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PubMed1997

Reversible dementia in elderly patients referred to a memory clinic.

Dementia has a reversible cause in some cases, and these should be diagnosed without over-investigating the many patients with irreversible disease. We prospectively studied the prevalence of reversible dementia in a memory clinic, determined the added value of investigations compared with clinical examination and assessed the outcome of treatment of potentially reversible causes by measuring (1) cognition, (2) disability in daily functioning, (3) behavioural changes and (4) caregiver burden. Two hundred patients aged 65 years and over were examined, using the CAMDEX-N. If they were demented, the probable cause was diagnosed clinically and confirmed or excluded by a standard set of investigations, which were done in all patients. Of the patients, 170 (mean age 79.2 years) were demented; 31 were treated for potentially reversible causes. At follow-up after 6 months, no patients showed complete reversal of dementia. Five patients improved on clinical impression, but only one on clinical measurement. Thirty patients were cognitively impaired, but not demented; seven were treated. Judged clinically, three patients improved, but on assessment only one did so; she recovered completely. Blood tests often produced diagnostic results that were not expected clinically, but electroencephalography and computed tomography of the brain did not. None of the investigations had an effect on outcome of dementia after treatment. We conclude that in elderly patients referred to a memory clinic, the prevalence of reversible dementia is of the order of 1%, if outcome after treatment is assessed by a standardized measurement. We recommend blood tests in all patients, to detect not only metabolic causes of dementia but also co-morbidity possibly worsening the dementia. Other investigations can be performed on clinical indication. Clinical evaluation remains the mainstay of diagnosis in dementia.

Aged↗
PubMed1999

Clozapine reversal of the deficits in coordinated movement induced by D2 receptor blockade does not depend upon antagonism of alpha2 adrenoceptors.

Alpha2 adrenoceptor antagonists have been shown to reverse D2-antagonist-induced catalepsy leading to the hypothesis that the alpha2 antagonistic properties of clozapine underlie the compound's lack of extrapyramidal symptoms in the clinic. The potential for alpha2 antagonists to reverse the motor deficits produced by D2 antagonists (loxapine and haloperidol) was further investigated using a rotating rod (3.5 rpm) test in male Sprague-Dawley rats that requires coordinated movement to perform the task. The effects of loxapine (0.3 mg/kg, s.c.) were dose-dependently and statistically significantly reversed by the administration of clozapine (1,3, 10 mg/kg, i.p., n=10). Isoloxapine (1 mg/kg, i.p.), RX 821002 (2-methoxy-idazoxan; 5.6 mg/kg, i.p.) and yohimbine (5.6 mg/kg, i.p.) did not reverse the effects of loxapine. Furthermore, the motor deficits produced by haloperidol could not be reversed by RX 821002 (5.6 mg/kg, i.p.) or yohimbine (5.6 mg/kg, i.p.). On the other hand, scopolamine (0.03-0.3 mg/kg, i.p.) dose-dependently and statistically significantly antagonised the effects of both loxapine and haloperidol. These results indicate that the anticholinergic rather than the alpha2 antagonistic properties of clozapine may mediate the reversal of the motor deficit induced by D2 antagonism in a rotating rod test.

Adrenergic alpha-2 Receptor Antagonists↗
PubMed2000

Pharmacological reversal of behavioral and cellular indices of cocaine sensitization in the rat.

RATIONALE AND OBJECTIVES: Behavioral sensitization has been proposed as an animal model for the intensification of drug craving in cocaine addiction. Interactions between dopamine and glutamate systems are important for the induction and maintenance of sensitization. The goal of this study was to determine if established cocaine sensitization could be reversed by pharmacological manipulation of these transmitter systems. METHODS: Rats received 15 mg/kg cocaine (IP) on days 1-10 and were challenged with cocaine (10 mg/kg) on day 13 to verify that sensitization had occurred. On days 14-20, separate groups of sensitized rats received daily injections of dopamine D1- or D2-class agonists, an NMDA receptor antagonist, or a dopamine agonist with an NMDA antagonist. Three days or 2 weeks later, all rats were again tested for their response to cocaine to determine if sensitization had been reversed. RESULTS: Reversal of sensitization was produced by repeated administration of either a D1-class agonist (SKF 81297) or the combination of an NMDA receptor antagonist and a D2-class agonist. Effective combinations were cocaine+MK-801, quinpirole+MK-801, quinpirole+CGS 19755, and pergolide+memantine. The latter drugs are approved for human use. Reversal of sensitization persisted for at least 2 weeks after cessation of drug treatment. Electrophysiological studies revealed that these drug treatments also reversed dopamine D1 receptor supersensitivity in the nucleus accumbens, a cellular correlate of sensitization. CONCLUSIONS: These results demonstrate that pharmacotherapy can reverse behavioral and cellular adaptations associated with repeated cocaine administration, and may do so without the need for continued medication.

