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George H Rothblat - One of the best experts on this subject based on the ideXlab platform.

  • use of cyclodextrins for manipulating cellular cholesterol content
    Journal of Lipid Research, 1997
    Co-Authors: A E Christian, M P Haynes, Michael C Phillips, George H Rothblat
    Abstract:

    Previous studies from this laboratory have demonstrated that exposure of tissue culture cells to cyclodextrins results in rapid cholesterol depletion. In the present study, we have developed experimental systems for using solutions of cyclodextrins, either 2-hydroxypropyl Beta-Cyclodextrin or methylated Beta-Cyclodextrin, complexed with varying amounts of free cholesterol to manipulate cell cholesterol content. Cholesterol delivered via the cyclodextrin has been found to be metabolically active, as measured by the acyl-coenzyme A:cholesterol acyltransferase (ACAT)-mediated esterification of [3H]cholesterol in Fu5AH rat hepatoma cells and Chinese hamster ovary cells. The methylated Beta-Cyclodextrin was found to be a more efficient donor in all cell types studied, with an average cholesterol uptake of at least 100 microg cholesterol/mg protein within 6 h. By modifying the cyclodextrin:cholesterol molar ratio, it is possible to manipulate the cellular cholesterol content of cells, producing conditions ranging from net cholesterol enrichment to depletion. The use of cyclodextrins provides a convenient, precise and reproducible method for modulating the cholesterol content of tissue culture cells.

  • cyclodextrins as catalysts for the removal of cholesterol from macrophage foam cells
    Journal of Clinical Investigation, 1997
    Co-Authors: V Atger, Michael C Phillips, Genevieve W Stoudt, M De La Llera Moya, W V Rodrigueza, George H Rothblat
    Abstract:

    Low concentrations of cyclodextrins (< 1.0 mM) added to serum act catalytically, accelerating the exchange of cholesterol between cells and lipoproteins. J774 macrophages incubated with serum and 2-hydroxypropyl-Beta-Cyclodextrin (< or = 1 mM) released fivefold more labeled cholesterol than with serum alone. Increased efflux was not accompanied by a change in cell cholesterol mass; thus, cyclodextrin functioned as a cholesterol shuttle, enhancing cholesterol bidirectional flux without changing the equilibrium cholesterol distribution between cells and medium. The addition of phospholipid vesicles to serum and cyclodextrin shifted the equilibrium distribution to favor the medium, producing rapid and extensive depletion of cell cholesterol mass. The combination of serum, phospholipid vesicles, and cyclodextrin also stimulated the rapid clearance of both free and esterified cholesterol from mouse peritoneal macrophages loaded with free and esterified cholesterol. This study: (a) demonstrates that a compound can function as a catalyst to enhance the movement of cholesterol between cells and serum, (b) illustrates the difference between cholesterol exchange and net transport in a cell/serum system, (c) demonstrates how net movement of cholesterol is linked to concentration gradients established by phospholipids, (d) provides a basis for the development of the shuttle/sink model for the first steps in reverse cholesterol transport, (e) validates the model using artificial shuttles (cyclodextrins) and sinks (large unilamellar vesicles), and (f) suggests that cyclodextrin-like cholesterol shuttles might be of pharmacological significance in treating unstable atherosclerotic plaques.

  • cellular cholesterol efflux mediated by cyclodextrins
    Journal of Biological Chemistry, 1995
    Co-Authors: E P C Kilsdonk, Michael C Phillips, Patricia G Yancey, Genevieve W Stoudt, Faan Wen Bangerter, William J Johnson, George H Rothblat
    Abstract:

    In this study, we compared cholesterol efflux mediated by either high density lipoproteins (HDL3) or Beta-Cyclodextrins, cyclic oligosaccharides that are able to dissolve lipids in their hydrophobic core. Beta-Cyclodextrin, 2-hydroxypropyl-Beta-Cyclodextrin, and methyl-Beta-Cyclodextrin at 10 mM induced the release of 50-90% of L-cell [3H]cholesterol after 8 h of incubation, with a major portion of this cholesterol being released in the first 1-2 h of incubation. The cholesterol efflux kinetics are different if cells are incubated with HDL3, which induces a relatively constant rate of release of cholesterol throughout an 8-h incubation. Cholesterol efflux to cyclodextrins was much greater than phospholipid release. To test the hypothesis that maximal efflux rate constants for a particular cell are independent of the type of acceptor, we estimated the maximal rate constants for efflux (Vmax) of cellular cholesterol from L-cells, Fu5AH cells, and GM3468A fibroblasts. The rate constant for HDL3-mediated efflux varied among cell lines in the order Fu5AH > L-cells > fibroblasts. However, these differences were not evident when cyclodextrins were used as cholesterol acceptors. The estimated Vmax values for cyclodextrin-mediated efflux were 3.5-70-fold greater than for HDL3 for the three cell lines. The very high efficiency of cyclodextrins in stimulating cell cholesterol efflux suggests that these compounds can be used in two general ways for studies of atherosclerosis: 1) as research tools to probe mechanisms of cholesterol transport and aspects of membrane structure or 2) as potential pharmacological agents that could modify in vivo cholesterol metabolism and influence the development of the atherosclerotic plaque.

Lajos Szente - One of the best experts on this subject based on the ideXlab platform.

  • protective effects of beta cyclodextrins vs zearalenone induced toxicity in hela cells and tg vtg1 mcherry zebrafish embryos
    Chemosphere, 2020
    Co-Authors: Zelma Faisal, Lajos Szente, Edina Garai, Rita Csepregi, Katalin Bakos, Eszter Fliszarnyul, Adrienn Balazs, Matyas Cserhati, Tamas Kőszegi, Bela Urbanyi
    Abstract:

    Abstract Zearalenone is a xenoestrogenic mycotoxin produced by Fusarium species. High exposure with zearalenone induces reproductive disorders worldwide. Cyclodextrins are ring-shaped host molecules built up from glucose units. The apolar cavity of cyclodextrins can entrap so-called guest molecules. The formation of highly stable host-guest type complexes with cyclodextrins can decrease the biological effect of the guest molecule. Therefore, cyclodextrins may be suitable to decrease the toxicity of some xenobiotics even after the exposure. In this study, the protective effect of Beta-Cyclodextrins against zearalenone-induced toxicity was investigated in HeLa cells and zebrafish embryos. Fluorescence spectroscopic studies demonstrated the formation of stable complexes of zearalenone with sulfobutyl-, methyl-, and succinyl-methyl-substituted Beta-Cyclodextrins at pH 7.4 (K = 1.4–4.7 × 104 L/mol). These chemically modified cyclodextrins considerably decreased or even abolished the zearalenone-induced loss of cell viability in HeLa cells and mortality in zebrafish embryos. Furthermore, the sublethal effects of zearalenone were also significantly alleviated by the co-treatment with Beta-Cyclodextrins. To test the estrogenic effect of the mycotoxin, a transgenic bioindicator zebrafish model (Tg(vtg1:mCherry)) was also applied. Our results suggest that the zearalenone-induced vitellogenin production is partly suppressed by the hepatotoxicity of zearalenone in zebrafish. This study demonstrates that the formation of stable zearalenone-cyclodextrin complexes can strongly decrease or even abolish the zearalenone-induced toxicity, both in vitro and in vivo. Therefore, cyclodextrins appear as promising new mycotoxin binders.

  • Removal of Zearalenone and Zearalenols from Aqueous Solutions Using Insoluble Beta-Cyclodextrin Bead Polymer
    MDPI AG, 2018
    Co-Authors: Miklós Poór, Zelma Faisal, Afshin Zand, Tímea Bencsik, Beáta Lemli, Sándor Kunsági-máté, Lajos Szente
    Abstract:

    Zearalenone (ZEN) is a Fusarium-derived mycotoxin, exerting xenoestrogenic effects in animals and humans. ZEN and its derivatives commonly occur in cereals and cereal-based products. During the biotransformation of ZEN, its reduced metabolites, α-zearalenol (α-ZEL) and β-zearalenol (β-ZEL), are formed; α-ZEL is even more toxic than the parent compound ZEN. Since previous studies demonstrated that ZEN and ZELs form stable complexes with β-cyclodextrins, it is reasonable to hypothesize that cyclodextrin polymers may be suitable for mycotoxin removal from aqueous solutions. In this study, the extraction of ZEN and ZELs from water, buffers, and corn beer was investigated, employing insoluble β-cyclodextrin bead polymer (BBP) as a mycotoxin-binder. Our results demonstrate that even relatively small amounts of BBP can strongly decrease the mycotoxin content of aqueous solutions (including beer). After the first application of BBP for mycotoxin binding, BBP could be completely reactivated through the elimination of ZEN from the cyclodextrin cavities by washing with a 50 v/v% ethanol-water mixture. Therefore, our study suggests that insoluble cyclodextrin polymers may be suitable tools in the future to deplete mycotoxins from contaminated drinks

