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PubMed1975

Regulation of carbohydrate metabolism in lymphoid tissue. Quantitative aspects of [U-14C]glucose oxidation by rat spleen slices.

When washed spleen slices from fed rats are incubated with 3 mm-[U-14C]glucose, the rate of glucose utilization (46.2 mumol/h per g dry wt.) is sufficient to account, theoretically, for 80% of the O2 consumption. Measurement of net lactate production, however, and the fate of the radioactive carbon, indicates that the contribution of glucose to the respiratory fuel of the tissue is only 25-30% whereas 60-70% of the glucose utilized is converted into lactate. At saturating glucose concentrations (above 5 mm) its contribution to the respiratory fuel of the slice is increased to a maximum value of 34-39%. Only 2% of the glucose utilized is metabolized via the oxidative steps of the pentose phosphate pathway. Starvation for 72 h marginally increases both the rate of glucose utilization (by 21%) and its net contribution to the respiratory fuel (by 29%). Insulin, glucagon, adrenaline and adenosine 3':5'-cyclic monophosphate have no significant effect on either the rate of glucose utilization or on the pattern of radioactive isotope distribution. The uptake of glucose is increased by only 20%, whereas the production of lactate doubles when slices are incubated under anaerobic conditions. In assessing the suitability of spleen slices for metabolic studies, the only serious major perturbation, compared with the freeze-clamped organ, is an elevated mitochondrial [NAD+]/[NADH] ratio (connected with increased endogenous NH3 production) that is partially restored to normal values on incubation with glucose. Equal proportions of erythrocytes and leucocytes are found in the washed spleen slice. Metabolic contributions of the constituent cell populations in the washed slice are calculated and it is concluded that lymphocytes account for the major part of the glycolytic metabolism (80-90%), whereas the contribution of erythrocytes is insignificant.

Adenosine Diphosphate↗
PubMed1977

Inhibition of protein degradation in isolated rat hepatocytes.

1. Isolated parenchymal cells were prepared by collagenase perfusion of livers from fed rats that had been previously injected with [(3)H]leucine to label liver proteins. When these cells were incubated in a salts medium containing glucose, gelatin and EDTA, cellular integrity was maintained over a period of 6h. 2. Cells incubated in the presence of 2mm-leucine to minimize radioactive isotope reincorporation released [(3)H]leucine into the medium at a rate accounting for the degradation of 4.5% of the labelled cell protein per h. 3. Degradation of [(3)H]protein in these cells was inhibited by insulin and by certain amino acids, of which tryptophan and phenylalanine were the most effective. 4. Protein degradation was decreased by several proteinase inhibitors, particularly those that are known to inhibit lysosomal cathepsin B, and by inhibitors of cell-energy production. 5. Ammonia inhibited degradation, but only at concentrations above 1.8mm. Aliphatic analogues of ammonia were effective at lower concentrations than was ammonia. 6. High concentrations of ammonia inhibited degradation by 50%. The extent of this inhibition could not be increased further by the addition of the cathepsin B inhibitor leupeptin, which by itself inhibited degradation by approx. 30%. 7. The sensitivity of proteolysis in isolated hepatocytes to these various inhibitory agents is discussed in relation to their possible modes of action.

Amino Acids↗
PubMed1978

Pyruvate-to-ethanol pathway in Entamoeba histolytica.

The pyruvate-to-ethanol pathway in Entamoeba histolytica is unusual when compared with most investigated organisms. Pyruvate decarboxylase (EC 4.1.1.1), a key enzyme for ethanol production, is not found. Pyruvate is converted into acetyl-CoA and CO2 by the enzyme pyruvate synthase (EC 1.2.7.1), which has been demonstrated previously in this parasitic amoeba. Acetyl-CoA is reduced to acetaldehyde and CoA by the enzyme aldehyde dehydrogenase (acylating) (EC 1.2.1.10) at an enzyme activity of 9 units per g of fresh cells with NADH as a reductant. Acetaldehyde is further reduced by either a previously identified NADP+-linked alcohol dehydrogenase or by a newly found NAD+-linked alcohol dehydrogenase at an enzyme activity of 136 units per g of fresh cells. Ethanol is identified as the product of soluble enzymes of amoeba acting on pyruvate or acetyl-CoA. This result is confirmed by radioactive isotopic, spectrophotometric and gas-chromatographic methods.

