In vitro and in vivo Effects of Recombinant Human Erythropoietin on Human Hemopoietic Progenitor Cells

Anemia is a severe complication in patients with chronic kidney disease (CKD). Treatment with exogenous erythropoietin (EPO) can correct anemia in many with CKD. We produced 5/6-nephrectomized rats that became uremic and anemic at 25 days post surgery. Injection of the anemic 5/6-nephrectomized rats with 2.8 mg zinc/kg body weight raised their red blood cell (RBC) levels from approximately 85% of the control to 95% in one day and continued for 4 days. We compared the effect of ZnSO4 and recombinant human erythropoietin (rHuEPO) injections on relieving anemia in 5/6-nephrectomized rats. After three consecutive injections, both the ZnSO4 and rHuEPO groups had significantly higher RBC levels (98 ± 6% and 102 ± 6% of the control) than the saline group (90 ± 3% of the control). In vivo, zinc relieved anemia in 5/6-nephrectomized rats similar to rHuEPO. In vitro, we cultured rat bone marrow cells supplemented with ZnCl2, rHuEPO, or saline. In a 4-day suspension culture, we found that zinc induced erythropoiesis similar to rHuEPO. When rat bone marrow cells were supplement-cultured with zinc, we found that zinc stimulated the production of EPO in the culture medium and that the level of EPO produced was dependent on the concentration of zinc supplemented. The production of EPO via zinc supplementation was involved in the process of erythropoiesis.

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In vivo Stimulatory Effects of Neopterin on Hematopoiesis

Pteridines ◽

10.1515/pteridines.1998.9.1.13 ◽

1998 ◽

Vol 9 (1) ◽

pp. 13-17 ◽

Cited By ~ 5

Author(s):

Shin Aizawa ◽

Masaki Hiramoto ◽

Sonoko Araki ◽

Hajime Hosh ◽

Shuji Kojima ◽

...

Keyword(s):

Bone Marrow ◽

Progenitor Cells ◽

Intraperitoneal Administration ◽

Extensive Study ◽

Blood Leukocytes ◽

Proliferation And Differentiation ◽

Blood Leukocyte ◽

Hemopoietic Progenitor ◽

Hemopoietic Progenitor Cells

Summary The pteridine neopterin (NP) was shown to be produced by monocytes and has been known as a useful maker of immunological activation, although, the biological activity of this agent is still unclear. To elucidate the biological function of NP, the changes in the number of blood leukocyte, bone marrow and spleen hemopoietic progenitor cells after intraperitoneal administration of NP into mice were investigated. Administration of NP increased the number of blood leukocytes about 2 fold higher than that of the control at days 7 and 14. Blood films, made by smearing samples of peripheral blood, showed the increment of granulocytes in blood of NP treated animals. The number of granulocyte-macrophage progenitor cells (CFU-GM) was also increased in both bone marrow and spleen in mice with NP administration. Extensive study showed that NP stimulated the hematological recovery in bone marrow transplanted animals. All these findings suggest that NP has a stimulating activity on hemopoiesis by affecting the proliferation and differentiation of hemopoietic progenitor cells in vivo.

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Effect of human recombinant erythropoietin on human hemopoietic progenitor cells in vitro

Klinische Wochenschrift ◽

10.1007/bf01748162 ◽

1988 ◽

Vol 66 (6) ◽

pp. 236-240 ◽

Cited By ~ 8

Author(s):

A. Ganser ◽

B. Völkers ◽

P. Scigalla ◽

D. Hoelzer

Keyword(s):

Progenitor Cells ◽

Recombinant Erythropoietin ◽

Human Recombinant Erythropoietin ◽

Hemopoietic Progenitor ◽

Hemopoietic Progenitor Cells

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Recombinant Human Erythropoietin: A Multipotential Hemopoietic Growth Factor in vivo and in vitro

Contributions to Nephrology - Erythropoietin in the 90s ◽

10.1159/000419472 ◽

2015 ◽

pp. 1-10 ◽

Cited By ~ 1

Author(s):