  • fluorescently labeled methyl beta cyclodextrin enters intestinal epithelial caco 2 cells by fluid phase endocytosis
    PLOS ONE, 2014
    Co-Authors: Ferenc Fenyvesi, Lajos Szente, Katalin Retinagy, Zsolt Bacso, Zsuzsanna Gutaytoth, Milo Malanga, Eva Fenyvesi, Judit Varadi, Zoltan Ujhelyi, Palma Feher
    Abstract:

    Cyclodextrins are widely used excipients for increasing the bioavailability of poorly water-soluble drugs. Their effect on drug absorption in the gastrointestinal tract is explained by their solubility- and permeability-enhancement. The aims of this study were to investigate penetration properties of fluorescently labeled randomly methylated-Beta-Cyclodextrin (FITC-RAMEB) on Caco-2 cell layer and examine the cellular entry of cyclodextrins on intestinal cells. The permeability of FITC-RAMEB through Caco-2 monolayers was very limited. Using this compound in 0.05 mM concentration the permeability coefficient was 3.35±1.29×10−8 cm/s and its permeability did not change in the presence of 5 mM randomly methylated-Beta-Cyclodextrin. Despite of the low permeability, cellular accumulation of FITC-RAMEB in cytoplasmic vesicles was significant and showed strong time and concentration dependence, similar to the characteristics of the macropinocytosis marker Lucifer Yellow. The internalization process was fully inhibited at 0°C and it was drastically reduced at 37°C applying rottlerin, an inhibitor of macropinocytosis. Notably, FITC-RAMEB colocalized with the early endosome organizer Rab5a. These results have revealed that FITC-RAMEB is able to enter intestinal epithelial cells by fluid-phase endocytosis from the apical side. This mechanism can be an additional process which helps to overcome the intestinal barrier and contributes to the bioavailability enhancement of cyclodextrins.

  • Fluorescently Labeled Methyl-Beta-Cyclodextrin Enters Intestinal Epithelial Caco-2 Cells by Fluid-Phase
    2014
    Co-Authors: Ferenc Fenyvesi, Milo Malanga, Lajos Szente
    Abstract:

    Cyclodextrins are widely used excipients for increasing the bioavailability of poorly water-soluble drugs. Their effect on drug absorption in the gastrointestinal tract is explained by their solubility- and permeability-enhancement. The aims of this study were to investigate penetration properties of fluorescently labeled randomly methylated-Beta-Cyclodextrin (FITC-RAMEB) on Caco-2 cell layer and examine the cellular entry of cyclodextrins on intestinal cells. The permeability of FITC-RAMEB through Caco-2 monolayers was very limited. Using this compound in 0.05 mM concentration the permeability coefficient was 3.3561.2961028 cm/s and its permeability did not change in the presence of 5 mM randomly methylated-Beta-Cyclodextrin. Despite of the low permeability, cellular accumulation of FITC-RAMEB in cytoplasmic vesicles was significant and showed strong time and concentration dependence, similar to the characteristics of the macropinocytosis marker Lucifer Yellow. The internalization process was fully inhibited at 0uC and it was drastically reduced at 37uC applying rottlerin, an inhibitor of macropinocytosis. Notably, FITC-RAMEB colocalized with the early endosome organizer Rab5a. These results have revealed that FITC-RAMEB is able to enter intestinal epithelial cells by fluid-phase endocytosis from the apical side. This mechanism can be an additional process which helps to overcome the intestinal barrier and contributes t

  • investigation of transport of genistein daidzein and their inclusion complexes prepared with different cyclodextrins on caco 2 cell line
    Journal of Pharmaceutical and Biomedical Analysis, 2013
    Co-Authors: Agnes Emma Daruhazi, Lajos Szente, Timea Kiss, Miklos Vecsernyes, Eva Szőke, E Lemberkovics
    Abstract:

    Abstract Isoflavonoids are widespread constituents in medical plants especially in legumes ( Fabaceae ), but occur in other different plant families as well ( Rosaceae , Iridaceae , Amaranthaceae ). Their antioxidant, estrogen-like, anti-inflammatory and analgesic effects make them promising compounds in therapy of important disorders especially in estrogen related diseases. Poor solubility in aqueous system of genistein and daidzein needs a solubility enhancement for pharmaceutical use. These compounds are suitable guest molecules for inclusion complex formation with cyclodextrins (CDs) considering matching their size and polarity. The molecular encapsulation with Beta-Cyclodextrin (β-CD), gamma-cyclodextrin (γ-CD), hydroxypropyl-Beta-Cyclodextrin (HP-β-CD) and random methyl-beta cyclodextrin (RAMEB-CD) results in a solid, molecularly dispersed form and in a significantly improved aqueous solubility of genistein and daidzein. Determining enhancement in solubility and bioavailability we investigated the transport of these inclusion complexes across Caco-2 cell line comparing that of the pure compounds and found significant improving effect of the different CD derivatives on membrane permeation of the two isoflavone aglycons.

Thierry Rabilloud - One of the best experts on this subject based on the ideXlab platform.

  • A single step protein assay that is both detergent and reducer compatible: The cydex blue assay.
    Electrophoresis, 2016
    Co-Authors: Thierry Rabilloud
    Abstract:

    Determination of protein concentration in often an absolute pre-requisite in preparing samples for biochemical and proteomic analyses. However, current protein assay methods are not compatible with both reducers and detergents, which are however present simultaneously in most denaturing extraction buffers used in proteomics and electrophoresis, and in particular in SDS electrophoresis. We found that inclusion of cyclodextrins in a Coomassie blue-based assay made it compatible with detergents, as cyclodextrins complex detergents in a 1:1 molecular ratio. As this type of assay is intrinsically resistant to reducers, we have thus developed a single step assay that is both detergent and reducer compatible. Depending on the type and concentration of detergents present in the sample buffer, either Beta-Cyclodextrin or alpha-cyclodextrin can be used, the former being able to complex a wider range of detergents and the latter being able to complex higher amounts of detergents due to its greater solubility in water. Cyclodextrins are used at final concentrations of 2-10 mg/mL in the assay mix. This typically allows to measure samples containing as little as 0.1 mg/mL protein, in the presence of up to 2% detergent and reducers such as 5 % mercaptoethanol or 50 mM DTT in a single step with a simple spectrophotometric assay. This article is protected by copyright. All rights reserved.

Palma Feher - One of the best experts on this subject based on the ideXlab platform.

  • fluorescently labeled methyl beta cyclodextrin enters intestinal epithelial caco 2 cells by fluid phase endocytosis
    PLOS ONE, 2014
    Co-Authors: Ferenc Fenyvesi, Lajos Szente, Katalin Retinagy, Zsolt Bacso, Zsuzsanna Gutaytoth, Milo Malanga, Eva Fenyvesi, Judit Varadi, Zoltan Ujhelyi, Palma Feher
    Abstract:

    Cyclodextrins are widely used excipients for increasing the bioavailability of poorly water-soluble drugs. Their effect on drug absorption in the gastrointestinal tract is explained by their solubility- and permeability-enhancement. The aims of this study were to investigate penetration properties of fluorescently labeled randomly methylated-Beta-Cyclodextrin (FITC-RAMEB) on Caco-2 cell layer and examine the cellular entry of cyclodextrins on intestinal cells. The permeability of FITC-RAMEB through Caco-2 monolayers was very limited. Using this compound in 0.05 mM concentration the permeability coefficient was 3.35±1.29×10−8 cm/s and its permeability did not change in the presence of 5 mM randomly methylated-Beta-Cyclodextrin. Despite of the low permeability, cellular accumulation of FITC-RAMEB in cytoplasmic vesicles was significant and showed strong time and concentration dependence, similar to the characteristics of the macropinocytosis marker Lucifer Yellow. The internalization process was fully inhibited at 0°C and it was drastically reduced at 37°C applying rottlerin, an inhibitor of macropinocytosis. Notably, FITC-RAMEB colocalized with the early endosome organizer Rab5a. These results have revealed that FITC-RAMEB is able to enter intestinal epithelial cells by fluid-phase endocytosis from the apical side. This mechanism can be an additional process which helps to overcome the intestinal barrier and contributes to the bioavailability enhancement of cyclodextrins.

Kaneto Uekama - One of the best experts on this subject based on the ideXlab platform.