Acetaldehyde↗
PubMed1978

Turnover of metallothioneins in rat liver.

Two electrophoretically distinguishable metallothioneins were isolated from the livers of Cd2+-treated rats and had thiol group/metal ratios of 3:1, a total metal content, in each of these proteins, of 3.6 atoms of Cd2+ + 2.4 atoms of Zn2+/molecule and 4.2 atoms of Cd2+ + 2.8 atoms of Zn2+/molecule and respective apoprotein mol.wts. of 5844 and 6251. Studies with 1 h pulse labels of [3H]cysteine, given after a single injection of ZnCl2 or CdCl2, showed that these metals stimulated radioactive isotope incorporation into the metallothioneins over the control value by 10- and 15-fold respectively. This stimulation was maximal at 4 h after a single CdCl2 injection and decreased to control values by 16 h, suggesting that either a translational event is responding to free intracellular Cd2+ or a short-lived mRNA is being produced or stabilized in response to the metal treatment. In rats chronically exposed to CdCl2, the metallothioneins increased to 0.2% of the liver wet weight from a control value of 2--4 mumol/kg of liver, with a maximum rate of accumulation of 2--3 mumol/h per kg of liver. The turnover of these proteins in control animals was 0.3--0.6 mumoles/h per kg of liver, measured by the rate of disappearance of 203Hg2+, which binds irreversibly to the metallothioneins. Pretreatment with CdCl2 completely stopped the rapid 203Hg turnover observed in untreated animals. Unlike CdCl2, treatment with ZnCl2 increased the concentration of metallothioneins to a new steady-state pool, 11 mumole/kg of liver, after 10 h. The increase in the zinc-thionein pool by exposure to ZnCl2 in vivo was determined to be primarily due to a stimulation of metallothionein biosynthesis.

Amino Acids↗
PubMed1980

Microbial metabolism of amino alcohols. Biosynthetic utilization of ethanolamine for lipid synthesis by bacteria.

1. Ten bacteria utilizing [2-14C]ethanol-2-amine as the sole or major source of nitrogen for growth on glycerol + salts medium incorporated radioactivity into a variety of bacterial substances. A high proportion was commonly found in lipid fractions, particularly in the case of Erwinia carotovora. 2. Detailed studies of [14C]ethanolamine incorporation into lipids by five bacteria, including E. carotovora, showed that all detectable lipids were labelled. Even where phosphatidylethanolamine was the major lipid labelled, radioactivity was predominantly in the fatty acid rather than the base moiety. The labelled fatty acids were identified in each case. 3. The addition of acetate to growth media decreased the incorporation of radioactivity from ethanolamine into both fatty acid and phosphatidyl-base fragments of lipids from all the bacteria except Mycobacterium smegmatis. Experiments with [3H]ethanolamine and [14C]acetate confirmed that unlabelled acetate decreased the incorporation of both radioactive isotopes into lipids, except in the case of M. smegmatis. 4. Enzyme studies suggested one of two metabolic routes between ethanolamine and acetyl-CoA for each of four bacteria. A role for ethanolamine O-phosphate was not obligatory for the incorporation of [14C]ethanolamine into phospholipids, but correlated with CoA-independent aldehyde dehydrogenase activity.

Acetates↗
PubMed1981

Zinc metalloenzyme properties of active and latent collagenase from rabbit bone.

