bluuebunny/crossref_metadata_2025_split huggingface dataset:
Characterization of insulin uptake into subcellular fractions of perfused rat liver using two different iodinated tracers
Authors: Christensen Jillian R. , Smith Geoffrey D. , Peters Timothy J. 
Published: May 2005

AbstractThe binding and uptake of insulin in perfused rat liver has been investigated with specifically labelled 125I‐A14‐tyrosyl insulin as a tracer and compared with a commercially available iodo‐insulin preparation. The commercial preparation did not show saturation uptake kinetics and the clearence from the perfusate remained low and constant throughtout a wide concentration range. A14 labelled insulin showed saturation kinetics and high clearence at low carrier concentration, falling rapidly with increasing carrier concentration and reaching a steady state value of 1 ml/min. These results emphasize the importance of using specifically labelled insulin in physiological and biochemical studies of hepatic insulin metabolism.Perfusion with A14 tyrosine‐labelled insulin at 4°C showed apparent saturation with binding to the plasma membrane fraction. Perfusion at 37°C also showed apparent saturation with uptake predominantly to the ligandosome fraction. These results implicate the plasma membrane‐ligandosome pathway in the hepatic uptake of insulin at both physiological and pharmacological concentrations of the hormone.
Incorporation of antigen 125I IgG into particualr cell compartments: Binding by chromatin
Authors: Rakowicz‐Szulczyńska Ewa M. , Horst Antoni 
Published: May 2005

AbstractIncorporation of [125I]IgG into spleen cells was studied in vivo and in vivo, the antigen after uptake into the cytoplasm migrated into cell nuclei, where it was bound to chromatin up to the saturation level. One day after immunization the constant level of [125I]IgG was 1·3 × 1012 molecules per spleen (108 cells). The same number of [125I]IgG molecules were bound to chromatin in cell cultures. The uptake of [125I]IgG was competitively inhibited by non‐labelled IgG. Binding of [125I]IgG molecules reextracted from cytoplasm and chromatin with specific anti‐human IgG serum argues against the uptake of degraded [125I]IgG molecules. [125I]IgG was tightly bound to DNA. Approximately 50 per cent of [125I]IgG was present in the residual chromatin fractin (after removal of 0·35 M and 2 M NaCl‐soluble frations) and 40 per cent was complexed with DNA (after removal of histones and non‐histones AP1, AP2, AP3 and AP4).Binding of [125I]IgG by isolated chromatin was inhibited by the cytoplasmic fraction but not by BSA. Binding of [125I]IgG by fractionated chromatin, (chromatins remaining after removal of 0·35M, and 2M NaCl‐soluble fractions or histones + non‐histones AP1 + AP2 + AP3 + AP4) occurred at a level similar to that observed with native chromatin. The results suggest that interaction of antigen with immunocompetent cells is not restricted to the cell surface but that antigen seems to be taken up into cytoplasm, migrates to the nuclei and is bound to chromatin, probably directly to DNA. The results are discussed in relation to the induction of the immune reaction.
Induction of plasma membrane alkaline phosphatase in rat liver
Authors: Bachs Oriol , Enrich Carles , Soriano Montserrat , Piñol Rosa , Domingo Jordi 
Published: May 2005

AbstractWe have determined alkaline phosphatase activity in total liver plasma membrane fractions from rats subjected to a partial hepatectomy and sham operated with or without manipulation of the liver. In all these cases, an increase of the enzyme activity was observed. Kinetic studies of alkaline phosphatase activity performed on plasma membrane fractions from rats subjected to a partial hepatectomy suggest that alkaline phosphatase increase is produced by de novo biosynthesis of enzyme molecules. Determination of alkaline phosphatase activity in purified plasma membrane subfractions corresponding to each of the three functional regions of the hepatocyte surface (blood sinusoidal, lateral and bile canalicular), indicates that the increase of the enzyme activity observed after partial hepatectomy is selectively induced in the bile canalicular domain of the hepatocyte plasma membrane.
Hormone‐Independent polyamine metabolism of squamous cell carcinoma of the prostate
Authors: Shukla Girja S. , Singhal R. L. 
Published: May 2005

