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Query: UNIPROT:P50583 (asymmetrical)
 12,197 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

A large number of hepatoma cell lines has been used to study expression and regulation of liver-specific function. However these cells, even the most differentiated, are morphologically far from hepatocytes. In no case is the typical hepatocyte cell polarity well maintained. Cell hybridization has been used as a potential means for turning on specific genes. From hybrids between well differentiated Fao rat hepatoma cells and WI 38 human fibroblasts, we have attempted to isolate segregated cells that are highly differentiated and polarized. Such cells, detected in aged cultures of only one hybrid (WIF12), were isolated by subcloning. One subclone, WIF12-1 was analyzed. Expression of liver-specific functions extinguished in the original hybrid is restored in all WIF12-1 cells at a very high level, similar to that of hepatocytes and 5-30 times higher that that of parental cells. Moreover human genes coding for liver-specific proteins (albumin, fibrinogen, and alcohol dehydrogenase) are actively expressed. WIF12-1 cells have acquired a polarized phenotype as attested by the presence of bile canaliculi between adjacent cells and by the asymmetrical localization of apical (Mg(2+)-ATPase, gamma-glutamyl transpeptidase) and basolateral membrane markers. The bile canaliculi formed are dynamic and functional structures, characterized by long periods of expansion followed by rapid contractions. The ability to polarize is a general and permanent property of WIF12-1 cells. These cells appear to constitute a valid model for the in vitro study of hepatocyte cell polarity, membrane domain formation and mechanisms of membrane protein sorting.
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PMID:Hybrid cell lines constitute a potential reservoir of polarized cells: isolation and study of highly differentiated hepatoma-derived hybrid cells able to form functional bile canaliculi in vitro. 195 80

The performance of the asymmetrical flow field-flow fractionation channel has been improved by the use of a much thinner (0.12-mm) channel than before and by flow programming (stepwise gradient elution). The thinner channel contributes to a decreased zone broadening which enables complete resolution in a shorter time. Three protein peaks, representing molecular weights from 12,000 to 136,000, wer completely resolved within 3 min. Flow programming speeds up the elution of the high-molecular-weight materials which occur late in a fractogram. This enabled separation of a small plasmid fragment (700 base pairs) from the large amounts of a big fragment (4600 base pairs) in 30 min and improved detection of a presumed trimer of albumin. Two viruses (1.8 10(6) and 50 10(6) daltons) were eluted as narrow peaks within 5 min.
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PMID:Improved separation speed and efficiency for proteins, nucleic acids and viruses in asymmetrical flow field flow fractionation. 277 88

The transport of radiolabelled albumin from tissue to blood was measured with an external detection technique in isolated, maximally vasodilated rat skeletal muscles. Initially, rat hindlimbs were perfused with albumin-serum solutions containing [99mTc]albumin for at least 2 h, during which time the tracer accumulated interstitially. The accumulated tracer albumin was then washed out over a period of 1 h, using a tracer-free, otherwise identical, perfusate. The wash-out curve was multi-exponential and the last 30-min period was used to calculate the turnover rate constant (k), which was 7.5 x 10(-4) min-1, (+/- 0.7 x 10(-4), n = 5). Moreover, if albumin was assumed to be distributed homogeneously within the interstitium, with a distribution volume (Vi) of 10 ml 100 g-1, a tissue-to-blood clearance of albumin (ClT-B) of 0.0075 ml min-1 100 g-1 could be calculated. By this approach ClT-B is probably slightly overestimated, but is still only 30% of the clearance from blood to tissue (ClB-T), as determined in several previous studies under similar conditions. Thus, transcapillary passage of albumin is highly asymmetrical, being at least three times greater from blood to tissue than in the opposite direction. This is in agreement with the concept of the capillary walls being composed of two populations of functional pores, where macromolecules are transported from blood to tissue mainly by convection through large pores, even at low filtration rates.
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PMID:Diffusional transport of albumin from interstitium to blood across small pores in the capillary walls of rat skeletal muscle. 322 5

