scholarly journals Serum‐free suspension cultured human cells can produce a high‐level of recombinant human erythropoietin

2020 ◽  
Vol 2 (6) ◽  
Author(s):  
Luciano Costa e Silva ◽  
Matheus Henrique Santos ◽  
Tarik Reis Heluy ◽  
Rafael Tagé Biaggio ◽  
Alexander Rodrigo Ferreira ◽  
...  
Keyword(s):  
Recombinant Human Erythropoietin ◽  
Human Cells ◽  
Serum Free ◽  
Human Erythropoietin ◽  
Cultured Human Cells ◽  
High Level ◽  
Serum Free Suspension

scholarly journals Induction of mixed erythroid-megakaryocyte colonies and bipotential blast cell colonies by recombinant human erythropoietin in serum-free culture

Blood ◽  
1990 ◽  
Vol 76 (7) ◽  
pp. 1330-1335
Author(s):  
N Nishi ◽  
T Nakahata ◽  
K Koike ◽  
M Takagi ◽  
K Naganuma ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Recombinant Human Erythropoietin ◽  
Bone Marrow Cells ◽  
Blast Cell ◽  
Mouse Bone Marrow ◽  
High Concentration ◽  
Erythroid Progenitors ◽  
Serum Free ◽  
Human Erythropoietin ◽  
Marrow Cells

The effects of recombinant human erythropoietin (rEp) on murine hematopoietic progenitors were studied using a serum-free culture. A high concentration of rEp stimulated the formation of mixed erythroid- megakaryocyte colonies (EM colonies) and blast cell colonies, as well as erythroid colonies, erythroid bursts, and megakaryocyte colonies from normal mouse bone marrow cells. Direct effects of rEp on EM colony, megakaryocyte colony, and erythroid burst formation were confirmed by depletion of accessory cells such as T cells, B cells, and macrophages from crude bone marrow cells, and inhibition of the colonies by the addition of rabbit anti-rEp antibody to the culture in a dose-dependent fashion. Replating experiments were performed to confirm the differentiating ability of blast cell colonies grown in the presence of rEp. Most of the blast cell colonies yielded not only secondary erythroid colonies but also megakaryocyte colonies in the presence of 2 IU/mL rEp. Some of the blast cell colonies produced secondary EM colonies in the presence of 16 IU/ml rEp of 2 IU/mL rEp plus interleukin-3, although no granulocyte-macrophage colonies were found in the secondary culture. These results suggest that Ep acts not only as a late-acting factor that is specific for erythroid progenitors, but also as a bipotential EM-stimulating factor for murine hematopoietic cells.

scholarly journals Induction of mixed erythroid-megakaryocyte colonies and bipotential blast cell colonies by recombinant human erythropoietin in serum-free culture

Blood ◽  
1990 ◽  
Vol 76 (7) ◽  
pp. 1330-1335 ◽  
Author(s):  
N Nishi ◽  
T Nakahata ◽  
K Koike ◽  
M Takagi ◽  
K Naganuma ◽  
...  
Keyword(s):  
Bone Marrow ◽  
Recombinant Human Erythropoietin ◽  
Bone Marrow Cells ◽  
Blast Cell ◽  
Mouse Bone Marrow ◽  
High Concentration ◽  
Erythroid Progenitors ◽  
Serum Free ◽  
Human Erythropoietin ◽  
Marrow Cells

Abstract The effects of recombinant human erythropoietin (rEp) on murine hematopoietic progenitors were studied using a serum-free culture. A high concentration of rEp stimulated the formation of mixed erythroid- megakaryocyte colonies (EM colonies) and blast cell colonies, as well as erythroid colonies, erythroid bursts, and megakaryocyte colonies from normal mouse bone marrow cells. Direct effects of rEp on EM colony, megakaryocyte colony, and erythroid burst formation were confirmed by depletion of accessory cells such as T cells, B cells, and macrophages from crude bone marrow cells, and inhibition of the colonies by the addition of rabbit anti-rEp antibody to the culture in a dose-dependent fashion. Replating experiments were performed to confirm the differentiating ability of blast cell colonies grown in the presence of rEp. Most of the blast cell colonies yielded not only secondary erythroid colonies but also megakaryocyte colonies in the presence of 2 IU/mL rEp. Some of the blast cell colonies produced secondary EM colonies in the presence of 16 IU/ml rEp of 2 IU/mL rEp plus interleukin-3, although no granulocyte-macrophage colonies were found in the secondary culture. These results suggest that Ep acts not only as a late-acting factor that is specific for erythroid progenitors, but also as a bipotential EM-stimulating factor for murine hematopoietic cells.

scholarly journals High-level expression of secreted complex glycosylated recombinant human erythropoietin in the Physcomitrella ?-fuc-t ?-xyl-t mutant

2007 ◽  
Vol 5 (3) ◽  
pp. 389-401 ◽  
Author(s):  
Andreas Weise ◽  
Friedrich Altmann ◽  
Marta Rodriguez-Franco ◽  
Eric R. Sjoberg ◽  
Wolfgang Bäumer ◽  
...  
Keyword(s):  
Recombinant Human Erythropoietin ◽  
High Level Expression ◽  
Human Erythropoietin ◽  
High Level

Serum-free suspension culturing of human cells: adaptation, growth, and cryopreservation

2015 ◽  
Vol 38 (8) ◽  
pp. 1495-1507 ◽  
Author(s):  
Rafael Tagé Biaggio ◽  
Mário Soares Abreu-Neto ◽  
Dimas Tadeu Covas ◽  
Kamilla Swiech
Keyword(s):  
Human Cells ◽  
Serum Free ◽  
Serum Free Suspension

scholarly journals High-level expression and purification of a recombinant human erythropoietin produced using a baculovirus vector