AbstractThe levels of polyamines and their synthesizing enzymes in squamous cell carcinoma of prostate implanted in intact as well as castrated male rats were determined after certain hormonal manipulations. The tumour was found to grow with an identical rate in non‐castrated and castrated rats. Polyamine content and activities of polyamine synthesizing enzymes in the tumour were found to be much lower compared to their values in ventral prostate. Moreover, the levels of these parameters were comparable in tumours whether implanted in non‐castrated or gonadectomized animals. The sequential analyses of putrescine and spermidine and activities of L‐ornithine decarboxylase and S‐adenosyl‐L‐methionine decarboxylase of tumours at different time intervals showed a significant reduction in their levels at 30 days compared to 10 days post implantation in non‐castrated as well as castrated rats. Daily intramuscular administration of tumour‐bearing intact or castrated animals with testosterone (50 μg/g), β‐estradiol (2 μg/g) or cyproterone (12·5 μg/g) for 10 days did not influence polyamine metabolism in tumour tissue. However, either β‐estradiol and cyproterone treatments or castration were found to decrease polyamine synthesis in ventral prostate. At the same time, the testosterone replacement therapy did not allow polyamine levels or activities of polyamine synthesizing enzymes to decline in the ventral prostate of castrated rats. Our results demonstrated that contrary to ventral prostate, the polyamine metabolism in squamous cell carcinoma of prostate is independent of hormonal control. The loss of hormonal sensitivity of polyamine metabolism in the prostatic tumour could be the result of qualitative changes that occurred during transformation.
The interaction between gluconeogenic metabolism and accumulation of phosphate by chick kidney tubule cells
Authors: Grahn Michael F. , Parveen Riffat , Butterworth Peter J. 
Published: May 2005

AbstractPyruvate promotes both phosphate uptake and glucose synthesis by isolated chick kidney proximal tubule cells. 3‐Mercaptopicolinate inhibits both glucose synthesis and the promoted phosphate accumulation to the same extent. Glycerol also stimulates glucose synthesis, but does not affect phosphate accumulation. Oxygen utilization by the tissue is slightly stimulated by glycerol and pyruvate, but the enhancement of uptake by pyruvate is unlikely to result from raised cellular oxidative phosphorylation. The action of pyruvate is not a direct effect on the phosphate transporter, or on the transport of phosphate across the basolateral membrane, but entails an obligatory flux to triose phosphate.
Distribution of tightly bound non‐histone proteins in chromatin fractions obtained by DNAase II digestion
Authors: Lonigro Renata I. , Altieri Fabio , Allegra Paola , Caiafa Paola 
Published: May 2005

AbstractDigestion of pig liver chromatin with DNAse II afforded three different fractions which were characterized in terms of their DNA, RNA and tightly bound non‐histone protein content, their DNA fragment size and their template activity. Two of these fractions are soluble after digestion with DNAase II and have been separated on the basis of their different solubility in MgCl2. A third fraction is not solubilized even after extensive digestion, although the size of its DNA is comparable to that of the enzyme solubilized fractions. The three fractions show qualitatively and quantitatively different distribution of tightly bound non‐histone proteins, with specific protein components in each fraction; furthermore the non‐solubilized fraction is greatly enriched in proteins tightly bound to DNA. From all the data obtained it can be suggested that the tightly bound proteins of the insoluble fraction may play, directly or indirectly, a role in maintaining an organized chromatin structure.
Effect of tricyclohexylhydroxytin on synaptosomal Ca2+‐dependent ATP hydrolysis and rat brain subcellular calmodulin
Authors: Rao K. S. Prasada , Chetty C. S. , Trottman C. H. , Uzodinma J. E. , Desaiah D. 
Published: May 2005

AbstractEffect of tricyclohexylhydroxytin (plictran) on Ca2+‐ATPase activity was studied in rat brain synaptosomes under in vitro and in vivo conditions. Plictran inhibited basal Ca2+‐ATPase activity with an IC50 value of 6 nM suggesting its interaction with calcium transport phenomenon. Plictran inhibited calmodulin (CaM) activated Ca2+‐ATPase in a concentration‐dependent manner. A complete reversal of calmodulin activation of Ca2+‐ATPase was observed with 2–3 nM plictran. A 50 per cent decrease of CaM activated Ca2+‐ATPase was observed with 0·5 nM plictran, a concentration at which no significant effect was observed on basel enzyme activity. Of all the brain fractions studied, calmodulin levels in P2 fractions alone were reduced significantly to about 75 per cent of control values in plictran treated rats. The synaptosomal Ca2+‐ATPase was also decreased by 35 per cent, 42 per cent and 65 per cent in 10, 20 and 40 mg plictran kg−1 day−1 treated rats for 3 days respectively. The activity levels of Ca2+‐ATPase in 10 and 20 mg plictran kg−1day−1 treated rats were restored to normal level by exogenously added calmodulin. These results suggest that plictran may disrupt synaptic function by altering calcium and calmodulin regulated processes in the central nervous system.
Studies on rat liver mitochondria. 6. The effect of contaminating particles in mitochondria stored at 0–4°C
Authors: Higuti Ilma Hiroko , Murata Lilia Yuko , Stencel Mario , Campello Annibal De Paiva , Nascimento Aguinaldo José Do 
Published: May 2005

