Cholate resistance in Lactococcus lactis is mediated by an ATP-dependent multispecific organic anion transporter. (9/61)

The cholate-resistant Lactococcus lactis strain C41-2, derived from wild-type L. lactis MG1363 through selection for growth on cholate-containing medium, displayed a reduced accumulation of cholate due to an enhanced active efflux. However, L. lactis C41-2 was not cross resistant to deoxycholate or cationic drugs, such as ethidium and rhodamine 6G, which are typical substrates of the multidrug transporters LmrP and LmrA in L. lactis MG1363. The cholate efflux activity in L. lactis C41-2 was not affected by the presence of valinomycin plus nigericin, which dissipated the proton motive force. In contrast, cholate efflux in L. lactis C41-2 was inhibited by ortho-vanadate, an inhibitor of P-type ATPases and ATP-binding cassette transporters. Besides ATP-dependent drug extrusion by LmrA, two other ATP-dependent efflux activities have previously been detected in L. lactis, one for the artificial pH probe 2',7'-bis-(2-carboxyethyl)-5(and 6)-carboxyfluorescein (BCECF) and the other for the artificial pH probe N-(fluorescein thio-ureanyl)-glutamate (FTUG). Surprisingly, the efflux rate of BCECF, but not that of FTUG, was significantly enhanced in L. lactis C41-2. Further experiments with L. lactis C41-2 cells and inside out membrane vesicles revealed that cholate and BCECF inhibit the transport of each other. These data demonstrate the role of an ATP-dependent multispecific organic anion transporter in cholate resistance in L. lactis.  (+info)

Genetic alterations of IL-1 receptor antagonist in mice affect plasma cholesterol level and foam cell lesion size. (10/61)

Inflammatory cytokines have been linked to atherosclerosis by using cell culture models and acute inflammation in animals. The goal of this study was to examine lipoprotein levels and early atherosclerosis in chronic animal models of altered IL-1 physiology by using mice with deficient or excess IL-1 receptor antagonist (IL-1ra). IL-1ra knockout C57BL/6J mice fed a cholesterol/cholate diet for 3 mo had a 3-fold decrease in non-high-density lipoprotein cholesterol and a trend toward increased foam-cell lesion area compared to wild-type littermate controls. IL-1ra transgenic/low-density lipoprotein receptor (LDLR) knockout mice fed a cholesterol-saturated fat diet for 10 wk showed a 40% increase in non-high-density lipoprotein cholesterol, consistent with the IL-1ra knockout data, although there was no change in lesion size. When these IL1-ra overexpressing transgenic mice on the LDLR knockout background were fed a high-cholesterol/high-fat diet containing cholate, however, a statistically significant 40% decrease in lesion area was observed compared to LDLR knockout mice lacking the transgene. By immunohistochemistry, IL-1ra was present in C57BL/6J and LDLR knockout aortae, absent in IL-1ra knockout aortae, and present at high levels in LDLR knockout/IL-1ra transgene aortae. In summary, IL-1ra tended to increase plasma lipoprotein levels and, when fed a cholate-containing diet, decrease foam-cell lesion size. These data demonstrate that in selected models of murine atherosclerosis, chronic IL-1ra depletion or overexpression has potentially important effects on lipoprotein metabolism and foam-cell lesion development.  (+info)

Iron chelators as antimalarial agents: in vitro activity of dicatecholate against Plasmodium falciparum. (11/61)

The present study was undertaken to explore the antimalarial effect of a series of dicatecholate iron chelators. They may be made more or less lipophilic by increasing or reducing the length of the R substituent on the nitrogen. In vitro activity against the W2 and 3D7 clones of Plasmodium falciparum, toxicity on Vero cells and toxicity on uninfected erythrocytes by measure of the released haemoglobin were assessed for each compound. These findings were compared with the ability of iron(III), iron(II) and ferritin to reverse the inhibitory effect of catecholates. This study shows that increased lipid solubility of catecholate iron chelators does not lead to improved antimalarial activity. However, their activity is well correlated with their interaction with iron and with their toxicity against Vero cells. This study demonstrates a potent antimalarial effect of FR160 (R = C9H19) on five different strains of P. falciparum in vitro. FR160 inhibited parasite growth with an IC50 between 0.8 and 1.5 micro M. The effects of FR160 on mammalian cells were minimal compared with those obtained with malaria parasites. FR160 acted on parasites at considerably higher rates than desferrioxamine, and at all stages of parasite growth. The drug was more effective at the late trophozoite and young schizont stages, although FR160 affected rings and schizonts as well. Ascorbic acid, a free radical scavenger, reduced the activities of FR160 and artesunate. FR160 might induce formation of free radicals, which could explain why FR160 antagonized the effects of artesunate and dihydroartemisinin.  (+info)

Physical properties of phosphatidylcholine vesicles containing small amount of sodium cholate and consideration on the initial stage of vesicle solubilization. (12/61)