Animals↗
PubMed2004

Muscle activation patterns in point-to-point and reversal movements in healthy, older subjects and in subjects with Parkinson's disease.

When young, healthy subjects perform rapid point-to-point and reversal movements over a range of distances, the patterns of muscle activation associated with accelerating the limb toward the target are modulated in the same way for both movement tasks. Differences in patterns of muscle activation for these two movement types are not observed until the deceleration phase of the movements. In this study, we first test the hypothesis that healthy, older subjects and subjects with Parkinson's disease will modulate the pattern of muscle activation in the same way during the acceleration phase of point-to-point and reversal elbow movements. Second, we test the hypothesis that healthy, older subjects and subjects with Parkinson's disease exhibit the same relationship in muscle activation patterns between the two movement types that have been observed for the young in the deceleration phase of the movements. Subjects performed point-to-point and reversal movements initiated in the direction of flexion over three distances (36, 54 and 72 degrees) "as fast as possible". Angle, velocity, acceleration and surface EMGs from biceps and triceps were recorded. With respect to the first hypothesis, the EMG, kinetic, and kinematic measures related to the acceleration phase of the movements were modulated in the same way for both movement types in the healthy older subjects. In the Parkinson's disease group, the kinematic and kinetic measures during the acceleration phase of the movements were the same in both movement types; however, the flexor and extensor EMG activation was smaller during reversal movements than during point-to-point movements. With respect to the second hypothesis, in contrast to that found in young subjects, in healthy older subjects, there was no significant difference between the movement types in the flexor EMG activity immediately after the time of peak velocity. This difference between younger and older subjects may be attributed to the fact that older subjects perform both movement types more slowly than do younger subjects. Although subjects with Parkinson's disease also move slowly, the flexor EMG shuts off more abruptly and more completely just after the time of peak velocity during reversal movements than during point-to-point movements. These results show that (1) for healthy subjects, when the task requirements are the same for the two movement types (acceleration phase), muscle activation patterns are modulated in the same way, and (2) both age and disease alter the relationship of muscle activation, kinetics and kinematics between point-to-point and reversal movements.

Aged↗
PubMed2005

Catalytic activity of lignin peroxidase and partition of veratryl alcohol in AOT/isooctane/toluene/water reverse micelles.

The activity of lignin peroxidase (LiP) and the partition of its optimum substrate veratryl alcohol (VA) in sodium bis(2-ethylhexyl)sulfosuccinate (AOT)/isooctane/toluene/water reverse micelles were studied in this paper to understand the microheterogeneous effect of the medium on the catalytic properties of LiP hosted in the reverse micelle. Results showed that LiP from Phanerochaete chrysosporium could express its activity in the reverse micelles, but its activity depended, to a great extent, on the composition of the reverse micelles. Optimum activity occurred at a molar ratio of water to AOT (omega0) of 11, a pH value of 3.6, and a volume ratio of isooctane to toluene of 7-9. Under optimum conditions, the half-life of LiP was circa 12 h. The dependence of LiP activity on the volume fraction of water in the medium (theta), at a constant omega0 value of 11, indicated that VA was mainly solubilized in the pseudophase of the reverse micelle. Based on the pseudobiphasic model and the corresponding kinetic method, a linear line can be obtained in a plot of apparent Michaelis constant of VA vs theta, and the partition coefficient of VA between the pseudophase and the organic solvent phase was determined to be 35.8, which was higher than that (22.3) between bulk water and the corresponding mixed organic solvent. H2O2 inhibited LiP at concentrations higher than 80 microM; this concentration value seems to be different from that in aqueous solution (about 3 mM). The differences mentioned above should be ascribed to the microheterogeneity and the interface of the AOT reverse micelle.