AbstractRat liver mitochondria were stored at 0–4°C for several days using an appropriate medium and energy source. The elimination of the majority of microsomes and lysosomes, that normally contaminate isolated mitochondria, had a positive effect in preservation of respiratory control, P:O ratio, and monoamine oxidase activity during long term storage.
Is the saccharide component of the insulin receptor involved in hormonal imprinting?
Authors: Kovács P. , Csaba G. 
Published: May 2005

AbstractHormonal imprinting takes place at the first interaction of a given hormone with the cell (the Tetrahymena in the present case) and accounts for a greater responsiveness to the hormone on re‐exposure(s). The Tetrahymena is able to bind insulin and Concanavalin‐A (Con‐A) as well. Exposure to both ligands—simultaneously or in sequence—enhances the binding of both in the progeny generations. It follows that the lectin, which inhibits insulin binding by direct action, enhances rather than depresses the effect of insulin‐induced imprinting.
Biochemical significance of enhanced activity of fluorinated 1,25‐dihydroxyvitamin D3 in human cultured cell lines
Authors: Eisman John A. , Frampton Richard J. , McLean Fiona J. 
Published: May 2005

AbstractSeveral human cancer cells possess receptors for 1,25‐dihydroxyvitamin D3[1,25‐(OH)2D3]. In these cells 1,25‐(OH)2D3 has a biphasic concentration‐dependent regulatory effect on cell replication and specifically induces its own metabolism. We have studied the effects on these parameters of the native hormone together with those of two analogues fluorinated at the 24‐carbon and of 1,24R,25‐trihydroxyvitamin D3[1,24R,25‐(OH)3D3]. The difluorinated analogue 24,24‐difluoro‐1,25‐(OH)2D3[24,24‐F2‐1,25‐(OH)2D3] is an approximately fivefold more potent inhibitor of cellular replication than the native hormone, while 1,24R,25‐(OH)3D3 is about fivefold less potent. This enhanced potency of the fluorinated analogue parallels its enhanced potency in in vivo studies of its effects on calcium and mineral metabolism. However, although the analogue retains replication stimulatory activity, it is clearly no more potent than the native hormone in this activity: 1,24R,25‐(OH)3D3 has no significant stimulatory activity. Exposure of the cells to 1,25‐(OH)2D3 at 0·05 nM for 6h increases the subsequent conversion of labelled hormone to aqueous phase soluble compounds by 6·7‐fold. None of the other compounds had a similar effect at this concentration. At 10nM all 1‐hydroxylated compounds increased aqueous phase radioactivity about equally (13 to 17‐fold); this effect is still specific since 25‐OH D3 had no such effect even at 10nM. Studies on the effects of the fluorinated analogues upon receptor binding of hormone in cell cytosols and uptake of hormone by intact cells clearly demonstrate that the enhanced activity of these analogues is not due to higher receptor affinity or more rapid access to intracellular receptor. These data suggest that this enhanced inhibitory potency, observed in these human cancer cells and by analogy their enhanced potency in vivo, is due to their resistance to metabolism by 24‐hydroxylation. In view of our previous observations that 24‐hydroxylation is part of the induced metabolic pathway, these data support the hypothesis that 24‐hydroxylation is part of an inactivation pathway in these human target cells.
Effect of Escherichia coli lipopolysaccharide on the microviscosity of liver plasma membranes and hepatocyte suspensions and monolayers
Authors: Portolés M. T. , Pagani R. , Díaz‐Laviada I. , Municio A. M. 
Published: May 2005