Blood ◽  
1989 ◽  
Vol 74 (2) ◽  
pp. 652-657
Author(s):  
FW Quelle ◽  
LF Caslake ◽  
RE Burkert ◽  
DM Wojchowski
Keyword(s):  
Mammalian Cells ◽  
Recombinant Human Erythropoietin ◽  
Insect Cell Line ◽  
Recombinant Baculovirus ◽  
Cell Surface Receptor ◽  
High Level Expression ◽  
Human Erythropoietin ◽  
Highly Active ◽  
High Level

Conditions presently have been established for the high-level expression and simplified purification of recombinant human erythropoietin produced in Spodoptera frugiperda cells. Expression, as mediated by infection with a recombinant baculovirus, was accomplished in suspension culture using reduced levels of serum and media supplements experimentally determined to provide optimum levels of factor production (500,000 U/L). Purification of this recombinant human erythropoietin to virtual homogeneity (greater than or equal to 99%) was accomplished via a simple three-step procedure involving isocratic elution from DEAE-Sephacel, reverse-phase high performance liquid chromatography (HPLC) on a C4 medium, and the single-step elution of purified hormone from concanavalin A agarose. Overall, an 890-fold purification was accomplished with a recovery of 80% as assayed in vitro. Biologically, this purified erythropoietin is highly active, possessing a specific activity in vitro of 200,000 U/mg protein. Chemically, this erythropoietin (molecular weight [mol wt] 26,200) appears exceptionally uniform in its oligosaccharide constitution (30%) as contrasted with heterogeneously glycosylated erythropoietins derived from mammalian cells (mol wt 30,000 to 38,000; 40% to 50% complex-type oligosaccharide). Thus, human erythropoietin as presently produced in an insect cell line comprises not only an abundant source of highly active, readily purified hormone for studies of its mechanism of action and cell surface receptor, but also represents a uniquely homogeneous form that should prove advantageous for direct structural analyses.

scholarly journals High-level expression and purification of a recombinant human erythropoietin produced using a baculovirus vector

Blood ◽  
1989 ◽  
Vol 74 (2) ◽  
pp. 652-657 ◽  
Author(s):  
FW Quelle ◽  
LF Caslake ◽  
RE Burkert ◽  
DM Wojchowski
Keyword(s):  
Mammalian Cells ◽  
Recombinant Human Erythropoietin ◽  
Insect Cell Line ◽  
Recombinant Baculovirus ◽  
Cell Surface Receptor ◽  
High Level Expression ◽  
Human Erythropoietin ◽  
Highly Active ◽  
High Level

Abstract Conditions presently have been established for the high-level expression and simplified purification of recombinant human erythropoietin produced in Spodoptera frugiperda cells. Expression, as mediated by infection with a recombinant baculovirus, was accomplished in suspension culture using reduced levels of serum and media supplements experimentally determined to provide optimum levels of factor production (500,000 U/L). Purification of this recombinant human erythropoietin to virtual homogeneity (greater than or equal to 99%) was accomplished via a simple three-step procedure involving isocratic elution from DEAE-Sephacel, reverse-phase high performance liquid chromatography (HPLC) on a C4 medium, and the single-step elution of purified hormone from concanavalin A agarose. Overall, an 890-fold purification was accomplished with a recovery of 80% as assayed in vitro. Biologically, this purified erythropoietin is highly active, possessing a specific activity in vitro of 200,000 U/mg protein. Chemically, this erythropoietin (molecular weight [mol wt] 26,200) appears exceptionally uniform in its oligosaccharide constitution (30%) as contrasted with heterogeneously glycosylated erythropoietins derived from mammalian cells (mol wt 30,000 to 38,000; 40% to 50% complex-type oligosaccharide). Thus, human erythropoietin as presently produced in an insect cell line comprises not only an abundant source of highly active, readily purified hormone for studies of its mechanism of action and cell surface receptor, but also represents a uniquely homogeneous form that should prove advantageous for direct structural analyses.

Alpha-Tocopherol Protects Cultured Human Cells from the Acute Lethal Cytotoxicity of Dioxin

2002 ◽  
Vol 72 (3) ◽  
pp. 147-153 ◽  
Author(s):  
Kei-Ichi Hirai ◽  
Jie-Hong Pan ◽  
Ying-Bo Shui ◽  
Eriko Simamura ◽  
Hiroki Shimada ◽  
...  
Keyword(s):  
Cell Death ◽  
Cell Damage ◽  
Smooth Endoplasmic Reticulum ◽  
Dehydroascorbic Acid ◽  
Human Cells ◽  
Alpha Tocopherol ◽  
Cervical Cancer Cells ◽  
Cultured Human Cells ◽  
Nuclear Damage ◽  
Conjunctival Epithelial Cells

The possible protection of cultured human cells from acute dioxin injury by antioxidants was investigated. The most potent dioxin, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), caused vacuolization of the smooth endoplasmic reticulum and Golgi apparatus in cultured human conjunctival epithelial cells and cervical cancer cells. Subsequent nuclear damage included a deep irregular indentation resulting in cell death. A dosage of 30–40 ng/mL TCDD induced maximal intracellular production of H2O2 at 30 minutes and led to severe cell death (0–31% survival) at two hours. A dose of 1.7 mM alpha-tocopherol or 1 mM L-dehydroascorbic acid significantly protected human cells against acute TCDD injuries (78–97% survivals), but vitamin C did not provide this protection. These results indicate that accidental exposure to fatal doses of TCDD causes cytoplasmic free radical production within the smooth endoplasmic reticular systems, resulting in severe cytotoxicity, and that vitamin E and dehydroascorbic acid can protect against TCDD-induced cell damage.

Sign in / Sign up

Export Citation Format

Share Document