Benzyl Alcohols↗
PubMed2004

Reversal of multidrug resistance of cancer through inhibition of P-glycoprotein by 5-bromotetrandrine.

PURPOSE: The present study aimed to evaluate the MDR reversal activity of bromotetrandrine (BrTet), a bromized derivative of tetrandrine (Tet), in vitro and in vivo. METHODS: Drug sensitivity was determined using the MTT assay. The in vivo effect of Tet was investigated using nude mice grafted with sensitive and resistant KB human epidermoid cancer cells. Doxorubicin (Dox) accumulation was analyzed by fluorospectrophotometry and the protein and mRNA levels of P-glycoprotein (P-gp) were determined by immunocytochemistry and RT-PCR, respectively. RESULTS: BrTet at 0.25, 0.5 and 1 micro M reversed Dox resistance in MDR human breast cancer MCF-7/Dox cells dose-dependently and its potency was greater than that of Tet at the same concentrations. BrTet reversed vincristine (VCR), Dox and paclitaxel resistance in MDR human oral epidermoid carcinoma KBv200 cells as well as innate VCR and Dox resistance in human hepatocellular carcinoma Bel(7402) cells. However, BrTet showed no effect on the IC(50) values of the above-mentioned anticancer drugs in sensitive MCF-7 and KB cells. No reversal effect of BrTet on the cytotoxicity of 5-fluorouracil and cisplatin, non-P-gp substrates, was observed. In nude mice bearing KBv200 xenografts on the left flank and KB xenografts on the right flank, i.p. injection of 5 mg/kg and 10 mg/kg BrTet significantly enhanced the antitumor activity of Dox against KBv200 xenografts with inhibitory rates of 33.0% and 39.2%, while Dox alone inhibited the growth of KBv200 xenografts by only 11.6%. No enhancement by BrTet was seen in KB xenografts. Moreover, BrTet at 5 mg/kg reversed paclitaxel resistance in KBv200 xenografts. Fluorospectrophotometric assay showed that BrTet significantly increased the intracellular accumulation of Dox in MCF-7/Dox cells in a dose-dependent manner. BrTet also inhibited the overexpression of P-gp in MCF-7/Dox cells, but had no effect on mdr1 expression. CONCLUSIONS: BrTet showed significant MDR reversal activity in vitro and in vivo. Its activity may be related to the inhibition of P-gp overexpression and the increase in intracellular accumulation of anticancer drugs. BrTet may be a promising MDR modulator for eventual assessment in the clinic.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1999

The bisindolylmaleimide protein kinase C inhibitor, Ro 32-2241, reverses multidrug resistance in KB tumour cells.

Ro 32-2241 is a bisindolylmaleimide that selectively inhibits protein kinase C (PKC) as compared with other protein kinases. Experiments were carried out to examine its potential as a multidrug resistance-reversing agent. Ro 32-2241 inhibited efflux, and increased accumulation, of [3H]-daunomycin in multidrug-resistant (MDR) KB-8-5 and KB-8-5-11 cells and had no effect on drug-sensitive KB-3-1 cells. Ro 32-2241 completely reversed the doxorubicin resistance of KB-8-5 and KB-8-5-11 cells, showing no effect on the sensitivity of drug-sensitive KB-3-1 cells. The potency of Ro 32-2241 was comparable with that of cyclosporin A and better than that of verapamil, known modulators of multidrug resistance. Ro 32-2241 also completely reversed the taxol resistance of KB-8-5 cells and partially reversed the resistance of KB-8-5-11 cells. Vinblastine resistance was also partially reversed. Mechanistic experiments were carried out to determine whether Ro 32-2241 interacted with P-glycoprotein (Pgp) directly. Increased efflux of [14C]-Ro 32-2241 was seen with the more resistant KB-8-5-11 cells (although the percentage effluxed was very low as compared with [3H]-daunomycin), suggesting that Ro 32-2241 can act as a substrate for Pgp. Direct interaction of Ro 32-2241 with Pgp was confirmed by demonstration that it inhibited binding of [3H]-azidopine to Pgp in KB-8-5-11 membranes. In conclusion, Ro 32-2241, acting directly on Pgp (rather than, or in addition to, an effect on PKC), is effective in reducing or reversing resistance to doxorubicin, taxol and vinblastine in human tumour cells with a clinically relevant degree of MDR. However, results of in vivo experiments conducted to investigate the effects of Ro 32-2241 on resistance to doxorubicin suggest that it may not be possible to achieve sufficiently high levels of Ro 32-2241 in vivo to modulate MDR.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1997