AbstractThe fluorescence probe 1,6‐diphenylhexa‐1,3,5‐triene (DPH) was used for monitoring structural perturbations induced by lipopolysaccharide (LPS) of Escherichia coli (0111 : B4) in plasma membranes of rat liver. Changes in microviscosity were observed in plasma membrane preparations from control rats after treatment with LPS and in plasma membrane preparations from liver perfused with LPS. In both systems fluorescence polarization was measured from which microviscosity was calculated. This parameter increases with LPS treatment. From temperature dependence studies was inferred that LPS interaction with plasma membrane preparations induces an increase of both the polarization term (r0/r–1)−1 and flow activation energy (ΔE). Addition of LPS to hepatocyte suspensions also induces an increase on microviscosity and a delay in the fall of microviscosity induced by a temperature rise in hepatocyte monolayers grown on microcover slides.These data suggest that LPS interaction can be attributed to its binding to membrane hydrophobic regions in a non‐specific manner.
Inhibition by elastase inhibitors of the formyl met leu phe‐induced chemotaxis of rat polymorphonuclear leukocytes
Authors: Hornebeck W. , Soleilhac J. M. , Tixier J. M. , Moczar E. , Robert L. 
Published: May 2005

AbstractRat leukocyte elastase has been purified successively by AH‐Sepharose Kappa‐elastin affinity chromatography and by ion exchange chromatography on a carboxymethyl Sephadex resin. It has great similarities with human leukocyte elastase in its molecular weight, substrate specificity and inhibitory profile.The effect of rat leukocyte elastase inhibitors in influencing the chemotactic response of rat PMN to fMetLeuPhe has been compared to that of other proteinase inhibitors. The results indicated that oleoyl (Ala)2ProValCH2Cl, a specific inhibitor of human and rat leukocyte elastases and Eglin C, which also inhibits human and rat cathepsin G, are among the powerful inhibitors of rat PMN chemotaxis induced by the formyl oligopeptide. This suggests that these neutral proteinases, in addition to their known participation in connective tissue catabolism, could play a role in PMN locomotion and chemotaxis.
Experimental conditions influence [3H]‐dihydroalprenolol binding characteristics to living HeLa cells due to morphological changes: A warning
Authors: Rademaker Ben , Stroes Jan W. , Bast Aalt , Timmerman Henk 
Published: May 2005

AbstractHarvesting of plated growing HeLa cells, followed by incubation of these cells without any addition at 37°C was found to cause changes in the cell shape. This phenomenon is accompanied by a diminished binding of the beta‐adrenergic antagonist [3H]‐dihydroalprenolol and the alpha‐adrenergic antagonist phentolamine to a binding compartment not representing beta‐adrenergic receptors. These binding sites have a high affinity for hydrophobic agents and most probably represent lipophilic structures in the cellular membrane. Changes in the cell shape obviously cause alterations in the physical properties of the plasma membrane. This might lead to misinterpretations of the results from experiments in which the redistribution of beta‐adrenergic receptors is followed during incubation with agonists, as receptor occupation with subsequent receptor redistribution is possibly accompanied by effects on the membrane microviscosity.It is concluded that investigations performed in order to follow physiological events like receptor redistribution and desensitization processes, may be obfuscated by changes in the normal physical state of the living cells.
Immunohistochemical localization of albumin and in situ hybridization of albumin mRNA
Authors: Shroyer Kenneth R. , Nakane Paul K. 
Published: May 2005

AbstractHepatocytes actively involved in albumin synthesis were identified by immunohistochemical method. In sections of perioidate–lysine–2 per cent (w/v) paraformaldehyde fixed normal rat liver, albumin was detected in all hepatocytes. At the untrastructural level, albumin was localized in the rough endoplasmic reticulum and in Golgi complexes located near the nucleus in only a small subpopulation of hepatocytes, while all other hepatocytes contained albumin only in Golgi complexes located near the bile canaliculi. Stimulation of albumin synthesis by puromycin aminonucleoside‐induced nephrosis resulted in an altered intracellular distribution of albumin at the light microscopic level. When examined at the ultrastructural level, albumin was localized in the rough endoplasmic reticulum as well as in Golgi complexes located near the nucleus in nearly all these hepatocytes.Hepatocytes with the potential to synthesize albumin were identified by in situ hybridization of albumin mRNA. In sections of 0·1 per cent (v/v) glutaraldehyde perfusion fixed normal rat liver, albumin mRNA was detected in the cytoplasm of only a few hepatocytes scattered throughout the lobule. Following stimulation of albumin synthesis by the induction of nephrosis, albumin mRNA was detected in the cytoplasm of the hepatocytes.The source of albumin in those hepatocytes which lacked albumin mRNA was identified in analbuminemic rats injected with rat albumin. At 6 h post injection, the light microscopic distribution of albumin in the liver of these animals was virtually indistinguishable from that in normal rat liver. At the ultrastructural level, injected albumin was localized in lysosomes and in Golgi complexes located near the bile canaliculi.
Differential time‐course of induction of rat liver gamma‐glutamyltransferase and drug‐metabolizing enzymes in the endoplasmic reticulum, golgi and plasma membranes after a single phenobarbital injection. Evaluation of protein variations by two‐dimensional electrophoresis
Authors: Antoine B. , Rahimi‐Pour A. , Siest G. , Magdalou J. , Galteau M. M. 
Published: May 2005