1. Inhibition of collagenase from rabbit bone cultures by the chelating agents 1,10-phenanthroline and EDTA is almost completely reversed by Zn2+; other metal cations are less effective in reversing the inhibition. Optimal restoration of activity is achieved at Zn2+ concentrations below that of the chelator, but excess of Zn2+ is inhibitory. 2. Prolonged incubation of collagenase with either chelator causes irreversible inactivation. This inactivation is prevented by Zn2+ at the same concentrations needed to reverse the primary inhibition. 3. Collagenase incorporates 65Zn by exchange when incubated with 1,10-phenanthroline and Zn2+ containing this radioactive isotope. The 65Zn2+ can be removed from its binding site in collagenase by 1,10-phenanthroline or EDTA. Irreversible inactivation of collagenase by chelators destroys its ability to incorporate 65Zn2+. 4. Latent collagenase, the inhibited form in which collagenase first appears in culture, behaves similarly to the active enzyme in 65Zn2+-exchange experiments, but is resistant to irreversible inactivation by chelators. 5. It is concluded that collagenase is a zinc metalloenzyme that forms an inactive and unstable apoenzyme on treatment with chelators. The bound inhibitor component of latent collagenase evidently stabilizes the apoenzyme.

Animals↗
PubMed2003

Therapeutic approaches to the modulation of apoptosis.

The appreciation of the role of apoptosis in the vast majority of diseases affecting humans has revolutionized the discovery and development of drugs targeting inflammation and oncology. Novel therapeutic approaches to modulate disease by regulating apoptosis are currently being tested in preclinical and clinical settings. Enthusiasm for some of these therapies is reflected in the fact that they have received U.S. Food and Drug Administration approval in record time. Approaches include the traditional use of small molecules to target specific players in the apoptosis cascade. They also include radical new approaches such as using antisense molecules to inhibit production of the Bcl-2 protein or antibodies that ligate either death receptors, such as TRAIL (tumour necrosis factor-related apoptosis-inducing ligand), or the MHC (HLA-DR), resulting in the initiation of apoptosis of target cells. Antibodies targeting cell-specific antigens are being used in conjunction with radioactive isotopes to deliver a more specific chemotherapy, particularly in the case of B-cell lymphomas. Other therapies target mitochondria, a key organelle in the apoptosis cascade. This diverse range of therapies includes photodynamic therapy, retinoic acid and arsenic trioxide, all of which induce apoptosis by generating reactive oxygen species. As our understanding of apoptosis increases, further opportunities will arise for tailor-made therapies that will result in improved clinical outcome without the devastating side effects of current interventions.

Animals↗
PubMed2003

Oxygen dependency of the adrenergic Na/H exchange in rainbow trout erythrocytes is diminished by a hydroxyl radical scavenger.

AIM: Potassium transport via the potassium chloride cotransporter in rainbow trout erythrocytes is increased by high oxygen tension. It appears that the effect of oxygen is mediated by reactive oxygen species, especially hydroxyl radicals. In contrast, the activity of adrenergically stimulated sodium proton exchange decreases with increasing oxygen tension. As available data suggest that the two transporters are regulated reciprocally, the present study was undertaken to evaluate, if hydroxyl radicals may inhibit sodium transport via the adrenergically stimulated sodium proton exchanger. METHODS: The effects of the hydroxyl radical scavenger, 2 mm mercaptopropionyl glycine (MPG), on the activity of the adrenergically activated sodium proton exchange in rainbow trout erythrocytes were examined by measuring unidirectional sodium flux, using radioactive isotope, and cellular water content. RESULTS: The activity of the sodium proton exchange increased with decreasing oxygen tension after adrenergic stimulation. When MPG was present during incubation, there was no statistically significant effect of oxygen tension on the adrenergically stimulated sodium proton exchange, whereby the activity of the transporter at atmospheric oxygen tension was markedly higher in the presence than in the absence of MPG. In the absence of adrenergic stimulation, MPG did not influence the transporter activity significantly at any oxygen tension. CONCLUSION: The data suggest that hydroxyl radicals are involved in the inhibition of the adrenergically stimulated sodium proton exchange at elevated oxygen tensions.

Animals↗
PubMed1999

Standardized intermittent static exercise increases peritendinous blood flow in human leg.

