Copper transport in rats involving a new plasma protein

1985 ◽  
Vol 249 (1) ◽  
pp. E77-E88 ◽  
Author(s):  
K. C. Weiss ◽  
M. C. Linder
Keyword(s):  
Time Course ◽  
Specific Activity ◽  
High Specific Activity ◽  
Apparent Molecular Weight ◽  
Female Rats ◽  
Whole Body ◽  
Peripheral Tissues ◽  
Chelex 100

The time course of distribution of high-specific activity 67CuCl2 to tissues and plasma components was followed in adult, female rats. Immediately after intubation or injection, tracer 67Cu associated with two components of the blood plasma separable on columns of Sephadex G-150: albumin and another (larger) component, which was not ceruloplasmin. The latter, tentatively named transcuprein, had an apparent molecular weight of 270,000 and a high affinity for Cu2+, as judged by processing through Chelex-100, dilution, and exchange with albumin copper, in vitro and in vivo. It was capable of donating copper to tumor cells in serum-free medium. Analysis of "cold" plasma by furnace atomic absorption confirmed the presence of 10-15% of plasma copper in this peak. Plots of percent dose and 67Cu specific activity against time showed that copper followed a very specific pathway after binding to albumin and transcuprein, entering mainly the liver, then reappearing in the plasma on ceruloplasmin, and then achieving peak distribution in peripheral tissues (muscles, brain, etc.). 67Cu disappeared from liver and kidney with an apparent half-life of 4.5 days, the same exponential rate found for whole body turnover. Apparent turnover of ceruloplasmin copper was more rapid. Even after 7-12 days, tracer copper in plasma was still found exclusively with ceruloplasmin. The results indicate that copper follows a carefully prescribed path, on entering the blood and binding to a new transport protein.

Effects of Heparin Oligosaccharides with High Affinity for Antithrombin III in Experimental Venous Thrombosis

1982 ◽  
Vol 47 (03) ◽  
pp. 244-248 ◽  
Author(s):  
D P Thomas ◽  
Rosemary E Merton ◽  
T W Barrowcliffe ◽  
L Thunberg ◽  
U Lindahl
Keyword(s):  
Antithrombin Iii ◽  
Specific Activity ◽  
High Specific Activity ◽  
Factor Xa ◽  
Blood Levels ◽  
Thrombin Time ◽  
High Affinity ◽  
Very High

SummaryThe in vitro and in vivo characteristics of two oligosaccharide heparin fragments have been compared to those of unfractionated mucosal heparin. A decasaccharide fragment had essentially no activity by APTT or calcium thrombin time assays in vitro, but possessed very high specific activity by anti-Factor Xa assays. When injected into rabbits at doses of up to 80 ¼g/kg, this fragment was relatively ineffective in impairing stasis thrombosis despite producing high blood levels by anti-Xa assays. A 16-18 monosaccharide fragment had even higher specific activity (almost 2000 iu/mg) by chromogenic substrate anti-Xa assay, with minimal activity by APTT. When injected in vivo, this fragment gave low blood levels by APTT, very high anti-Xa levels, and was more effective in preventing thrombosis than the decasaccharide fragment. However, in comparison with unfractionated heparin, the 16-18 monosaccharide fragment was only partially effective in preventing thrombosis, despite producing much higher blood levels by anti-Xa assays.It is concluded that the high-affinity binding of a heparin fragment to antithrombin III does not by itself impair venous thrombogenesis, and that the anti-Factor Xa activity of heparin is only a partial expression of its therapeutic potential.

scholarly journals A New Technic for the Production of an in Vivo Labeled Fibrinogen

Blood ◽  
1967 ◽  
Vol 29 (4) ◽  
pp. 517-525 ◽  
Author(s):  
HENRY GANS ◽  
JAMES MC LEOD ◽  
JAMES T. LOWMAN
Keyword(s):  
Amino Acid ◽  
Specific Activity ◽  
High Specific Activity ◽  
Entire Period ◽  
Turnover Rates ◽  
Prolonged Infusion ◽  
Slow Infusion

Abstract The fact that in vitro labeled proteins, as a rule, exhibit faster turnover rates than in vivo labeled materials led us to explore means of obtaining in vivo labeled fibrinogen of high specific activity. It was found that defibrination of the rat provides a stimulus for the liver to regenerate fibrinogen at an accelerated rate. Administration of seleno75 methionine shortly after thrombin-induced defibrination of the animal resulted in the incorporation of large quantities of the label. The rate of incorporation was further increased if the amino acid was administered as a slow infusion during the entire period of fibrinogen regeneration. In addition, prior nephrectomy of the animal would appear to result in a slight increase in specific activity of the fibrinogen preparation obtained. The results of these studies indicate that defibrination, nephrectomy, and the prolonged infusion of the labeled amino acid selenomethionine provided us with a technic for obtaining a biosynthetically labeled, γ-emitting, fibrinogen preparation of high specific activity.

