical and biologic properties of the Sl17H-encoded protein and its impact in vivo and in vitro on hematopoiesis and on c-Kit signaling. We demonstrate that compound heterozygous Sl/Sl17H mice manifest multiple hematopoietic abnormalities in vivo, including red blood cell deficiency, bone marrow hypoplasia, and defective thymopoiesis. In vitro, both S and MA Sl17H isoforms of SCF exhibit reduced cell surface expression on stromal cells and diminished biological activity in comparison to wild-type (wt) SCF isoforms. These alterations in presentation and biological activity are associated with a significant reduction in the proliferation of an SCF-responsive erythroid progenitor cell line and in the activation of phosphatidylinositol 3-Kinase/Akt and mitogen-activated protein-Kinase signaling pathways. In vivo, transgene expression of the membrane-restricted (MR) (SCFX9/D3) SCF in Sl/Sl17H mutants results in a significant improvement in peripheral red blood cell counts in comparison toSl/Sl17H mice.

Erythroid Development in the Absence of Hemoglobin.

Blood ◽  
2007 ◽  
Vol 110 (11) ◽  
pp. 1262-1262
Author(s):  
Shan-Run Liu ◽  
Sean C. McConnell ◽  
Yongliang Huo ◽  
Ting-Ting Zhang ◽  
Rui Yang ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Blood Cell ◽  
Es Cells ◽  
Embryonic Stem ◽  
Globin Genes ◽  
Erythroid Cells ◽  
Erythroid Progenitors ◽  
Erythroid Progenitor ◽  
Stage Of Development

Abstract The mammalian erythrocyte is a highly specialized blood cell that differentiates via an orderly series of committed progenitors in the bone marrow in a process termed erythropoiesis. Homeostasis of the erythron is carefully maintained by balancing the proliferation and destruction of early and late erythroid progenitors. In mature red blood cells over ninety-five percent of the protein is hemoglobin (Hb). What happens to committed erythroid cells in the absence of hemoglobin? To answer this question we have derived a novel line of embryonic stem (ES) cells from mouse embryos that have all eight adult alpha and beta globin genes knocked out. These “Null” Hb ES cells were injected into wild-type blastocysts to examine their in vivo potential to contribute to the tissues of developing chimeric mice. Examination of the peripheral blood and bone marrow of these chimeras by flow cytometry revealed that the “Null” Hb ES cells were able to produce normal levels of each type of white blood cell analyzed. However, “Null” erythrocytes were absent from the circulation and only early committed progenitors were found in the bone marrow. Very few “Null” erythroid cells matured beyond the proerythoblast to the basophilic erythroblast stage (Ter119low, CD71hi). To study this maturational block in more detail, an erythroid culture system was established by in vitro differentiation of the “Null” Hb ES cells. These pure erythroid progenitor (EP) cultures support and amplify the proerythroblast stage of development. Interestingly, EP cells could be derived from “Null” Hb ES cells demonstrating that Hb is not required for the development of proerythroblasts. “Null” derived EP cells express erythroid lineage markers (EKLF, GATA1, GypA, EpoR, Tal1), but express no adult globins or markers of other hematopoietic lineages (Mpl, GATA3, IL7R, PAX5, CEBPα, CD41b). Upon terminal differentiation most “Null” derived EP cells undergo apoptosis by 48 hours (7AAD−, Annexin V+) and are dead (7AAD+) by 72 hours. These “Null” Hb ES cells provide a novel experimental system to elucidate the role of hemoglobin during erythroid differentiation, maturation, and homeostasis.

scholarly journals Interleukin-11 stimulates multiple phases of erythropoiesis in vitro

Blood ◽  
1992 ◽  
Vol 80 (5) ◽  
pp. 1218-1223 ◽  
Author(s):  
VF Quesniaux ◽  
SC Clark ◽  
K Turner ◽  
B Fagg
Keyword(s):  
Bone Marrow ◽  
Erythroid Differentiation ◽  
Erythroid Progenitors ◽  
Murine Bone Marrow ◽  
Kit Ligand ◽  
B Cell Maturation ◽  
Stromal Cell Line ◽  
Interleukin 11