Alteration in tendinous and peritendinous blood flow during and after exercise is suggested to contribute to the development of Achilles tendon injury and inflammation. In the present study a method for evaluating the influence of standardized workload on peritendinous flow is presented. The radioactive isotope xenon-133 was injected just ventrally to the Achilles tendon 5 cm proximal to the tendon's insertion on the calcaneous. The disappearance of 133Xe was used to determine blood flow during intermittent static exercise of the calf muscle (1.5 s exercise/1.5 s rest) for 30 min at a workload equivalent to individual body weight (1 BW) in six healthy volunteers around both Achilles tendons (n = 12). During intermittent static exercise, blood flow was increased from 1.8 +/- 0.3 ml 100 g tissue-1 min-1 (mean value and SEM) (rest) to 6.1 +/- 1.3 ml 100 g tissue-1 min-1 (exercise) (P < 0.05). The exercise induced an average increase in blood flow (3.4-fold) equivalent to results previously obtained during regular dynamic heel raises (P > 0.05). It is concluded that the method is well suited to study the influence of standardized workload on the physiology and pathophysiology of the tissue around the Achilles tendon in humans.

Achilles Tendon↗
PubMed2003

Subcellular localization of boron in cultured melanoma cells by electron energy-loss spectroscopy of freeze-dried cryosections.

Boron neutron capture therapy (BNCT) is based on the ability of the non-radioactive isotope 10B to capture thermal neutrons and to disintegrate instantaneously. This reaction opens a way to selectively destroy tumour cells after specific uptake of 10B. In this paper, a method based on electron energy-loss spectroscopy is presented for detecting and quantifying boron in freeze-dried cryosections of human melanoma cells. A practical detection limit of around 6 mmol kg-1 in 0.1- micro m2 areas is estimated using specimens prepared from standard boron solutions. Preliminary results of boron mapping in the spectrum-imaging acquisition mode reveal boron penetration and probably spot-like accumulation within melanoma cells when exposed to culture medium containing sodium borocaptate.

Boron↗
PubMed1999

Biocompatible magnetic polymer carriers for in vivo radionuclide delivery.

The magnetic guidance of antiplastic and antibacterial agents as well as x-ray and MRI contrast substances in vivo by means of magnetic particles has been attempted repeatedly during the last 2 decades with more or less success. When using microparticles, the circulation time in the blood, the biodistribution, and to a greater or lesser extent, the specific targeting are determined by the uniformity of size, chemical composition, surface modification, and the electric surface charge. The electrophoretic mobility is an important parameter for the prediction of the usefulness of the prepared particle, modified by chemical and biological molecules. For its success, radionuclide therapy depends on the critical relationship between the amount of radioactive isotopes in the target tissue and in critical normal tissue. Because the implementation of radioimmunotherapy for the treatment of cancer has proven to be considerably more difficult than initially anticipated, we propose the use of magnetic nanospheres for the well directed delivery of radionuclides to a tumor after the intravenous administration of the biodegradable colloidal suspension.

Biocompatible Materials↗
PubMed2000

[19F-MRT of pulmonary ventilation in the breath-hold technic using SF6 gas].

OBJECTIVE: Development of a method to analyze lung ventilation by 19F-magnetic resonance imaging (MRI) of inspired SF6 gas during breath hold. MATERIAL AND METHODS: Measurements were performed with a Siemens Magnetom Vision 1.5 T scanner using the conventional gradient overdrive. Coronal images of the lung were acquired using ultrafast gradient-echo pulse sequences with TR/TE/alpha = 1.4 ms/0.48 ms/40 degrees without slice selection. With NEX = 200 averages and MA = 32 x 64 raw data matrix, the acquisition time was 9 s/image. Higher spatial resolution of 4.7 x 6.3 x 15 mm3 was obtained with a three-dimensional pulse sequence (TR/TE/alpha = 1.6 ms/0.48 ms/65 degrees, NEX = 20) running for 49 s. Measurements wer performed in three anesthetized and ventilated pigs (18 kg). RESULTS: A nearly linear relation between SF6 concentration and 19F signal intensity was observed. The signal-to-noise ratio in images obtained without slice selection was 30.9, with slice selection it was 14.9. No differences between SF6 distribution to both lungs were observed in the animals. CONCLUSION: Breath-hold MRI of SF6 gas distribution in the lung was demonstrated for the first time. The low spin-density was compensated for by highly repetitive signal averaging. Breath-hold 19F-MR imaging of ventilated airspaces to assess SF6 distribution in the human lung appears to be an interesting new method, which can be implemented with little technical efforts, and does not rely on radioactive isotopes.