Human placental lactogen directly inhibits rat cartilage growth processes in vivo and in vitro

1993 ◽  
Vol 128 (1) ◽  
pp. 65-68 ◽  
Author(s):  
Mimi H Chiang ◽  
Kevin M Kelley ◽  
Charles S Nicoll
Keyword(s):  
Cartilage Explants ◽  
Female Rats ◽  
Placental Lactogen ◽  
Human Placental Lactogen ◽  
Cartilage Growth ◽  
Growth Processes ◽  
Peripheral Tissues ◽  
Placental Hormone

We have recently reported that human placental lactogen inhibits the growth of young female rats without changing the serum levels of insulin like growth factor-I. Accordingly, experiments were conducted to determine whether human placental lactogen could directly inhibit cartilage growth processes in vivo and in vitro. Osmotic minipumps with attached polyethylene catheters were used to infuse the hormone for seven days into the left hindlimb of three-month-old female rats via the common iliac artery. The right hindlimb of each animal served as an internal control. Infusion of the placental lactogen at 10 μg/rat/day caused a slight (10%) but significant decrease in the width of the tibial epiphysial cartilage plate and a higher dose (100 μg/rat/day) caused a greater degree of inhibition (25%). However, the higher dose also inhibited the tibial cartilage plate of the contralateral (non-infused) limb. The possibility that human placental lactogen could directly inhibit cartilage anabolic activity in vitro was evaluated by measuring the incorporation of 35SO4 into costal cartilage explants from three to four-month-old female rats. The placental hormone inhibited the incorporation of 35SO4 in a dose-related manner at concentrations ranging from 1.0 to 100 μg/l. As a test of the specificity of this inhibition the effect of the hormone on the incorporation of 35SO4 into cartilage explants from Coho salmon was determined. The placental lactogen did not affect incorporation of the sulfate into the fish cartilage over a range of doses from 1.0 to 1000 μg/l. These results indicate that at least some of the inhibitory effects of human placental lactogen on the growth of rats is direct on peripheral tissues, such as cartilage. This effect may be mediated by the well-established anti-insulin action of the placental hormone.

Studies on the toxicity of copper. I. The toxic action of copper in vivo and in vitro

1966 ◽  
Vol 166 (1004) ◽  
pp. 273-284 ◽  
Keyword(s):  
Microsomal Fraction ◽  
Specific Activity ◽  
Membrane Surface ◽  
High Specific Activity ◽  
Rapid Onset ◽  
Significant Inhibition ◽  
Low Concentrations ◽  
The Brain

With the object of throwing light upon the brain damage found in patients with Wilson’s disease (hepato-lenticular degeneration) due to the accumulation of copper, the effect of Cu 2+ has been investigated in pigeons. Subarachnoid injections of Cu 2+ (10 to 25 µ g) led to rapid onset of convulsions and death. These concentrations of Cu 2+ inhibited pigeon and rat b rain mitochondria; more organized tissue breis or slices showed no significant inhibition of oxygen up take at Cu +2 concentration inducing convulsions in vivo . Studies with radioactive copper ( 64 Cu) showed that the injected copper was widely distributed in the brain, though maximal near the site of injection. Centrifugation showed a high specific activity in the ATP -ase-rich microsomal fraction. Thorium in concentrations similar to Cu 2+ was not toxic. From this we suggest that the Cu 2+ does not alter the charge on some membrane surface. Since the effect of the copper is immediate, and since it does not affect respiration of slices in these low concentrations, we conclude that it is exerting its convulsive effect directly upon the cell surfaces.

scholarly journals Role of 11β-hydroxysteroid dehydrogenase type 1 in the development of atopic dermatitis

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Noo Ri Lee ◽  
Beom Jun Kim ◽  
Chung Hyeok Lee ◽  
Young Bin Lee ◽  
Solam Lee ◽  
...  
Keyword(s):  
Atopic Dermatitis ◽  
De Novo ◽  
Hydroxysteroid Dehydrogenase ◽  
Whole Body ◽  
Epidermal Keratinocytes ◽  
Human Epidermal Keratinocytes ◽  
Peripheral Tissues