K. Geissler ◽

F. Stockenhuber ◽

W. Hinterberger ◽

B. Balcke ◽

K. Lechner

Keyword(s):

Growth Factor ◽

Recombinant Human Erythropoietin ◽

Human Erythropoietin ◽

Hemopoietic Growth Factor

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Factor-independent erythropoietic progenitor cells in leukemia induced by the myeloproliferative leukemia virus

Blood ◽

10.1182/blood.v73.5.1161.1161 ◽

1989 ◽

Vol 73 (5) ◽

pp. 1161-1167 ◽

Cited By ~ 15

Author(s):

F Wendling ◽

JF Penciolelli ◽

M Charon ◽

P Tambourin

Keyword(s):

Progenitor Cells ◽

Leukemia Virus ◽

Erythroid Progenitor ◽

Adult Mice ◽

Cell Compartments ◽

Erythroid Progenitor Cells ◽

Semi Solid ◽

In Vivo Effects

Abstract The myeloproliferative leukemia virus (MPLV), a novel murine retroviral complex that does not transform fibroblasts, has been shown to cause an acute leukemia in adult mice accompanied by a progressive polycythemia. The present study demonstrates that, on in vivo inoculation, MPLV induces a rapid suppression of growth factor requirement for in vitro colony formation by both the late and the primitive erythroid progenitor cells. CFU-e-derived erythrocytic colonies developed and differentiated in semi-solid medium without the addition of erythropoietin (Epo). In addition, the formation of CFU-e colonies was not altered by the presence of specific neutralizing Epo antibodies. In the spleen, the CFU-e pool size increased rapidly up to 30-fold. By day 6 postinfection, 100% of these progenitor cells were Epo-independent. The in vivo effects of MPLV-infection on early erythroid progenitor cell compartments were examined in cultures grown for seven days. The concentration of erythroid progenitor cells was twofold elevated in spleen from MPLV-infected mice. As early as day 4 postinfection, 50% of these progenitors produced fully hemoglobinized colonies in serum-free cultures without the addition of interleukin-3 (IL-3) and Epo. Most spontaneous colonies were large and contained up to 10(5) cells per colony. They were composed of either erythroblasts only (16%) or erythroblasts and megakaryocytes (70%); few of them were multipotential (14%). In the marrow, the total number of BFU-e was reduced and only few factor-independent bursts were observed, suggesting a rapid migration of infected progenitors from marrow to spleen. Furthermore, the data show that abnormal erythropoiesis was due to the replication defective MPLV information and was not influenced by the Fv-2 locus.

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Antibody stimulation of hemopoietic progenitor cells

Blood ◽

10.1182/blood.v65.4.869.869 ◽

1985 ◽

Vol 65 (4) ◽

pp. 869-876 ◽

Cited By ~ 4

Author(s):

RM Crapper ◽

JW Schrader

Keyword(s):

Bone Marrow ◽

Progenitor Cells ◽

Bone Marrow Cells ◽

Exchange Chromatography ◽

Staphylococcal Protein A ◽

Igg Antibodies ◽

Hemopoietic Progenitor ◽

Hemopoietic Progenitor Cells ◽

Stimulation Of

Abstract Antisera were raised by immunizing rabbits with cloned lines of murine hemopoietic progenitor cells (P cells) that depended on the presence of a specific hemopoietic growth factor, persisting cell-stimulating factor (PSF), for their growth and survival. The unabsorbed antiserum was inhibitory, but after absorption with murine spleen cells and the mastocytoma, P815, significant stimulation of both P cell growth and thymidine incorporation was evident. IgG antibodies isolated from the antiserum by staphylococcal protein A chromatography or further purified by diethylaminoethyl anion exchange chromatography, ammonium sulphate precipitation, and gel filtration using Sephacryl S-300 were responsible for the stimulation. The absorbed antiserum promoted the survival of normal murine bone marrow cells in liquid culture over a four-day period, and the inclusion of IgG antibodies in agar cultures of normal bone marrow promoted the in vitro survival, over a 48-hour period, of cells capable of subsequently generating, in the presence of a source of PSF, colonies of neutrophils, macrophages, and megakaryocytes. It is postulated that the antibodies act by stimulating the PSF receptor on both the factor-dependent cell lines and normal myeloid progenitor cells.