Animals↗
PubMed1992

[Comparative DNA hybridization, cytology and histology of condylomatous and precancerous lesions of the cervix uteri].

Cell scrapings from 552 consecutive women attending the colposcopy outpatient unit were examined by radioactive isotopic DNA hybridisation using the Dot-Blot technique, and by conventional Papanicolaou-smear cytology. In 315 out of these patients, punch biopsies were submitted for histological examination. HPV DNA-sequences were detected in 72% (n = 167) of 232 cytologically diagnosed CIN cases and in 77% (n = 97) out of 126 histological diagnoses of CIN. The proportion of HPV positive cases increased proportionally to the grade of CIN. The frequency of HPV type 6/11 decreased proportionally to the grade of the CIN, and HPV type 16/18 increased at the same time. The most frequent HPV type was 16/18 (26.3%), followed by the HPV types 31/33/35 (16.3%) and 6/11 (4.2%). Furthermore, HPV positivity was detected in 16.5% of patients with negative cytologic smears, in 21.7% of those with negative histology and in 8.9% of women with both negative histology and cytology. Histological and moreover cytological criteria of HPV infection decreased dramatically and proportionally inverse to the grade of the CIN. These changes were not followed entirely by HPV positivity. The discrepancy between HPV positivity (66.1%/67.6%) and morphological evidence of HPV infection (cytology 4.9%, histology 21.6%) was higher in CIN III than in lower grade CIN. Neither cytology nor histology is able to detect all HPV-related cervical lesions. Hybridisation techniques are helpful in the detection of latent HPV infections as well as of CIN lesions have been missed either by cytology or by histology or both. Thus both morphological methods are supplemented by HPV hybridisation.(ABSTRACT TRUNCATED AT 250 WORDS)

Cervix Uteri↗
PubMed1984

[Visualization of malignant tumors in the abdomen using radiolabelled monoclonal antibodies].

The production of monoclonal antibodies by the hybrid technique introduced by Köhler and Milstein in 1975, led to new dimensions in the diagnosis and therapy of malignant diseases. Labelling such monoclonal antibodies, directed against tumour cells, by a radioactive isotope, makes them usable for visualising malignant tumours and their metastases. Early detection of cancer may represent a major step forward in cases where early detection is not possible with common noninvasive methods, such as ovarian cancer in gynaecological oncology. Radioimmunoscintigrams (RIS) were performed in seven patients, all of whom were suspect of having ovarian cancer. The RIS-results showed very good correlations with the operation reports. According to these first results we assume that the RIS method using I123-labelled monoclonal antibodies directed against tumour cells may be successful in the early detection of ovarian cancer. Other important information supplied by radioimmunoscanning is determination of tumour status, localisation, extent or metastatic spread.

Abdominal Neoplasms↗
PubMed1995

Endoscopic management of craniopharyngiomas: a review of 3 cases.

The authors describe the endoscopic management of cystic craniopharyngiomas in 3 cases. This method has been attempted in cystic craniopharyngiomas using a rigid endoscope. The instrument has been described earlier (7-9). All these operations were done under general anaesthesia. Criteria for endoscopic extirpation and removal included type D, E, F classification according to Yasargil (17). All 3 cases fitted in the F category. There were one female and two male patients. In the female patient an aspiration of cyst contents was performed as a first attempt to relieve her hydrocephalus. Two months later recurrent symptomology necessitated a larger intervention. All cysts were opened using the laser, drained by a Fogarthy balloon-catheter, and the capsule removed by forceps. This technique is safe and provides a reasonable alternative to open microsurgery, radioactive isotope instillation, or radiotherapy. In our series we achieved total removal in one case after the second intervention and partial removal in two cases. There was no mortality directly associated with this procedure and the female patient developed severe electrolyte disturbances after macroscopic total removal. Our results suggest that endoscopic of management of cystic craniopharyngiomas is a safe and effective procedure which could be considered as the initial management for cystic craniopharyngiomas of the intraventricular type.