AbstractGlucocorticoids (GCs) are potent anti-inflammatory drugs, the secretion of which is mediated and controlled by the hypothalamic–pituitary–adrenal axis. However, they are also secreted de novo by peripheral tissues for local use. Several tissues express 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1), including the skin. The inactive GC cortisone is converted by 11β-HSD1 to active GC cortisol, which is responsible for delayed wound healing during a systemic excess of GC. However, the role of 11β-HSD1 in inflammation is unclear. We assessed whether 11β-HSD1 affects the development of atopic dermatitis (AD) in vitro and in vivo. The expression of 11β-HSD1 in the epidermis of AD lesions was higher than that in the epidermis of healthy controls. Knockdown of 11β-HSD1 in human epidermal keratinocytes increased the production of thymic stromal lymphopoietin. In an oxazolone-induced mouse model of AD, localized inhibition of 11β-HSD1 aggravated the development of AD and increased serum cytokine levels associated with AD. Mice with whole-body knockout (KO) of 11β-HSD1 developed significantly worse AD upon induction by oxazolone. We propose that 11β-HSD1 is a major factor affecting AD pathophysiology via suppression of atopic inflammation due to the modulation of active GC in the skin.

Very stable 188Re-S4 chelates for labelling biomolecules

2007 ◽  
Vol 46 (05) ◽  
pp. 181-184 ◽  
Author(s):  
C. Jentsche ◽  
R. Bergmann ◽  
H.-J. Pietzsch ◽  
G. Wunderlich ◽  
J. Kotzerke ◽  
...  
Keyword(s):  
Ascorbic Acid ◽  
Specific Activity ◽  
Direct Reduction ◽  
High Specific Activity ◽  
Amine Group ◽  
Stability Studies ◽  
Therapeutic Application ◽  
Carboxylic Groups

SummaryAim: The preparation and stability of a new 188Re-S4-complex [S4 = (1-aza-18-crown-6)(O)C-C(SH)-C(SH)- C(O)NH-(CH2)3-NH-(CH2)3-NHC(O)-C(SH)-C(SH)- C(O)(1-aza-18-crown-6] was studied at therapeutic relevant radioactive concentrations. The results were compared with 188Re-MAG3 (MAG3: mercaptoacetyltriglycine) and 188Re-DMSA preparations (DMSA: dimercaptosuccinic acid) performed with the same highly concentrated [188Re]perrhenate solution (12-15 GBq/ml). Methods: The 188Re complexes were prepared by direct reduction of perrhenate (188Re-S4-complex) as well as via the 188Re- EDTA precursor complex (188Re-MAG3, 188Re-DMSA). The preparations were stabilised with 15 mg of ascorbic acid and analysed after 1, 2, and 24 hours by TLC and HPLC. Additionally, in vitro and in vivo stability studies were performed with the purified complexes. Results: After stabilisation with 15 mg of ascorbic acid, all of the complexes were nearly stable under nitrogen for hours, and only 2–8 % of perrhenate was observed after 24 h. In contrast, only the 188Re-S4 complex was completely stable in vitro and in all investigated in vivo samples after separation of ligand excess and reducing agent by HPLC. Conclusion: The bridging amine group or free carboxylic groups of the S4-ligand framework make available reactive positions for coupling biomolecules to the chelate. Thus it appears that the new 188Re-S4 complexes offer the possibility of stable and high specific activity labelling of biomolecules for therapeutic application.

Studies in Man and Experimental Animals of a Low Molecular Weight Heparin Fraction

1981 ◽  
Vol 45 (03) ◽  
pp. 214-218 ◽  
Author(s):  
D P Thomas ◽  
R E Merton ◽  
W E Lewis ◽  
T W Barrowcliffe
Keyword(s):  
Molecular Weight ◽  
Low Molecular Weight Heparin ◽  
Specific Activity ◽  
Ex Vivo ◽  
High Specific Activity ◽  
Factor Xa ◽  
In Vivo Studies ◽  
Low Molecular Weight

SummaryIn vitro and in vivo studies were carried out on a commercially prepared low molecular weight heparin fraction. By APTT assay the fraction had a specific activity of half that of unfractionated mucosal heparin, yet retained full potency by anti-Xa assay (both clotting and chromogenic substrate). When administered intravenously to human volunteers, the anti-Xa/APTT ratio remained the same as it was in vitro. However, after subcutaneous injection, the ratio increased and anti-Xa activity could not be fully neutralized ex vivo by PF4. The fraction was as effective as unfractionated heparin in preventing experimental serum-induced thrombosis, suggesting that a heparin fraction with high specific activity by anti-Factor Xa assay compared to APTT activity may be an effective drug for the prophylaxis of venous thrombosis.

