Recombinant human GM-CSF induces leukocytosis and activates peripheral blood polymorphonuclear neutrophils in nonhuman primates

Abstract The in vivo efficacy of glycosylated and nonglycosylated recombinant human granulocyte macrophage colony-stimulating factor (rh GM-CSF) expressed in Chinese hamster ovary cells and Escherichia coli respectively was studied in rhesus monkeys following a daily subcutaneous (SC; three times) or intravenous (IV; over six hours) dose for seven consecutive days. The monkeys responded to the rh GM-CSF with a prompt (within 24 hours) rise in circulating white blood cells (WBCs). Thereafter the total cell counts increased steadily in a dose- dependent manner with repeated dosing to numbers six times over the pretreatment levels. Overall, granulocyte counts increased fivefold, lymphocytes twofold to fourfold, and monocytes threefold to fourfold. Platelets and erythrocytes were unaffected. Within 1 week after the end of treatment the leukocytosis had disappeared. Of the two routes of treatment, SC (three times daily)-administered rh GM-CSF was more effective than the same dose given by a six-hour IV infusion. In addition to inducing leukocytosis, parenterally administered rh GM-CSF primed mature circulating granulocytes for enhanced oxidative metabolism and killing of an E coli strain. These results show that exogenously administered glycosylated or nonglycosylated rh GM-CSF is both an effective stimulator of leukocytosis and a potent activator of the phagocytic function of mature granulocytes in monkeys.

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Antithrombotic effect of recombinant human thrombomodulin on thrombin- induced thromboembolism in mice

Blood ◽

10.1182/blood.v75.7.1396.1396 ◽

1990 ◽

Vol 75 (7) ◽

pp. 1396-1399 ◽

Cited By ~ 98

Author(s):

K Gomi ◽

M Zushi ◽

G Honda ◽

S Kawahara ◽

O Matsuzaki ◽

...

Keyword(s):

Chinese Hamster Ovary Cells ◽

Chinese Hamster ◽

Lethal Effect ◽

Dependent Manner ◽

Antithrombotic Effect ◽

Concentration Dependent Manner ◽

Ovary Cells ◽

Recombinant Thrombomodulin

Abstract Antithrombotic effect of recombinant human thrombomodulin in mice, both in vitro and in vivo, was studied. The soluble recombinant human thrombomodulin was expressed in Chinese hamster ovary cells and purified from the conditioned medium by a modification of the conventional method. Recombinant thrombomodulin prolonged thrombin clotting time for mouse plasma in a dose-dependent manner. Thrombin was injected into the lateral tail vein of mice and caused acute thromboembolism. All mice injected with thrombin died of thromboembolism; however, preinjection with recombinant human thrombomodulin neutralized the lethal effect of thrombin in a concentration-dependent manner. Histologic examination showed that fibrin deposits were found in all large and small arteries in the lung from mice injected with thrombin; however, fibrin deposits were not detected in any large arteries from the mouse preinjected with thrombomodulin.

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Sialyltransferase specificity in selectin ligand formation

Blood ◽

10.1182/blood-2002-04-1007 ◽

2002 ◽

Vol 100 (10) ◽

pp. 3618-3625 ◽

Cited By ~ 101

Author(s):

Lesley G. Ellies ◽

Markus Sperandio ◽

Gregory H. Underhill ◽

James Yousif ◽

Michael Smith ◽

...

Keyword(s):

Chinese Hamster Ovary Cells ◽

Chinese Hamster ◽

Dependent Manner ◽

Neuraminidase Treatment ◽

Ovary Cells ◽

Rolling Velocity ◽

Selectin Ligands ◽

Selectin Ligand

Selectin ligands are glycan structures that participate in leukocyte trafficking and inflammation. At least 6 ST3Gal sialyltransferases (I-VI) have been identified that may contribute to selectin ligand formation. However, it is not known which of these sialyltransferases are involved in vivo and whether they may differentially regulate selectin function. We have produced and characterized mice genetically deficient in ST3Gal-I, ST3Gal-II, ST3Gal-III, and ST3Gal-IV. Unlike mice bearing severe defects in selectin ligand formation, there was no finding of leukocytosis with these single ST3Gal deficiencies. Among neutrophils, only ST3Gal-IV was found to play a role in the synthesis of selectin ligands. In vitro rolling of marrow-derived neutrophils on E- or P-selectins presented by Chinese hamster ovary cells was reduced in the absence of ST3Gal-IV. However, in a tumor necrosis factor α (TNF-α)–induced inflammation model in vivo, no defect among P-selectin ligands was observed. Nevertheless, the number of leukocytes rolling on postcapillary venules in an E-selectin–dependent manner was decreased while E-selectin–dependent rolling velocity was increased. We propose that multiple ST3Gal sialyltransferases contribute to selectin ligand formation, as none of these ST3Gal deficiencies recapitulated the degree of E- and P-selectin ligand deficit observed on neuraminidase treatment of intact neutrophils. Our findings indicate a high degree of functional specificity among sialyltransferases and a substantial role for ST3Gal-IV in selectin ligand formation.