Abstract Interleukin-11 (IL-11), a pleiotropic cytokine originally isolated from a primate bone marrow stromal cell line, has been shown to stimulate T- cell-dependent B-cell maturation, megakaryopoiesis, and various stages of myeloid differentiation, but to inhibit adipogenesis. Because stromal cells are essential for the maintenance of early hematopoietic progenitor cells in long-term culture, we investigated the effects of IL-11 on multipotent and erythroid precursors from murine bone marrow in vitro in suspension and semisolid cultures. Our results show that in the presence of IL-3 or c-kit ligand (KL), IL-11 has profound stimulatory effects on primitive multilineage hematopoietic progenitors, pre-CFC(multi), as well as on precursors representing various stages of erythroid differentiation observable in vitro, including CFC(multi), BFU-E, and CFU-E. In addition, the combination of KL with IL-11 also stimulated highly proliferative erythroid progenitors that yield remarkable macroscopic erythroblast colonies in culture. These results indicate that IL-11 is likely to play a pivotal role in early hematopoiesis and at multiple stages of erythropoiesis.

The Role of Stathmin, a Regulator of Mitosis, in Hematopoiesis

Blood ◽  
2012 ◽  
Vol 120 (21) ◽  
pp. 3453-3453
Author(s):  
Charmaine A. Ramlogan-Steel ◽  
Jason C. Steel ◽  
Hassana Fathallah ◽  
Camelia Iancu-Rubin ◽  
Manoocher Soleimani ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Blood Cell ◽  
Red Blood Cell ◽  
Megaloblastic Anemia ◽  
Platelet Production ◽  
Cell Counts ◽  
Blood Cell Counts ◽  
The Mean

Abstract Abstract 3453 Introduction Stathmin is a 17KDa cytosolic protein that plays an important role in the regulation of microtubule dynamics, mitotic spindle formation, cell cycle progression and cell differentiation. Stathmin knockout (KO) mice were initially reported to have a normal phenotype but were subsequently shown to develop an age-related neurological phenotype with axonopathy evident in both central and peripheral nervous systems. These mice were also shown to have a defect in recovery from acute ischemic renal injury. We had previously shown that stathmin plays an important role in the differentiation and proliferation of megakaryocytes (MK) and that down-regulation of stathmin is necessary for the maturation of MK and platelet production in vitro. In this study, we investigated the role of stathmin in megakaryopoiesis and hematopoiesis in vivo using the stathmin KO mouse as an experimental model. Results Stathmin KO mice had lower platelet (PLT) counts at 3 weeks of age when compared to WT mice. The WT mice had a mean PLT count of 662 ± 27 K/μL while KO mice had a mean PLT count of 543 ± 37 K/μL. This correlated with larger and fewer MK in the bone marrow of KO mice (WT: 4.2 ± 0.7 MK/40X field; KO: 3.6 ± 0.2 MK/40X field). Furthermore, in the spleen, there was a 10 fold decrease in the number of MK in KO mice compared to WT mice (6.6 ± 0.6 vs 0.7 ± 0.1 MK/40X field). By 8 weeks, PLT counts and MK size and numbers in the bone marrow and spleen were similar in WT and KO mice. Interestingly, by 16 weeks, the mean PLT of KO mice became significantly higher than that of WT and by 40 weeks, the mean PLT count of KO mice was 1379 ± 100K/μL compared to 1045 ± 120K/μL in WT mice (P<0.05). Microscopic analysis of the bone marrow at 46 weeks of age showed approximately 50% more MK in KO mice compared to WT mice. Differences in red blood cell counts (RBC) were also observed. While at 3 weeks, there were no significant differences between the 2 groups, at 8 weeks, KO mice had significantly lower RBC counts, hemoglobin levels (Hb) and hematocrit (HCT). This trend continued until the last measurement recorded at 40 weeks. Mean RBC in WT mice was 10.5 ± 0.1M/μL compared to 8.9 ± 0.2M/μL in KO mice. The mean corpuscular volume (MCV) and the red blood cell distribution width (RDW) were consistently higher in KO mice than in WT mice. No significant differences were noted in white blood cell counts. Bone marrow cell counts were significantly lower in KO mice when compared to WT mice at different ages from 3–40 weeks. Progenitor cell assays from 10–12 week old animals have shown that bone marrow from KO mice produce significantly fewer BFU-E and Pre-B colonies while no differences were observed in CFU-GMs. Conclusions The phenotypic characteristics of stathmin KO mice confirmed our prior in vitro findings that suggested a role for stathmin in megakaryopoiesis. We expected to see a decrease in the number of platelets and MK coupled with an increase in MK size. This was confirmed in stathmin KO mice at 3 weeks of age. However, we did not expect to see the marked increase in the number of platelets and MK that was observed as the mice aged. The exact mechanism for this has not been identified. Interestingly, the stathmin KO mice exhibited characteristic features of megaloblastic anemia including mild anemia and a significant increase in MCV and RDW. The megaloblastic anemia that is seen in the presence of B12 and folate deficiency results from interference with DNA synthesis resulting in asynchronous maturation of the nucleus and the cytoplasm. We believe a similar phenomenon is occurring in the stathmin KO mice. The deficiency of stathmin results in aberrant exit from mitosis, thereby delaying nuclear maturation and resulting in the megaloblastic features. Thus, the deficiency of stathmin in the KO mice results in two hematopoietic phenotypes that are seen in humans, megaloblastic anemia and thrombocytosis. It is unclear whether mutations of stathmin in humans might result in similar phenotypes. This is a question that will require further investigation. Future studies will investigate the compensatory mechanisms that result in the switch from decreased to increased platelet production as the mice age. Furthermore, examining the effects of hematopoietic stress (e.g. response to chemotherapy or bleeding) in stathmin KO mice might also elucidate a role for stathmin in the recovery from hematopoietic injury as was seen in acute ischemic renal injury. Disclosures: No relevant conflicts of interest to declare.