New multidrug-resistance-reversing drugs, MS-209 and SDZ PSC 833.

The emergence of multidrug resistance (MDR) is a major problem in cancer chemotherapy. Many compounds have been developed to reverse MDR, and some of them are undergoing clinical trials. Among them, MS-209, a novel quinoline derivative, is one of the most potent MDR-reversing agents: MS-209 at 3 microM effectively reverses MDR in various cell lines in vitro. MS-209 directly interacts with P-glycoprotein (Pgp) and inhibits Pgp-mediated drug transport. Oral administration of MS-209 combined with anticancer drugs significantly increases the life span of mice bearing MDR tumor cells without causing serious side effects. SDZ PSC 833, a non-immunosuppressive analogue of cyclosporin A (CsA), is another potent MDR-reversing drug. Interestingly, the MDR-reversing activity of SDZ PSC 833 is enhanced in vitro and in vivo by MRK-16, a monoclonal antibody that recognizes an extracellular epitope of Pgp. Since MRK-16 promotes immune responses to MDR tumor cells expressing Pgp, the combined use of MRK-16, SDZ PSC 833, and antitumor drugs could be an effective therapeutic modality to reverse MDR.

Animals↗
PubMed2001

Neuronal control of heart reversal in the hawkmoth Manduca sexta.

Cardiograms demonstrate that heart activity of Manduca sexta changes from larva, to pupa, to adult. The larval heart has only anterograde contractions. During metamorphosis, heart activity becomes a cyclic alternation of anterograde and retrograde contractions. Thus, the adult heart has both an anterograde and a retrograde pacemaker. External stimuli also can initiate cardiac reversal. Cardiac reversal is blocked by tetrodotoxin, indicating that reversal is under neuronal control. A branch of each dorsal nerve 8 innervates the posterior chamber of the heart, the location of the anterograde pacemaker. Only retrograde contractions occur when dorsal nerves 8 are cut. Stimulation of ml(-1) 8 initiates anterograde contractions; when stimulation ceases, the heart reverses to retrograde contractions. These experiments indicate that the anterograde pacemaker receives neural input that makes it the dominant pacemaker. In the absence of neural input this pacemaker is inactive, and the retrograde pacemaker becomes active. Application of crustacean cardioactive peptide accelerates the heart but does not eliminate cardiac reversal. The terminal chamber of the heart is also innervated by a branch of each dorsal nerve 7; stimulation of this nerve increases the strength of contraction of the terminal chamber but has no effect on contractions of the remainder of the heart or on cardiac reversal.

Animals↗
PubMed2003

Reverse transcriptase-related proteins in telomeres and in certain chromosomal loci of Rhynchosciara (Diptera: Sciaridae).