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Antithrombotic effect of recombinant human thrombomodulin on thrombin- induced thromboembolism in mice

Blood ◽

10.1182/blood.v75.7.1396.bloodjournal7571396 ◽

1990 ◽

Vol 75 (7) ◽

pp. 1396-1399 ◽

Cited By ~ 2

Author(s):

K Gomi ◽

M Zushi ◽

G Honda ◽

S Kawahara ◽

O Matsuzaki ◽

...

Keyword(s):

Chinese Hamster Ovary Cells ◽

Chinese Hamster ◽

Lethal Effect ◽

Dependent Manner ◽

Antithrombotic Effect ◽

Concentration Dependent Manner ◽

Ovary Cells ◽

Recombinant Thrombomodulin

Antithrombotic effect of recombinant human thrombomodulin in mice, both in vitro and in vivo, was studied. The soluble recombinant human thrombomodulin was expressed in Chinese hamster ovary cells and purified from the conditioned medium by a modification of the conventional method. Recombinant thrombomodulin prolonged thrombin clotting time for mouse plasma in a dose-dependent manner. Thrombin was injected into the lateral tail vein of mice and caused acute thromboembolism. All mice injected with thrombin died of thromboembolism; however, preinjection with recombinant human thrombomodulin neutralized the lethal effect of thrombin in a concentration-dependent manner. Histologic examination showed that fibrin deposits were found in all large and small arteries in the lung from mice injected with thrombin; however, fibrin deposits were not detected in any large arteries from the mouse preinjected with thrombomodulin.

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Formycin B-resistant mutants of Chinese hamster ovary cells: novel genetic and biochemical phenotype affecting adenosine kinase

Molecular and Cellular Biology ◽

10.1128/mcb.3.8.1468-1477.1983 ◽

1983 ◽

Vol 3 (8) ◽

pp. 1468-1477

Author(s):

K D Mehta ◽

R S Gupta

Keyword(s):

Chinese Hamster Ovary ◽

Chinese Hamster Ovary Cells ◽

Chinese Hamster ◽

Complex Form ◽

Single Step ◽

Adenosine Kinase ◽

Cross Resistance ◽

Ovary Cells ◽

Cell Extracts

Stable mutants which are approximately three- and eightfold resistant to the pyrazolopyrimidine nucleosides formycin A and formycin B (FomR) have been selected in a single step from mutagenized Chinese hamster ovary cells. In cell extracts, the two FomR mutants which were examined were both found to contain no measurable activity of the enzyme adenosine kinase (AK). However, cross-resistance studies with other adenosine analogs such as toyocamycin and tubercidin show that these mutants are distinct from toyocamycin or tubercidin resistant (Toyr) mutants which also contain no measurable AK activity in cell extracts. Studies on the uptake and incorporation of [3H]adenosine and [3H]tubercidin by various mutants and parental cell lines show that unlike the Toyr mutants, which are severely deficient in the phosphorylation of these compounds, the FomR mutants possess nearly normal capacity to phosphorylate these compounds and incorporate them into cellular macromolecules. These results suggest that the FomR mutants contain normal levels of AK activity in vivo. In cell hybrids formed between FomR X FomS cells and FomR X Toyr cells, the formycin-resistant phenotype of both of the FomR mutants behaved codominantly. However, the extracts from these hybrid cells contained either congruent to 50% (FomR X FomS) or no measurable (FomR X Toyr) AK activity, indicating that the lesion in these mutants neither suppresses the wild-type AK activity nor complements the AK deficiency of the Toyr mutants. The presence of AK activity in the FomR mutants in vivo, but not in their cell extracts, along with the codominant behavior of the mutants in hybrids, indicates that the lesions in the FomR mutant are of a novel nature. It is suggested that the genetic lesion in these mutants affects AK activity indirectly and that it may involve an essential cellular function which exists in a complex form with AK. Some implications of these results regarding the mechanism of action of formycin B are discussed.

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Evaluation of the Toxicity of Intravenous Delivery of Auroshell Particles (Gold–Silica Nanoshells)

International Journal of Toxicology ◽

10.1177/1091581812465969 ◽

2012 ◽

Vol 31 (6) ◽

pp. 584-594 ◽

Cited By ~ 113

Author(s):

Shayne C. Gad ◽

Kelly L. Sharp ◽

Charles Montgomery ◽

J. Donald Payne ◽

Glenn P. Goodrich

Keyword(s):

Water Solution ◽

Chinese Hamster Ovary Cells ◽

Chinese Hamster ◽

Gold Nanoshells ◽

Sprague Dawley ◽

Ovary Cells ◽

Good Laboratory ◽

Chromosomal Aberration Assay

Gold nanoshells (155 nm in diameter with a coating of polyethylene glycol 5000) were evaluated for preclinical biocompatibility, toxicity, and biodistribution as part of a program to develop an injectable device for use in the photothermal ablation of tumors. The evaluation started with a complete good laboratory practice (GLP) compliant International Organization for Standardization (ISO)-10993 biocompatibility program, including cytotoxicity, pyrogenicity (US Pharmacopeia [USP] method in the rabbit), genotoxicity (bacterial mutagenicity, chromosomal aberration assay in Chinese hamster ovary cells, and in vivo mouse micronucleus), in vitro hemolysis, intracutaneous reactivity in the rabbit, sensitization (in the guinea pig maximization assay), and USP/ISO acute systemic toxicity in the mouse. There was no indication of toxicity in any of the studies. Subsequently, nanoshells were evaluated in vivo by intravenous (iv) infusion using a trehalose/water solution in a series of studies in mice, Sprague-Dawley rats, and Beagle dogs to assess toxicity for time durations of up to 404 days. Over the course of 14 GLP studies, the gold nanoshells were well tolerated and, when injected iv, no toxicities or bioincompatibilities were identified.