scholarly journals Retrovirus-induced feline pure red blood cell aplasia: pathogenesis and response to suramin

Blood ◽  
1991 ◽  
Vol 77 (7) ◽  
pp. 1442-1451 ◽  
Author(s):  
JL Abkowitz
Keyword(s):  
Blood Cell ◽  
Red Blood Cell ◽  
Mononuclear Cells ◽  
Leukemia Virus ◽  
Transcriptase Inhibitor ◽  
Feline Leukemia Virus ◽  
Erythroid Progenitors ◽  
Marrow Cultures

Abstract Feline leukemia virus, subgroup C/Sarma (FeLV-C/Sarma) induces pure red blood cell aplasia in cats. Although erythroid (BFU-E and CFU-E) and granulocyte/macrophage (CFU-GM) progenitors are infected with this virus, only erythropoiesis is impaired. Two to 3 weeks before the onset of anemia, CFU-E become undetectable in marrow cultures while earlier erythroid progenitors (BFU-E) persist, suggesting that FeLV-C/Sarma (presumably via its envelope glycoprotein gp70) inhibits the differentiation of BFU-E to CFU-E in vivo. To correlate in vitro observations with the progression of disease, prospective studies were performed in six cats. These studies showed that at the time that the frequencies of CFU-E decreased in marrow cultures, BFU-E no longer responded to hematopoietic growth factor(s), although the responses of CFU-GM were unchanged. In further studies, anemic cats received suramin, a reverse-transcriptase inhibitor with other diverse effects. Within 4 to 14 days, erythropoiesis improved and up to 1,616 CFU-E were detected per 10(5) marrow mononuclear cells. However, progenitor cells remained infected, suggesting that suramin modulated erythroid differentiation without inhibiting progenitor infection. These observations led to the hypothesis that the gp70 of FeLV-C/Sarma impairs BFU-E differentiation by interference with ligand/receptor interactions or signal transduction pathways unique to erythroid cells. Understanding this mechanism should provide insights into the interactions controlling early erythropoiesis.

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