Mycoplasma gallisepticum was adapted to grow with delta 5-sterols modified in the aliphatic side chain, and stopped-flow kinetic measurements of filipin association were made to estimate the sterol distribution between the two leaflets of the membrane. Cholesterol derivatives with unsaturated side chains (desmosterol, cis- and trans-22-dehydrocholesterol, and cholesta-5,22E,24-trien-3 beta-ol) or an alkyl substituent (beta-sitosterol) were predominantly (86-94%) localized in the outer leaflet of the bilayer. However, cholesterol, 20-isocholesterol, and sterols with side chains of varying lengths (in the 20(R)-n-alkylpregn-5-en-3 beta-ol series where the alkyl group ranged from ethyl to undecyl) were distributed nearly symmetrically between the two halves of the bilayer. Kinetic measurements of beta-[14C]sitosterol and [14C]desmosterol exchange between M. gallisepticum cells and an excess of sonicated sterol/phosphatidylcholine vesicles confirmed the filipin-binding studies. More than 90% of these radiolabeled sterols underwent exchange at 37 degrees C with unlabeled sterols in vesicles over a period of 12-14 h in the presence of 2% (w/v) albumin. beta-[14C]Sitosterol exchange was characterized by biphasic exchange kinetics, indicative of two pools of sitosterol molecules in the cell membrane. Only a single kinetic pool was detected for [14C]desmosterol exchange. Stopped flow measurements of filipin binding to beta-sitosterol and stigmasterol also revealed an asymmetrical localization of these sterols in membranes of growing Mycoplasma. capricolum cells. When an early exponential culture of beta-sitosterol- or stigmasterol-adapted M. capricolum was transferred to a sterol-rich medium at 37 degrees C, approximately three-quarters of the beta-sitosterol or stigmasterol was localized in the outer leaflet after growth was continued for 6 h; in contrast, cholesterol was distributed symmetrically after about 1 h. The asymmetric localization of sterols with alkylated or unsaturated side chains suggests that growth-supporting sterols need not be translocated extensively into the inner leaflet of the bilayers of M. gallisepticum and M. capricolum.
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PMID:Distribution and movement of sterols with different side chain structures between the two leaflets of the membrane bilayer of mycoplasma cells. 670 46

Nerve terminals as well as glial cells are thought to possess high-affinity Na(+)-dependent transport sites for excitatory amino acids. However, recent immunocytochemical results with antibodies against such a transporter isolated from rat brain showed a selective labelling of glial cells [Danbolt et al. (1992) Neuroscience 51, 295-310]. Critical evaluation of the literature indicates that previous evidence for nerve terminal uptake of acidic amino acids might possibly be attributed to glia. To find out whether there is indeed a glutamate transporter in nerve endings, we incubated hippocampal slices with D-aspartate (10 and 50 microM), a metabolically inert substrate for the high-affinity glutamate transport system. After fixation by glutaraldehyde/formaldehyde the slices were processed immunocytochemically with specific polyclonal antibodies raised against D-aspartate coupled to albumin by glutaraldehyde/formaldehyde. The electron-microscopic postembedding immunogold technique demonstrated a large accumulation of gold particles in nerve terminals making asymmetrical synapses, compared to their postsynaptic dendritic spines, as well as in glial cell processes. The labelled terminals include those of the glutamatergic Schaffer collaterals. Axosomatic boutons appeared unlabelled. Comparison with a test conjugate with known concentration of fixed D-aspartate (94 mM) suggests that the concentration attained in the terminals after incubation with 50 microM D-aspartate was in the lower millimolar range. The uptake was totally dependent on Na+, blocked by L-threo-3-hydroxyaspartate, and had a high affinity for D-aspartate (apparent Km about 20 microM). There was no labelling in slices incubated without D-aspartate. Compared to glia, the nerve terminals had a higher D-aspartate density and accounted for a much higher proportion of the total tissue uptake, but this relationship may be different in vivo. At the light-microscopic level the D-aspartate-like immunoreactivity showed a distinct laminar distribution, identical to that shown autoradiographically for D-[3H]aspartate and L-[3H]glutamate uptake sites [Taxt and Storm-Mathisen (1984) Neuroscience 11, 79-100], and corresponding to the terminal fields of the major excitatory fibre systems in the hippocampal formation. The novel approach described here establishes that glutamatergic nerve terminals as well as glia do sustain sodium-dependent high-affinity transport of excitatory amino acids, implying that more than one glutamate transporter must be present in the brain. Immunogold detection of D-aspartate gives a much higher anatomical resolution than electron microscopic autoradiography of D-[3H]aspartate or L-[3H]glutamate uptake, the only method that has been available previously for ultrastructural demonstration of uptake activity.(ABSTRACT TRUNCATED AT 400 WORDS)
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PMID:Demonstration of glutamate/aspartate uptake activity in nerve endings by use of antibodies recognizing exogenous D-aspartate. 790 57