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In vitro and in vivo functional characterization of bovine vitamin K-dependent gamma-carboxylase expressed in Chinese hamster ovary cells.

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.90.10.4611 ◽

1993 ◽

Vol 90 (10) ◽

pp. 4611-4615 ◽

Cited By ~ 51

Author(s):

A. Rehemtulla ◽

D. A. Roth ◽

L. C. Wasley ◽

A. Kuliopulos ◽

C. T. Walsh ◽

...

Keyword(s):

Vitamin K ◽

Chinese Hamster Ovary ◽

Chinese Hamster Ovary Cells ◽

Functional Characterization ◽

Chinese Hamster ◽

Ovary Cells

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Formycin B-resistant mutants of Chinese hamster ovary cells: novel genetic and biochemical phenotype affecting adenosine kinase.

Molecular and Cellular Biology ◽

10.1128/mcb.3.8.1468 ◽

1983 ◽

Vol 3 (8) ◽

pp. 1468-1477 ◽

Cited By ~ 12

Author(s):

K D Mehta ◽

R S Gupta

Keyword(s):

Chinese Hamster Ovary ◽

Chinese Hamster Ovary Cells ◽

Chinese Hamster ◽

Complex Form ◽

Single Step ◽

Adenosine Kinase ◽

Cross Resistance ◽

Ovary Cells ◽

Cell Extracts

Stable mutants which are approximately three- and eightfold resistant to the pyrazolopyrimidine nucleosides formycin A and formycin B (FomR) have been selected in a single step from mutagenized Chinese hamster ovary cells. In cell extracts, the two FomR mutants which were examined were both found to contain no measurable activity of the enzyme adenosine kinase (AK). However, cross-resistance studies with other adenosine analogs such as toyocamycin and tubercidin show that these mutants are distinct from toyocamycin or tubercidin resistant (Toyr) mutants which also contain no measurable AK activity in cell extracts. Studies on the uptake and incorporation of [3H]adenosine and [3H]tubercidin by various mutants and parental cell lines show that unlike the Toyr mutants, which are severely deficient in the phosphorylation of these compounds, the FomR mutants possess nearly normal capacity to phosphorylate these compounds and incorporate them into cellular macromolecules. These results suggest that the FomR mutants contain normal levels of AK activity in vivo. In cell hybrids formed between FomR X FomS cells and FomR X Toyr cells, the formycin-resistant phenotype of both of the FomR mutants behaved codominantly. However, the extracts from these hybrid cells contained either congruent to 50% (FomR X FomS) or no measurable (FomR X Toyr) AK activity, indicating that the lesion in these mutants neither suppresses the wild-type AK activity nor complements the AK deficiency of the Toyr mutants. The presence of AK activity in the FomR mutants in vivo, but not in their cell extracts, along with the codominant behavior of the mutants in hybrids, indicates that the lesions in the FomR mutant are of a novel nature. It is suggested that the genetic lesion in these mutants affects AK activity indirectly and that it may involve an essential cellular function which exists in a complex form with AK. Some implications of these results regarding the mechanism of action of formycin B are discussed.

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Properties of the kinetochore in vitro. I. Microtubule nucleation and tubulin binding.

The Journal of Cell Biology ◽

10.1083/jcb.101.3.755 ◽

1985 ◽

Vol 101 (3) ◽

pp. 755-765 ◽

Cited By ~ 69

Author(s):

T J Mitchison ◽

M W Kirschner

Keyword(s):

Chinese Hamster Ovary ◽

Chinese Hamster Ovary Cells ◽

Chinese Hamster ◽

Lag Phase ◽

Ovary Cells ◽

Tubulin Binding ◽

Mixed Polarity ◽

Tubulin Immunofluorescence

We have isolated chromosomes from Chinese hamster ovary cells arrested in mitosis with vinblastine and examined the interactions of their kinetochores with purified tubulin in vitro. The kinetochores nucleate microtubule (MT) growth with complex kinetics. After an initial lag phase, MTs are continuously nucleated with both plus and minus ends distally localized. This mixed polarity seems inconsistent with the formation of an ordered, homopolar kinetochore fiber in vivo. As isolated from vinblastine-arrested cells, kinetochores contain no bound tubulin. The kinetochores of chromosomes isolated from colcemid-arrested cells or of chromosomes incubated with tubulin in vitro are brightly stained after anti-tubulin immunofluorescence. This bound tubulin is probably not in the form of MTs. It is localized to the corona region by immunoelectron microscopy, where it may play a role in MT nucleation in vitro.

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