Isolation of several fractions from human serum, inhibiting rabbit cartilage metabolism in vitro

1990 ◽  
Vol 122 (2) ◽  
pp. 277-282 ◽  
Author(s):  
Marc V. L. Du Caju ◽  
Raoul P. Rooman
Keyword(s):  
Human Serum ◽  
Thymidine Incorporation ◽  
Inhibitory Effect ◽  
Cartilage Metabolism ◽  
Molecular Weights ◽  
Rabbit Cartilage ◽  
Molecular Sieving

Abstract In contrast to the well characterized serum stimulators of cartilage metabolism, information is scarce on the nature of circulating inhibitors. Human serum was fractionated by molecular sieving chromatography on Sephadex G-200, G-75, G-50 and Biogel P-4 gels. Six fractions with molecular weights of 150, 45, 30, 16, 9 and 1.2 kD inhibited [35S]sulphate incorporation into rabbit cartilage segments. All fractions but the smallest one exhibited their inhibitory effect only in the serum-stimulated cartilage. The 1.2 kD fraction impaired [35S]sulphate and [3H]methyl-thymidine incorporation into the cartilage segments in both stimulated and basal conditions. A seventh inhibitory fraction corresponded to the serum salt peak.

Effect of steroids on cartilage metabolism "in vitro"

1986 ◽  
Vol 113 (4_Suppl) ◽  
pp. S35-S40 ◽  
Author(s):  
Marc V.L. DU CAJU ◽  
Raoul P. ROOMAN
Keyword(s):  
Growth Factor ◽  
Thymidine Incorporation ◽  
Normal Serum ◽  
Synthetic Analogue ◽  
Factor Activity ◽  
Poor Growth ◽  
Cartilage Metabolism ◽  
Glucocorticoid Administration

ABSTRACT Conditions characterized by high levels of glucocorticoids are associated with poor growth. Serum somatomedin or insulin-like growth factor activity measured by cartilage bioassay systems is low, but is generally not accompanied by a fall in somatomedin concentration. Hydrocortisone and a synthetic analogue, dexamethasone, impaired the serum stimulated "in vitro" 35S sulphate and 3H-thymidine incorporation in porcine rib cartilage at physiological concentrations. Hydrocortisone added at a concentration of 0,1 μg/ml decreased the potency of normal serum to 50 % of controls. Dexamethasone was at least 10 times more potent. Removal of "in vitro" or "in vivo" administered hydrocortisone with dextran-coated charcoal restored the sulphate and thymidine activity to normal. We conclude that physiological amounts of glucocorticoids inhibit the "in vitro" porcine cartilage metabolism. Glucocorticoid administration "in vivo" does not abolish the activity of the cartilage stimulating effect of serum but affects cartilage metabolism directly or by the induction of locally produced inhibitors of cartilage metabolism.

On the Complex Formation of Antithrombin III with Thrombin

1973 ◽  
Vol 30 (02) ◽  
pp. 280-283 ◽  
Author(s):  
B Binder
Keyword(s):  
Complex Formation ◽  
Human Serum ◽  
Blood Coagulation ◽  
Antithrombin Iii ◽  
Normal Human Serum ◽  
Molecular Weights ◽  
At Iii ◽  
Normal Human

SummaryBased on gelfiltration studies, the part of AT III which becomes bound to thrombin during the process of in vitro blood coagulation was calculated to be about 40% of total AT III. Complexes consisting of one AT III and one thrombin molecule could not be detected while fractions corresponding to molecular weights of about 190,000 Dalton show AT III as well as thrombin activities. The AT III - thrombin complex in normal human serum consists, therefore, of either 2 AT III and 2 thrombin molecules or of one AT III and 4 thrombin molecules.

scholarly journals Phytohemagglutinin-stimulated normal human peripheral blood lymphocytes in folate-depleted medium: an in vitro model for megaloblastic hemopoiesis

Blood ◽  
1980 ◽  
Vol 55 (5) ◽  
pp. 863-865 ◽  
Author(s):  
RJ Hayman ◽  
MB Van Der Weyden
Keyword(s):  
Peripheral Blood ◽  
Thymidine Incorporation ◽  
Human Peripheral Blood ◽  
Inhibitory Effect ◽  
Peripheral Blood Lymphocytes ◽  
Blood Lymphocytes ◽  
Human Peripheral Blood Lymphocytes ◽  
Normal Human ◽  
Normal Human Peripheral Blood