The localization of reverse transcriptase-related proteins in polytene chromosomes of dipterans was investigated using previously characterized antibodies to a recombinant polypeptide containing conserved motifs of insect reverse transcriptases. The immunoreactions were carried out with polytene chromosome squashes of eight sciarids, one chironomid and three Drosophila species. Telomeric staining was regularly observed on chromosomes of the sciarid Rhynchosciara americana under normal growth conditions. Five of eight chromosomal tips were labelled except for the heterochromatic ends that are occasionally found associated forming a chromocentre in the salivary gland. Reverse transcriptase-related proteins were detected at chromosomal tips of young larvae and remained bound to the telomeres throughout larval development. As in salivary gland chromosomes, five non-telocentric ends of the chromosomes from Malpighian tubules of R. americana appeared clearly stained with anti-reverse transcriptase. The occurrence of telomeric reverse transcriptase in R. americana correlates with the presence of RNA in addition to an unusual enrichment with homopolymeric dA/dT DNA associated with the telomeric heterochromatin. The antibodies also reacted with a few interstitial sites in chromosomes of four Rhynchosciara species, one band overlapping the histone gene locus of three species in the americana -like group. The results provide evidence for a reverse transcriptase-related protein as a constitutive component in telomeres of R. americana and also in certain interstitial loci of Rhynchosciara species in which RNA was immunologically detected in the form of RNA:DNA hybrids.

Animals↗
PubMed2000

Alpha activity decreases during the perception of Necker cube reversals: an application of wavelet transform.

Since the first observation of perceptual reversal by Necker, many theoretical approaches have been proposed. In a previous study, we showed that a positive wave appeared approximately 250 ms prior to the button press of the subjects, indicating perceptual reversal during the observation of the Necker cube figure. A basic difficulty in this type of study is the possible jitter in the latency of the button press due to the variability of the subjects' reaction time during a recording session. To overcome this difficulty, a pattern selection method based on the wavelet transform was proposed in the previous study. A dominant positive wavelet coefficient in the delta band was found to represent the perceptual-reversal-related positivity. In the present study, we aim to analyze the changes in the alpha frequency band during perceptual reversal by using the Necker cube. The RMS values of the alpha frequency band were measured for two time periods: +/- 3 SD around the mean peak latency of the perceptual-reversal-related positivity and a time window of the same length before the positive wave. We found significantly increased delta power and decreased alpha power during the perceptual-reversal-related positivity.

Adult↗
PubMed2006

Reversed actrocytic GLT-1 during ischemia is crucial to excitotoxic death of neurons, but contributes to the survival of astrocytes themselves.

During ischemia, the operation of astrocytic/neuronal glutamate transporters is reversed and glutamate and Na(+) are co-transported to the extracellular space. This study aims to investigate whether this reversed operation of glutamate transporters has any functional meanings for astrocytes themselves. Oxygen/glucose deprivation (OGD) of neuron/astrocyte co-cultures resulted in the massive death of neurons, and the cell death was significantly reduced by treatment with either AP5 or DHK. In cultured astrocytes with little GLT-1 expression, OGD produced Na(+) overload, resulting in the reversal of astrocytic Na(+)/Ca(2+)-exchanger (NCX). The reversed NCX then caused Ca(2+) overload leading to the damage of astrocytes. In contrast, the OGD-induced Na(+) overload and astrocytic damage were significantly attenuated in PACAP-treated astrocytes with increased GLT-1 expression, and the attenuation was antagonized by treatment with DHK. These results suggested that the OGD-induced reversal of GLT-1 contributed to the survival of astrocytes themselves by releasing Na(+) with glutamate via reversed GLT-1.

Animals↗
PubMed2004

Identification of shareholder ethics and responsibilities in online reverse auctions for construction projects.

The increasing number of companies providing internet services and auction tools helped popularize the online reverse auction trend for purchasing commodities and services in the last decade. As a result, a number of owners, both public and private, accepted the online reverse auctions as the bidding technique for their construction projects. Owners, while trying to minimize their costs for construction projects, are also required to address their ethical responsibilities to the shareholders. In the case of online reverse auctions for construction projects, the ethical issues involved in the bidding technique directly reflects on the owner's ethical and social responsibilities to their shareholders. The goal of this paper is to identify the shareholder ethics and responsibilities in online reverse auctions for construction projects by analyzing the ethical issues for the parties involved in the process. The identification of the ethical issues and responsibilities requires clear definition and understanding of professional ethics and the roles of the involved parties. In this paper, first, the concept of professional ethics and social responsibility is described in a general form. To illustrate the ethical issues and responsibilities, a sample case of bidding for a construction project using online reverse auction techniques is presented in which the shareholders were actively involved in questioning the ethical issues. The issues involved in the bidding process and their reflection on the shareholder responsibilities are described and analyzed for each stage of the process. A brief discussion of the overall process is also included to address the general ethical issues involved in online reverse auctions.