Adult↗
PubMed1985

Biological effects of background radiation: mutagenicity of 40K.

The naturally occurring radioactive isotope 40K is the single largest contributor to the internal background radiation dose in living organisms. We examined cell growth and mutation rate or frequency in several strains of Escherichia coli in (i) media containing the natural content of 40K, (ii) media containing potassium from which essentially all of the 40K had been removed by isotope separation, and (iii) media highly enriched in 40K. Growth rates (doubling times) were identical in the present or absence of 40K. In more than 40 chemostat experiments, we were unable to detect any significant differences in mutation rate to bacteriophage T5 resistance or in mutation frequency to valine resistance or tryptophan prototrophy attributable to 40K. We conclude that, in the bacterial systems we have studied, 40K does not make a significant contribution to spontaneous mutation.

Dose-Response Relationship, Radiation↗
PubMed1997

Molecular and functional characterization of a novel low-affinity cation transporter (LCT1) in higher plants.

The transport of cations across membranes in higher plants plays an essential role in many physiological processes including mineral nutrition, cell expansion, and the transduction of environmental signals. In higher plants the coordinated expression of transport mechanisms is essential for specialized cellular processes and for adaptation to variable environmental conditions. To understand the molecular basis of cation transport in plant roots, a Triticum aestivum cDNA library was used to complement a yeast mutant deficient in potassium (K+) uptake. Two genes were cloned that complemented the mutant: HKT1 and a novel cDNA described in this report encoding a cation transporter, LCT1 (low-affinity cation transporter). Analysis of the secondary structure of LCT1 suggests that the protein contains 8-10 transmembrane helices and a hydrophilic amino terminus containing sequences enriched in Pro, Ser, Thr, and Glu (PEST). The transporter activity was assayed using radioactive isotopes in yeast cells expressing the cDNA. LCT1 mediated low-affinity uptake of the cations Rb+ and Na+, and possibly allowed Ca2+ but not Zn2+ uptake. LCT1 is expressed in low abundance in wheat roots and leaves. The precise functional role of this cation transporter is not known, although the competitive inhibition of cation uptake by Ca2+ has parallels to whole plant and molecular studies that have shown the important role of Ca2+ in reducing Na+ uptake and ameliorating Na+ toxicity. The structure of this higher plant ion transport protein is unique and contains PEST sequences.

Amino Acid Sequence↗
PubMed1998

Ionophores and receptors using cation-pi interactions: collarenes.

Cation-pi interactions are important forces in molecular recognition by biological receptors, enzyme catalysis, and crystal engineering. We have harnessed these interactions in designing molecular systems with circular arrangement of benzene units that are capable of acting as ionophores and models for biological receptors. [n]Collarenes are promising candidates with high selectivity for a specific cation, depending on n, because of their structural rigidity and well-defined cavity size. The interaction energies of [n]collarenes with cations have been evaluated by using ab initio calculations. The selectivity of these [n]collarenes in aqueous solution was revealed by using statistical perturbation theory in conjunction with Monte Carlo and molecular dynamics simulations. It has been observed that in [n]collarenes the ratio of the interaction energies of a cation with it and the cation with the basic building unit (benzene) can be correlated to its ion selectivity. We find that collarenes are excellent and efficient ionophores that bind cations through cation-pi interactions. [6]Collarene is found to be a selective host for Li+ and Mg2+, [8]collarene for K+ and Sr2+, and [10]collarene for Cs+ and Ba2+. This finding indicates that [10]collarene and [8]collarene could be used for effective separation of highly radioactive isotopes, 137Cs and 90Sr, which are major constituents of nuclear wastes. More interestingly, collarenes of larger cavity size can be useful in capturing organic cations. [12]Collarene exhibits a pronounced affinity for tetramethylammonium cation and acetylcholine, which implies that it could serve as a model for acetylcholinestrase. Thus, collarenes can prove to be novel and effective ionophores/model-receptors capable of heralding a new direction in molecular recognition and host-guest chemistry.

Benzene Derivatives↗