Abstract Normal human peripheral blood lymphocytes cultured with phytohemagglutinin in folate-free RPMI 1640 medium supplemented with normal human serum exhibit megaloblastic maturation. These changes are accompanied by a decrease in intracellular folate content, and when compared to folate-replete cells, reveal increased 3H-thymidine incorporation into DNA together with a decreased inhibitory effect of deoxyuridine on 3H-thymidine incorporation. This in vitro system of cellular folate deficiency is a convenient model for analysis of biochemical events accompanying megaloblastic maturation.

scholarly journals Phytohemagglutinin-stimulated normal human peripheral blood lymphocytes in folate-depleted medium: an in vitro model for megaloblastic hemopoiesis

Blood ◽  
1980 ◽  
Vol 55 (5) ◽  
pp. 863-865
Author(s):  
RJ Hayman ◽  
MB Van Der Weyden
Keyword(s):  
Peripheral Blood ◽  
Thymidine Incorporation ◽  
Human Peripheral Blood ◽  
Inhibitory Effect ◽  
Peripheral Blood Lymphocytes ◽  
Blood Lymphocytes ◽  
Human Peripheral Blood Lymphocytes ◽  
Normal Human ◽  
Normal Human Peripheral Blood

Normal human peripheral blood lymphocytes cultured with phytohemagglutinin in folate-free RPMI 1640 medium supplemented with normal human serum exhibit megaloblastic maturation. These changes are accompanied by a decrease in intracellular folate content, and when compared to folate-replete cells, reveal increased 3H-thymidine incorporation into DNA together with a decreased inhibitory effect of deoxyuridine on 3H-thymidine incorporation. This in vitro system of cellular folate deficiency is a convenient model for analysis of biochemical events accompanying megaloblastic maturation.

scholarly journals Soluble forms of the interleukin-6 signal-transducing receptor component gp130 in human serum possessing a potential to inhibit signals through membrane-anchored gp130

Blood ◽  
1993 ◽  
Vol 82 (4) ◽  
pp. 1120-1126 ◽  
Author(s):  
M Narazaki ◽  
K Yasukawa ◽  
T Saito ◽  
Y Ohsugi ◽  
H Fukui ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
Human Serum ◽  
Interleukin 6 ◽  
Inhibitory Effect ◽  
Oncostatin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Healthy Human ◽  
Molecular Weights ◽  
Human Sera ◽  
Neurotropic Factor

Abstract The interleukin-6 (IL-6) signal is transduced through membrane-anchored gp130, which is associated with IL-6 receptor (IL-6R) in the presence of IL-6. Soluble forms of gp130 (sgp130) with molecular weights of 90 and 110 Kd were found in human serum. In the presence of recombinant IL- 6 (rIL-6), serum sgp130 were capable of associating with serum sIL-6R. By the sandwich enzyme-linked immunosorbent assay, healthy human sera was shown to contain 390 +/- 72 ng/mL of sgp130. A mouse pro-B-cell line-derived transfectant, BAF-130, expressing human gp130 was used to examine the function of serum sgp130. When supplemented with rIL-6, human serum induced DNA synthesis in BAF-130 cells, whereas the serum deprived of sIL-6R did not. In contrast, the DNA synthesis induced in BAF-130 cells by rIL-6-supplemented serum was increased when the serum was deprived of sgp130. These results indicated that serum sgp130 could negatively regulate the IL-6 signal. Recently, gp130 has been shown to be involved in the signaling processes of oncostatin M, leukemia inhibitory factor, and ciliary neurotropic factor, in addition to those of IL-6. Recombinant sgp130 showed inhibitory effect on the biologic function of such cytokines. This work implies physiologic roles of naturally produced serum sgp130 in modulating signals through gp130.

scholarly journals Soluble forms of the interleukin-6 signal-transducing receptor component gp130 in human serum possessing a potential to inhibit signals through membrane-anchored gp130

Blood ◽  
1993 ◽  
Vol 82 (4) ◽  
pp. 1120-1126 ◽  
Author(s):  
M Narazaki ◽  
K Yasukawa ◽  
T Saito ◽  
Y Ohsugi ◽  
H Fukui ◽  
...  
Keyword(s):  
Dna Synthesis ◽  
Human Serum ◽  
Interleukin 6 ◽  
Inhibitory Effect ◽  
Oncostatin M ◽  
Enzyme Linked Immunosorbent Assay ◽  
Healthy Human ◽  
Molecular Weights ◽  
Human Sera ◽  
Neurotropic Factor