AbstractThis study was conducted to follow as a function of time the activity of gamma‐glutamyltransferase in the various membranes of rat liver cells after a single dose of phenobarbital (PB) (75 mg kg−1 body weight). Gamma‐glutamyltransferase induction was maximal 24 h after PB treatment in both the rough endoplasmic reticulum and the plasma membranes. This pattern of induction differed from that of some drug metabolizing enzymes. While total cytochrome P‐450 content was enhanced mainly in endoplasmic reticulum until 48 h after PB treatment, UDP‐glucuronosyltransferase activity was not greatly altered by PB under the same conditions.The comparison of two‐dimensional electrophoretic polypeptide profiles of each subcellular membrane isolated from control and phenobarbital‐treated rats revealed important variations induced by PB. In plasma membranes, the heaviest subunit (apparent Mr = 60 × 103) of hepatic gamma‐glutamyltransferase was provisionally identified as a collection of polypeptides which differ only by their pI. The concentration of these polypeptides was smaller in the endoplasmic reticulum where they were of lower apparent molecular mass. This suggests that the gamma‐glutamyltransferase precursor is already processed at the level of the endoplasmic reticulum but it is still not completely mature or glycosylated. Five days of continuous PB treatment induced the appearance of new gamma‐glutamyltransferase isoforms in plasma membranes.We demonstrate that after a single injection of PB, gamma‐glutamyltransferase activity increases simultaneously with some drug‐metabolizing enzymes, such as total cytochrome P‐450 but not with others, such as UDP‐glucuronosyltransferases.
Δ53β hydroxysteroid dehydrogenase activity of the rat corpus luteum exhibits positive substrate‐binding co‐operativity: A continuous monitoring microdensitometric study with unfixed ovarian tissue sections
Authors: Gordon M. , Robertson W. R. 
Published: May 2005

AbstractΔ53β hydroxysteroid dehydrogenase activity transforms biologically inactive Δ53β hydroxy steroids into the active Δ43‐keto products (e.g. pregnenolone to progesterone). Using a cytochemical procedure which allows for the continuous microdensitometric monitoring of an enzyme reaction as it proceeds and a well described cytochemical assay for Δ53β HSD we have analysed the initial velocity rates (Vo) for dehydroepiandrosterone (DHEA) binding to this enzyme in regressing (i.e. 20α hydroxy steroid dehydrogenase positive) corpus luteum (CL) cells in unfixed tissue sections (5 μm) of the dioestrous and proestrous rat ovary. The results are mean ± S.E.M. The relationship between DHEA concentration (0 to 50 μM) and Δ53β HSD activity in the dioestrous corpora lutea was sigmoidal and had an atypical 1/Vo versus 1/S plot, the x intercept being positive. Using a 1/Vo versus 1/S2 plot the Vmax was determined to be 1·0 ± 0·08 μmol min−1 mg−1 CL (n = 6). The Hill constant was 2·7 ± 0·02 (n = 6) suggesting a high degree of positive co‐operativity for DHEA binding. The S concentration for half maximal activity was 17 ± 1 μmoles (n = 6). In the corpora lutea cells of the proestrous ovary, the Vmax for DHEA transformation was unchanged (0·95 ± 0·04 μmol min−1 mg−1, n = 3) whilst the S0·5 was significantly increased to 27 ± 0·1 (p < 0·01, n = 3). The Hill constant remained positive being 2·9 ± 0·2 (n = 3).NAD+ binding to 3β HSD in regressing corpora lutea of the proestrous ovary has been demonstrated previously to be hyperbolic and fit the classical Michaelis‐Menten model.1 Extending the analysis of NAD+ binding to the regressing corpus luteum of the dioestrous rat ovary revealed similar kinetic characteristics to that seen with the proestrous enzyme, the apparent Vmax and Km being 0·84 ± 0·04 μmol min−1 mg−1 CL (n = 3) and 27 ± 7 μmol 1−1 (n = 3) respectively. The Hill constant was 1·1 ± 0·03 (n = 3), indicating no co‐operativity of co‐factor binding.