Trichloroacetic acid (TCA) is a contaminant of drinking water. It induces peroxisome proliferation in livers of rats and mice and is hepatocarcinogenic in the latter species. Previous experimental studies of the kinetics of TCA in the isolated perfused rat liver (IPRL) at two doses have been reported. To gain more insight into the mechanistic processes controlling TCA kinetics in the liver a biologically based kinetic (BBK) model for the IPRL was used to analyze the experimental data. The IPRL was exposed to 25, 250, or 1000 microM TCA for 2 h in a recirculating perfusion system. These doses were not cytotoxic. The BBK model simulated the TCA concentration in perfusion medium and liver, and the biliary excretion of TCA. Separate protein binding studies showed that over 90% of TCA was bound to albumin in the perfusion medium whereas binding in liver homogenate was much lower. Integrating the information on protein binding into the BBK model, the hepatic uptake of TCA and its biliary excretion could be fitted assuming asymmetrical saturable transport at the sinusoidal membrane and linear transport at the bile canalicular membrane. To validate the BBK model, additional washout experiments were conducted in which the perfusion medium was replaced with TCA-free medium after 30 min of exposure of the liver to 1000 microM TCA. This approach illustrates the usefulness of BBK modeling for analyzing experimental kinetic data and gaining insight in kinetic mechanisms controlling the behavior of a chemical in the liver.
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PMID:Simulation of trichloroacetic acid kinetics in the isolated perfused rat liver using a biologically based kinetic model. 1238 32

During fetal life, there are periods of rapid cell proliferation, which are uniquely sensitive to nutritional perturbation. Feeding the pregnant rat a protein-restricted diet alters the growth trajectory of major fetal organs such as the kidney. By day 21 of gestation, the ratio of kidney weight to total body weight is reduced in the fetuses of dams fed a protein-deficient diet. In contrast, the ratio of fetal liver weight to total body weight is unchanged. To investigate the mechanisms underlying this disproportionate change in organ growth in the low-protein group, cell proliferation and differentiation have been assessed in the liver and kidney. The steady-state levels of mRNA for the growth-arrest and DNA-damage gene gadd153/CHOP-10, CCAAT enhancer-binding proteins alpha and beta were unaffected by maternal diet in both fetal liver and kidney. The mRNA for alpha-fetoprotein, albumin and hepatic glucokinase were unchanged in the liver, suggesting that maternal protein deficiency does not alter the state of differentiation. The steady-state levels of the mRNA coding for the cyclin-dependent protein kinase inhibitors (p15(INK4a), p19(INK4d), p21(CIP1), p27(KIP1) and p57(KIP2)) were unchanged in the fetal livers but were significantly increased in the kidneys of fetuses from dams fed the low-protein diet. These results show that the asymmetrical growth of the kidney is associated with increases in mRNA for the Cip/Kip cyclin-dependent kinase inhibitors and that these may reflect specific lesions in organ development.
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PMID:The expression of growth-arrest genes in the liver and kidney of the protein-restricted rat fetus. 1611 27