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Near-optimal Conditions for the In Vitro Culture of Hemopoietic Progenitor Cells in Bone Marrow from the Rat

Toxicologic Pathology ◽

10.1177/0192623308328133 ◽

2009 ◽

Vol 37 (2) ◽

pp. 170-174 ◽

Cited By ~ 1

Author(s):

Gemma Molyneux ◽

Sian Rizzo ◽

John Turton ◽

Parvinder Phul ◽

Frances Gibson

Keyword(s):

Bone Marrow ◽

In Vitro Culture ◽

Progenitor Cells ◽

Optimal Conditions ◽

Hemopoietic Progenitor ◽

Hemopoietic Progenitor Cells

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225 RECOMBINANT HUMAN ERYTHROPOIETIN PRODUCED IN THE MILK OF TRANSGENIC MICE: FUNCTIONAL CHARACTERIZATION AND PHARMACEUTICAL APPROACH

Reproduction Fertility and Development ◽

10.1071/rdv17n2ab225 ◽

2005 ◽

Vol 17 (2) ◽

pp. 263

Author(s):

D.N. Kwon ◽

J.Y. Park ◽

S.Y. Lee ◽

S.J. Kang ◽

J.H. Kim

Keyword(s):

Mammary Gland ◽

Transgenic Mouse ◽

Cho Cells ◽

Recombinant Human Erythropoietin ◽

Mouse Mammary Gland ◽

Human Erythropoietin ◽

Mouse Milk ◽

Bioreactor System

The proper post-transcriptional modification of recombinant human erythropoietin (rhEPO) is critical to retain its biological functions, either in vivo or in vitro. The major glycosyltransferases for the determinant of glycosylation patterns of rhEPO are N-acetylglycosaminyltransferase (GnT) and α-1-3/4 fucosyltransferase (Fut). GnT-III expression (388 ± 19.09) in the mouse mammary gland has been shown to be dramatically different from that in CHO cells, although FuT-VIII expression in CHO cells (1970 ± 255.9) is comparable to mouse mammary gland (272 ± 14.8), suggesting that the mammary gland may proceed with the proper glycosylation of rhEPO as shown in CHO cells. To identify this hypothesis and establish the rhEPO bioreactor system for mass production of protein in transgenic animals, we have generated two transgenic mouse lines that express rhEPO in milk. Both lines of transgenic mouse express only rhEPO in the lactating mammary gland, and the protein yield of rhEPO in lactating milk is comparable to that in CHO cells. After determining the protein expression in lactating milk, using three different methods – enzymatic release of oligosaccharide analysis, two-dimensional electrophoresis, and 2-aminobenzamide-labeled analysis – we report that the rhEPO produced by the animal bioreactor system has the proper glycosylation patterns as shown in CHO cell-derived Epoietin α, and has more tetra-acidic oligosaccharide structures than Epoietin α, which is the widely used rhEPO for therapeutic purposes. The in vitro biological property of transgenic mouse milk-derived rhEPO has been tested by measuring luciferase activity in MCF-7 cells, indicating that rhEPO from mammary gland up-regulates the EPO-receptor-mediated STAT5 gene expression in a dose-dependent manner the same as Epoietin α does. In addition, in vivo biological activity demonstrated that direct injection of rhEPO into a mouse vein increases blood components such as RBC and HCT. In light of these findings, we suggest that high levels of tetra-acidic structures observed in transgenic mouse milk-derived rhEPO may be related to the high level of expression of glycosiltransferases (GnT-III and FuT-VIII) in mammary gland; thus the bioreactor system using the mammary gland of a transgenic animal could be a good candidate for production of rhEPO for pharmaceutical purposes. This work was supported in part by a grant program from RDA(Biogreen21) and Cho-A, Republic of Korea. D.N. Kwon is the recipient of a scholarship from the BK21 program, granted by the Ministry of Education, Korea.

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