  • crystallization and polymorphic transitions of chlorpropamide in aqueous 2 hydroxybutyl β cyclodextrin solution
    European Journal of Pharmaceutical Sciences, 2010
    Co-Authors: Fumitoshi Hirayama, Takako Ishiguro, Daisuke Iohara, Hidetoshi Arima, Kaneto Uekama
    Abstract:

    Effects of cyclodextrins on crystallization of chlorpropamide and the polymorphic transition mechanism of the drug in aqueous solution were investigated. In the presence of 2-hydroxybutyl-Beta-Cyclodextrin, chlorpropamide was exclusively crystallized to metastable Form II and III polymorphs, whereas it was crystallized to stable Form A in the absence of the Beta-Cyclodextrin at 4 degrees C. The crystallization to metastable Form II or III polymorph was dependent upon 2-hydroxybutyl-Beta-Cyclodextrin concentrations employed, i.e. crystallization to Form III at a lower concentration (0.5 mM), whereas to Form II in a higher concentration (5 mM). At an intermediate concentration (2 mM), the least stable Form II crystal was initially precipitated, but it was transformed to Form III crystal. At higher temperature, Form III crystal was converted to stable Form A crystal. In aqueous solution, chlorpropamide crystallized to stable Form A crystal consecutively through metastable Forms II and III, according to "Ostwald's Rule of Stages". 2-Hydroxybutyl-Beta-Cyclodextrin inhibits the transition of Form II to Form III at higher concentrations and that of Form III to Form A at lower concentrations. The results suggest that 2-hydroxybutyl-Beta-Cyclodextrin is useful for selective preparation of metastable chlorpropamide polymorphs occurring during crystallization according to the Ostwald's rule.

  • improvement of nasal bioavailability of luteinizing hormone releasing hormone agonist buserelin by cyclodextrin derivatives in rats
    Journal of Pharmaceutical Sciences, 1995
    Co-Authors: Kazutaka Matsubara, Tetsumi Irie, Kazuya Abe, Kaneto Uekama
    Abstract:

    The effects of chemically modified cyclodextrins on the nasal absorption of buserelin, an agonist of luteinizing hormone-releasing hormone, were investigated in anesthetized rats. Of the cyclodextrins tested, dimethyl-Beta-Cyclodextrin (DM-beta-CyD) was the most effective in improving the rate and extent of the nasal bioavailability of buserelin. Fluorescence spectroscopic studies indicated that the cyclodextrins formed inclusion complexes with buserelin, which may reduce the diffusibility of buserelin across the nasal epithelium and may participate in the protection of the peptide against enzymatic degradation in the nasal mucosa. Additionally, the cyclodextrins increased the permeability of the nasal mucosa, which was the primary determinant based on the multiple regression analysis of the nasal absorption enhancement of buserelin. Scanning electron microscopic observations revealed that DM-beta-CyD induced no remarkable changes in the surface morphology of the nasal mucosa at a minimal concentration necessary to achieve substantial absorption enhancement. The present results suggest that DM-beta-CyD could improve the nasal bioavailability of buserelin and is well-tolerated by the nasal mucosa of the rat.

  • peracylated β cyclodextrins as novel sustained release carriers for a water soluble drug molsidomine
    Journal of Pharmacy and Pharmacology, 1994
    Co-Authors: Kaneto Uekama, Takashi Horikawa, Masayuki Yamanaka, Fumitoshi Hirayama
    Abstract:

    Peracylated Beta-Cyclodextrins with different alkyl chains (acetyl-octanoyl) were prepared by acylating all hydroxyl groups of Beta-Cyclodextrin (beta-CyD), and their physical properties were evaluated. These hydrophobic beta-CyDs decreased the release rate of molsidomine, a peripheral vasodilator, in proportion to the lengthening of alkyl chain and suppressed a peak plasma level of molsidomine following oral administration of peracylated beta-CyD complexes to dogs. Among the peracylated beta-CyDs tested, perbutanoyl-beta-CyD maintained sufficient plasma drug levels for a long period of time, while other peracylated beta-CyDs having shorter or longer chains were inappropriate to control the in-vivo release behaviour of molsidomine. The prominent retarding effect of perbutanoyl-beta-CyD was ascribable to the appropriate mucoadhesive property and hydrophobicity, compared with other peracylated beta-CyDs. The present results suggest that perbutanoyl-beta-CyD is particularly useful in modifying the release rate of water-soluble drugs as a novel slow-release carrier.