The interleukin-6 (IL-6) signal is transduced through membrane-anchored gp130, which is associated with IL-6 receptor (IL-6R) in the presence of IL-6. Soluble forms of gp130 (sgp130) with molecular weights of 90 and 110 Kd were found in human serum. In the presence of recombinant IL- 6 (rIL-6), serum sgp130 were capable of associating with serum sIL-6R. By the sandwich enzyme-linked immunosorbent assay, healthy human sera was shown to contain 390 +/- 72 ng/mL of sgp130. A mouse pro-B-cell line-derived transfectant, BAF-130, expressing human gp130 was used to examine the function of serum sgp130. When supplemented with rIL-6, human serum induced DNA synthesis in BAF-130 cells, whereas the serum deprived of sIL-6R did not. In contrast, the DNA synthesis induced in BAF-130 cells by rIL-6-supplemented serum was increased when the serum was deprived of sgp130. These results indicated that serum sgp130 could negatively regulate the IL-6 signal. Recently, gp130 has been shown to be involved in the signaling processes of oncostatin M, leukemia inhibitory factor, and ciliary neurotropic factor, in addition to those of IL-6. Recombinant sgp130 showed inhibitory effect on the biologic function of such cytokines. This work implies physiologic roles of naturally produced serum sgp130 in modulating signals through gp130.

scholarly journals The Presence of Chemical Cross-Linking Stabilises HIV-1 Envelope Glycoprotein Trimer Antigens in a Model of Intramuscular Immunisation

2021 ◽  
Author(s):  
Luiza Farache Trajano ◽  
Rebecca Moore ◽  
Quentin Sattentau
Keyword(s):  
Human Serum ◽  
Envelope Glycoprotein ◽  
Proteolytic Enzymes ◽  
Neutrophil Infiltration ◽  
Inhibitory Effect ◽  
Cross Linking ◽  
The Stability ◽  
Antibody Epitopes ◽  
Hiv 1

Background: The HIV-1 envelope glycoprotein (Env) is the target of antigen design for antibody- based vaccination. In 2019, four trimeric Env vaccines entered an experimental trial: ConM, ConS, and their cross-linked counterparts. The trimers were formulated with MPLA adjuvant. Studies have demonstrated that adjuvants trigger neutrophil infiltration. Neutrophils activate and degranulate releasing proteases, namely elastase and cathepsinG. Aims: To assess the stability and immunogenicity of these vaccines in the presence of adjuvant- recruited neutrophils and their proteolytic enzymes. Methods: Trimers were incubated with commercially-sourced proteases. To analyse stability, samples were reduced, denatured and separated using gel electrophoresis. To assess antibody binding, a trimer-protease incubation was followed by an ELISA. To establish more physiologically relevant conditions, harvested neutrophils were exposed to various adjuvants. The supernatant, shown to contain elastase, was incubated alongside the vaccines. The reducing and denaturing gels, as well as the ELISA, was repeated. Results: Gel analysis revealed that un-crosslinked trimers underwent significant digestion whereas cross-linking conferred enhanced stability. In the presence of neutrophil-sourced protease-containing-supernatant, trimers displayed resistance to digestion. The differential stability profile of Env trimers when exposed to commercially sourced compared to supernatant- derived proteases may be due to the inhibitory effect of human serum on elastase. Antibody epitopes were maintained in vitro. Conclusion: The vaccine antigens are sensitive to enzymatic degradation. This is reduced by cross-linking and human serum.

Neutrophil Oxygen Metabolite Inhibition of Cultured Chinchilla Middle Ear Epithelial Cell Growth

1994 ◽  
Vol 103 (10) ◽  
pp. 812-816 ◽  
Author(s):  
Masahiro Kawana ◽  
Chihiro Kawana ◽  
Steven K. Juhn ◽  
Ryo Amesara ◽  
G. Scott Giebink
Keyword(s):  
Cell Growth ◽  
Middle Ear ◽  
Thymidine Incorporation ◽  
Inhibitory Effect ◽  
Human Neutrophils ◽  
Middle Ear Effusion ◽  
Metabolic Products ◽  
Activated Neutrophils ◽  
Ear Effusion

Middle ear inflammation in acute bacterial otitis media is characterized by accumulation of neutrophils in middle ear effusion. Since neutrophils release products that may injure surrounding tissues, we studied the effect of neutrophil metabolic products on middle ear epithelial cells (MEECs) in vitro. Chinchilla MEECs were incubated with phorbol myristate acetate (PMA)–activated human neutrophils or with hydrogen peroxide (H2O2). Cell growth, which was measured by 3H-thymidine incorporation, was inhibited by activated neutrophils and by H2O2. Unstimulated neutrophils, PMA alone, and catalase alone did not affect the viability of MEECs. Catalase, an enzyme that reduces H2O2, partially blocked the inhibitory effect of activated neutrophils and completely blocked the inhibitory effect of H2O2. Inhibition of MEEC metabolism by neutrophil-reactive oxygen species may contribute to epithelial injury, which may prolong the middle ear inflammatory response and lead to chronic tissue damage.

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