Competitive Bidding↗
PubMed1987

Comparisons between diastolic fluttering and reverse doming of anterior mitral leaflet in aortic regurgitation.

In 27 patients with aortic regurgitation and nine patients with associated aortic stenosis, the incidence of reverse doming was studied by two-dimensional echocardiography and was compared with diastolic fluttering of the mitral valve. Reverse doming could be identified in four of six patients (66.7%) with mild aortic regurgitation and in 20 of 21 patients (95.2%) with moderate or severe aortic regurgitation (p less than 0.05). Fluttering of the anterior mitral leaflet was observed in all patients with mild aortic insufficiency and in 19 of 21 patients (90.5%) with moderate or severe aortic insufficiency. In patients with associated aortic stenosis, reverse doming was observed in three patients (33.3%), whereas fluttering was seen in six patients (66.7%). After aortic valve replacement, reverse doming persisted in one patient with a paravalvular leak. Reverse doming or fluttering was not seen in healthy volunteers or in 10 patients with congestive cardiomyopathy. Our data indicate that reverse doming is a helpful parameter for the diagnosis of aortic regurgitation.

Adolescent↗
PubMed1988

Use of reverse transcription to determine the exact locations of psoralen photochemical crosslinks in RNA.

We have studied the properties of avian myeloblastosis virus reverse transcriptase on Escherichia coli 16S ribosomal RNA that was known to contain a psoralen crosslink in a restricted region around residue 920. These crosslinked RNA molecules were purified on the basis of loop size by gel electrophoresis. Reverse transcription stopped at specific positions in the crosslinked molecules but not in control linear molecules. With the particular crosslink that was studied, at the earliest time of reverse transcription, the most frequent stopping site was G925. At later times two nearly equal stops at G925 and C924 were seen. The crosslinked site was an absolute stop since even at long times of incubation the reverse transcriptase was not able to proceed beyond G925/C924. An independent determination of the crosslinked site by directly sequencing a section of the crosslinked RNA indicated that C924 was the sole nucleotide involved in the crosslink. Therefore, reverse transcriptase was able to synthesize cDNA all the way up to and including the nucleotide that contained the psoralen crosslink. Thus, reverse transcription can be a rapid and precise method for determining psoralen crosslinking sites when prefractionated, crosslinked RNA templates are used.

Avian Myeloblastosis Virus↗
PubMed1992

Exogenous primer-independent cDNA synthesis with commercial reverse transcriptase preparations on plant virus RNA templates.

Upon reverse transcription and cloning manipulations with virion RNAs of several plant viruses, namely beet yellows virus, brome mosaic virus, and potato virus X, we came across a significant background synthesis of cDNA on the virion RNA template in vitro independent of exogenous primers added. When tested with beet yellow virus RNA template, several commercial preparations of avian myeloblastosis virus (AMV) reverse transcriptase showed the background activity monitored by the [alpha-32P]dNTP incorporation in vitro, while the enzyme from murine moloney leukemia virus (MMLV) was found strictly exogenous-primer-dependent. To detect possible nucleic acid contaminations in reverse transcriptase, the enzyme preparations from several commercial sources were incubated with [gamma-32P]ATP and polynucleotide kinase. The labeled material from AMV reverse transcriptase preparations comigrated with a tRNA marker in polyacrylamide gels and was found to be RNase-sensitive. The MMLV reverse transcriptase preparations were free from such a contamination. These results indicate that the exogenous-primer-independent cDNA synthesis by some AMV reverse transcriptases could be due to a contaminating tRNA (or its low-molecular-weight degradation products) serving as an endogenous primer.

Artifacts↗
PubMed1986

Uncoupler-inhibitor titrations of ATP-driven reverse electron transfer in bovine submitochondrial particles provide evidence for direct interaction between ATPase and NADH:Q oxidoreductase.