In vitro and in vivo characterization of high specific activity S-(−)[11C]CGP-12177, a radioligand for β-adrenoreceptor, in rats

2004 ◽  
Vol 1264 ◽  
pp. 261-266 ◽  
Author(s):  
Ken-ichi Nishijima ◽  
Yuji Kuge ◽  
Noriko Motoki ◽  
Koh-ichi Seki ◽  
Kazue Ohkura ◽  
...  
Keyword(s):  
Specific Activity ◽  
High Specific Activity ◽  
Cgp 12177 ◽  
In Vivo Characterization

Insulin stimulation of glucose uptake in skeletal muscles and adipose tissues in vivo is NO dependent

1998 ◽  
Vol 274 (4) ◽  
pp. E692-E699 ◽  
Author(s):  
Denis Roy ◽  
Mylène Perreault ◽  
André Marette
Keyword(s):  
Glucose Uptake ◽  
Skeletal Muscles ◽  
Whole Body ◽  
Glucose Disposal ◽  
Adipose Tissues ◽  
Peripheral Tissues ◽  
Nos Inhibitor ◽  
Oxide Synthase

The purpose of this study was to investigate whether in vivo nitric oxide synthase (NOS) inhibition influences insulin-mediated glucose disposal in rat peripheral tissues. The NOS inhibitor N G-nitro-l-arginine methyl ester (l-NAME) or saline was infused constantly during a hyperinsulinemic-euglycemic clamp in normal rats. Glucose utilization rates of insulin-sensitive tissues (individual muscles, heart, and adipose tissues) were simultaneously determined using tracer infusion of 2-deoxy-d-[3H]glucose (2-[3H]DG). NOS blockade with l-NAME resulted in significant ( P < 0.05) reduction in both whole body glucose disposal (−16%, P < 0.01) and plasma 2-[3H]DG disappearance rate (−30%, P < 0.05) during hyperinsulinemic-euglycemic clamp.l-NAME significantly decreased insulin-stimulated glucose uptake in heart (−62%, P = 0.01), soleus (−42%, P = 0.05), red (−53%, P < 0.001) and white (−62%, P < 0.001) gastrocnemius, tibialis (−57%, P < 0.01), and quadriceps (−33%, P < 0.05) muscles. The NOS inhibitor also decreased insulin action in brown interscapular (−47%, P < 0.01), retroperitoneal (−52%, P = 0.07), and gonadal (−66%, P = 0.06) adipose tissues. In contrast to in vivo NOS blockade,l-NAME failed to affect basal or insulin-stimulated 2-[3H]DG transport in isolated soleus or extensor digitorum longus muscles in vitro. These results support the hypothesis that the action of insulin to augment glucose uptake by skeletal muscles and other peripheral insulin-sensitive tissues in vivo is NO dependent.

scholarly journals Biochemical characterization of the active heterodimer form of human heparanase (Hpa1) protein expressed in insect cells

2003 ◽  
Vol 373 (2) ◽  
pp. 423-435 ◽  
Author(s):  
Edward McKENZIE ◽  
Kathryn YOUNG ◽  
Margaret HIRCOCK ◽  
James BENNETT ◽  
Maina BHAMAN ◽  
...  
Keyword(s):  
Insect Cells ◽  
Biochemical Characterization ◽  
Specific Activity ◽  
Expression System ◽  
Heparan Sulphate ◽  
High Specific Activity ◽  
Vitro Assay ◽  
Tumour Metastasis

The mammalian endoglycosidase heparanase (Hpa1) is primarily responsible for cleaving heparan sulphate proteoglycans (HSPGs) present on the basement membrane of cells and its potential for remodelling the extracellular matrix (ECM) could be important in embryonic development and tumour metastasis. Elevated expression of this enzyme has been implicated in various pathological processes including tumour cell proliferation, metastasis, inflammation and angiogenesis. The enzyme therefore represents a potential therapeutic target. Hpa1 protein is initially synthesized as an inactive 65 kDa proenzyme that is then believed to be subsequently activated by proteolytic cleavage to generate an active heterodimer of 8 and 50 kDa polypeptides. By analysis of a series of Hpa1 deletion proteins we confirm that the 8 kDa subunit is essential for enzyme activity. We present here for the first time an insect cell expression system used for the generation of large amounts of recombinant protein of high specific activity. Individual subunits were cloned into baculoviral secretory vectors and co-expressed in insect cells. Active secreted heterodimer protein was recovered from the medium and isolated by a one-step heparin–Sepharose chromatography procedure to give protein of >90% purity. The recombinant enzyme behaved similarly to the native protein with respect to the size of HS fragments liberated on digestion, substrate cleavage specificity and its preference for acidic pH. A significant amount of activity, however, was also detectable at physiological pH values, as measured both by an in vitro assay and by in vivo degradation of cell-bound heparan sulphate.

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