To evaluate the contribution of neural pathways to the determination of the circadian oscillator phase in peripheral organs, we assessed lateralization of clock gene expression in Syrian hamsters induced to split rhythms of locomotor activity by exposure to constant light. We measured the ratio of haPer1, haPer2, and haBmal1 mRNA on the high vs. low (H/L) side at 3-h intervals prior to the predicted activity onset (pAO). We also calculated expression on the sides ipsilateral vs. contralateral (I/C) to the side of the suprachiasmatic nucleus (SCN) expressing higher haPer1. The extent of asymmetry in split hamsters varied between specific genes, phases, and organs. Although the magnitude of asymmetry in peripheral organs was never as great as that in the SCN, we observed significantly greater lateralization of clock gene expression in the adrenal medulla and cortex, lung, and skeletal muscle, but not in liver or kidney, of split hamsters than of unsplit controls. We observed fivefold lateralization of expression of the clock-controlled gene, albumin site D-element binding protein (Dbp), in skeletal muscle (H/L: 10.7 +/- 3.7 at 3 h vs. 2.2 +/- 0.3 at 0 h pAO; P = 0.03). Furthermore, tyrosine hydroxylase expression was asymmetrical in the adrenal medulla of split (H/L: 1.9 +/- 0.5 at 0 h) vs. unsplit hamsters (1.2 +/- 0.04; P < 0.05). Consistent with a model of neurally controlled gene expression, we found significant correlations between the phase angle between morning and evening components (psi(me)) and the level of asymmetry (H/L or I/C). Our results indicate that neural pathways contribute to, but cannot completely account for, SCN regulation of the phase of peripheral oscillators.
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PMID:Lateralization of the central circadian pacemaker output: a test of neural control of peripheral oscillator phase. 2059 76

Intravascular tracer washout data obtained from gastrocnemius muscle of lean Zucker rats (LZRs) and obese Zucker rats (OZRs) were analysed to investigate flow distributions in the OZR, a model of non-atherosclerotic peripheral vascular disease. A computer model used to simulate the network washout curves was developed based on experimentally observed relative dispersions in large vessels and asymmetrical flow distributions at bifurcations in dichotomous microvascular networks. The model results of simulations were compared to experimental washout data of (125)I-labelled albumin, an intravascular tracer, to uncover flow distributions on the arterial-network and capillary levels. The lean and obese Zucker rats demonstrated distinct capillary-level flow distributions, with higher dispersion and significantly more low-flow capillaries in the OZRs than in the LZRs. Targeted pharmacological treatments against identified sites of vascular dysfunction in OZRs (adrenoreceptor blockade with phentolamine, antioxidant treatment with Tempol and thromboxane receptor antagonism with SQ-29548) were shown to improve the capillary-level flow distributions in treated OZRs toward distributions determined in control LZRs. Combination therapy with multiple pharmacological interventions resulted in a greater degree of recovery. This study demonstrates that the enhanced perfusion heterogeneity at arteriole bifurcations is a potential mechanism underlying perfusion-demand mismatching in OZRs, and suggests that amelioration of this dysfunction must involve a multi-faceted interventional approach.
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PMID:Computational analyses of intravascular tracer washout reveal altered capillary-level flow distributions in obese Zucker rats. 2178 50

Nutritional factors such as magnesium, folic acid, vitamins B12 and B6, L-arginine, and polyunsaturated fatty acids (PUFAs) appear to be significantly beneficial for patients with coronary artery disease (CAD), and in the prevention and arresting the progression of HF and cardiac arrhythmias. Additionally, ingestion of adequate amounts of protein and maintaining normal concentrations of plasma albumin seem to be essential for these patients. These nutrients closely interact with the metabolism of L-arginine-nitric oxide (NO) system, essential fatty acids, and eicosanoids such that beneficial products such as NO, prostaglandin E1, prostacyclin, prostaglandin I3, lipoxins, resolvins, and protectins are generated and synthesis of proinflammatory cytokines is suppressed that results in platelet anti-aggregation, vasodilation, angiogenesis, and prevention of CAD, cardiac arrhythmias, and stabilization of HF. This implies that individuals at high risk for CAD, cardiac arrhythmias, and HF and those who have these diseases need to be screened for plasma levels of magnesium, folic acid, vitamins B12 and B6, L-arginine, NO, various PUFAs, lipoxin A4, resolvins, protectins, asymmetrical dimethylarginine (an endogenous inhibitor of NO), albumin, and various eicosanoids and cytokines and correct their abnormalities to restore normal physiology.
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PMID:Nutritional factors in the prevention and management of coronary artery disease and heart failure. 2559 5


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