From the chemiosmotic hypothesis it follows that no change is expected in potency of an uncoupler to inhibit an energy-driven reaction in an energy-transducing membrane if the energy-requiring part of the reaction, the so-called secondary proton pump, is partially inhibited by a specific, tightly bound inhibitor. An increase in potency upon inhibition of the primary pump may be expected, due to a lower rate of the total proton flow that can be used by the secondary pump and dissipated by the uncoupler. Contrary to this prediction several uncouplers (S13, SF6847, 2,4-dinitrophenol, valinomycin + nigericin) show an increase in uncoupling efficiency in ATP-driven reverse electron transfer (reversal) upon inhibition of the secondary pump in this reaction, the NADH:Q oxidoreductase, by rotenone. The increase in uncoupling efficiency is proportional to the decrease in the rate of reversal, that is to the decrease in concentration of active secondary pump. Similarly, upon inhibition of the primary pump, the ATPase, with oligomycin, an increase in uncoupling efficiency was found, also proportional to the decrease in the rate of reversal. When the pore-forming uncoupler gramicidin was used, no change in uncoupling potency was found upon inhibition of NADH:Q oxidoreductase. Inhibition of the ATPase, however, resulted in a proportionally lower uncoupling titre for gramicidin, just as was found for S13 in the presence of oligomycin. A difference was also found in the relative concentrations of S13 and gramicidin required to stimulate ATP hydrolysis or to inhibit reversal. The amount of S13 needed to stimulate ATP hydrolysis was clearly higher than the amount needed to inhibit reversal. On the contrary, the titre of gramicidin for both actions was about the same. To explain these results we propose that gramicidin uncouples via dissipation of the bulk delta mu H+, whereas the carrier-type uncouplers preferentially interfere with the direct energy transduction between the ATPase and redox enzymes. This is in accordance with the recently developed collision hypothesis.

Adenosine Triphosphatases↗
PubMed1993

A pool of reversibly cell-associated cholesteryl esters involved in the selective uptake of cholesteryl esters from high-density lipoproteins by Hep G2 hepatoma cells.

Selective uptake of high-density lipoprotein (HDL) cholesteryl esters without parallel uptake of HDL apolipoproteins occurs by a non-endocytotic pathway that results in net delivery of cholesteryl esters to cells. With respect to the cellular mechanism of this pathway, previous studies with adrenal cells showed a cholesteryl ester pool that is reversibly associated with cells and which appears to mediate irreversible selective uptake. A cholesteryl ester pool with similar properties was observed in plasma membranes isolated from adrenal cells, suggesting that this is the site of the cellular pool. Human Hep G2 hepatoma cells also selectively take up HDL cholesteryl esters. Therefore we asked if these cells have a reversibly cell-associated cholesteryl ester pool as well that could mediate irreversible selective uptake. To do this, human HDL3 (d = 1.125-1.21 g/ml) was labeled in both its protein and cholesteryl ester moieties. Uptake of HDL3 tracers by Hep G2 cells was then studied. After an uptake incubation in the presence of labeled HDL3, either cellular uptake of tracers was immediately determined or cells were 'chase' incubated in the presence of unlabeled HDL before determination of cellular tracer content. Hep G2 cells selectively took up HDL3 cholesteryl esters under these conditions. However, a fraction of cholesteryl ester tracer selectively taken up was chased from the cells by subsequent incubation in the presence of unlabeled HDL. This reversible pool of cholesteryl ester tracer was distinct from that irreversibly internalized, and in excess of that accounted for by dissociation of labeled HDL3 particles bound to the cell surface. Selective uptake was down-regulated by prior incubation with LDL, and cholesteryl ester tracer in the reversible pool was down-regulated in parallel. Plasma membranes were isolated from Hep G2 cells and incubated with doubly labeled HDL3. HDL3 particles bound to these membranes, as indicated by the apolipoprotein tracer. However, HDL cholesteryl esters associated with plasma membranes in excess on that accounted for by HDL3 particles. This selective association of HDL3 cholesteryl ester tracer with membranes was reversible, and the tracer was chased during incubation in the presence of unlabeled HDL. These results suggest that, as with steroidogenic cells, a reversible pool of cholesteryl esters localized in the plasma membrane is involved in selective uptake of HDL3 cholesteryl esters by hepatic cells at a step prior to irreversible internalization.

Animals↗