The effects of sub-solubilizing concentrations of sodium cholate (Na-chol) on several physicochemical properties of phosphatidylcholine (PC) small unilamellar vesicles (SUV) were considered in connection with the initial stage of membrane solubilization. ESR spectra of 12-doxylstearic acid (12-DS) in phosphatidylcholine from egg yolk (EPC) or dimyristoylphosphatidylcholine (DMPC) SUV at low concentrations (insufficient to destroy the vesicles) of Na-chol were composed of two (a strongly immobilized and an additional weakly immobilized) immiscible components. The origin of the additional bands was phase separation which occurred in the hydrophobic parts of PC SUV in the presence of Na-chol. Differential scanning calorimetry measurements demonstrated that the mixed DMPC/Na-chol SUV possessed two (a sharp low-temperature and a broad high-temperature) endothermic peaks, which is consistent with the coexistence of two immiscible phases in the vesicular membranes. zeta Potentials of the EPC/Na-chol SUV revealed that high anionic densities appeared on the surfaces of the SUV at a Na-chol concentration slightly below the upper boundary of the vesicle region. Thus, the initial stage of the solubilization of PC SUV by Na-chol was caused by the aggregation of hydrophobic parts of PC membranes, followed by the occurrence of high anionic densities on the surfaces of the vesicles. The fact that removal of Na-chol from PC/Na-chol mixed systems preferentially resulted in the formation of small vesicles might originate from these anionic charges.  (+info)

Antifolate resistance associated with loss of MRP1 expression and function in Chinese hamster ovary cells with markedly impaired export of folate and cholate. (13/61)

Export of folates from a Chinese hamster ovary PyrR100 cell line is markedly impaired, resulting in expansion of cellular folate pools and high-level antifolate resistance. We now report that MRP1 expression is absent in PyrR100 cells along with a marked decrease in MRP5 expression with 3-fold cross-resistance to thiopurines. PyrR100 and wild-type cells had comparable low levels of MRP2 expression; both lacked the breast cancer resistance protein. PyrR100 cells showed a 4-fold decrease in cholate (an MRP substrate) efflux with a 6-fold increase in cellular cholate accumulation compared with wild-type cells. Prostaglandin A1 increased cholate accumulation in wild-type cells to levels comparable with PyrR100 cells. Calcein (an MRP1 substrate) fluorescence increased 5-fold in PyrR100 cells; probenecid increased the intracellular calcein level in wild-type cells to that of PyrR100 cells. Consistent with the loss of MRP1 expression, PyrR100 cells showed modest collateral sensitivity to cholate, etoposide, doxorubicin, and vincristine. Transfection of MRP5 into PyrR100 cells did not alter sensitivity to pyrimethamine or MTX but restored sensitivity to mercaptopurines, indicating that decreased MRP5 expression did not play a role in antifolate resistance. Hence, although MRP-mediated anticancer drug resistance has been associated with gain of function (i.e., overexpression), this is the first report that loss of MRP1 efflux function can expand intracellular folate pools to result in acquired antifolate resistance. The data also suggest that MRP1, and possibly other MRPs that transport folates, can play a role in the maintenance of cellular folate homeostasis.  (+info)

Cholesterol and cholate components of an atherogenic diet induce distinct stages of hepatic inflammatory gene expression. (14/61)

Atherosclerosis in inbred mouse strains has been widely studied by using an atherogenic (Ath) diet containing cholesterol, cholic acid, and fat, but the effect of these components on gene expression has not been systematically examined. We employed DNA microarrays to interrogate gene expression levels in liver of C57BL/6J mice fed the following five diets: mouse chow, the Ath diet, or modified versions of the Ath diet in which either cholesterol, cholate, or fat were omitted. Dietary cholesterol and cholate produced discrete gene expression patterns. Cholesterol was required for induction of genes involved in acute inflammation, including three genes of the serum amyloid A family, three major histocompatibility class II antigen genes, and various cytokine-related genes. In contrast, cholate induced expression of genes involved in extracellular matrix deposition in hepatic fibrosis, including five collagen family members, collagen-interacting proteins, and connective tissue growth factor. The gene expression findings were confirmed by biochemical measurements showing that cholesterol was required for elevation of circulating serum amyloid A, and cholate was required for accumulation of collagen in the liver. The possibility that these gene expression changes are relevant to atherogenesis in C57BL/6J mice was supported by the observation that the closely related, yet atherosclerosis-resistant, C57BL/6ByJ strain was largely resistant to dietary induction of the inflammatory and fibrotic response genes. These results establish that cholesterol and cholate components of the Ath diet have distinct proatherogenic effects on gene expression and suggest a strategy to study the contribution of acute inflammatory response and fibrogenesis independently through dietary manipulation.  (+info)

Effect of mixed micelle formulations including terpenes on the transdermal delivery of diclofenac. (15/61)

The significant inhibitory action of diclofenac formulated in mixed micelles of lecithin with cholate or deoxycholate was observed on the rat hind paw edema induced by carrageenan. In the primary stage, mixed micelle formulation of deoxycholate was more effective compared with that of cholate. However, in the final term, the inhibitory action was similar in both formulations. In a previous study, the flux of diclofenac was greater in the mixed micelle formulation of deoxycholate compared with that of cholate. It was suggested that the permeation rate of diclofenac through skin was proportional to the pharmacological activity. The hind paw edema was quickly inhibited when cyclic monoterpene such as d-limonene or l-menthol was included in the formulations. All the micelle formulations significantly decreased the value of AUC estimated the hind paw thickness-time profile. This suggests that the micelle formulation of cholate in addition to deoxycholate showed significant anti-inflammatory activity to hind paw edema of rats. Incorporation of d-limonene or l-menthol was more effective on the decrease of AUC. A pharmacological study revealed that micelle formulations were able to reduce the skin irritation of chemicals.  (+info)

The farnesoid X receptor controls gene expression in a ligand- and promoter-selective fashion. (16/61)

Farnesoid X receptor (FXR) is a nuclear receptor for bile acids. Ligand activated-FXR regulates transcription of genes to allow feedback control of bile acid synthesis and secretion. There are five major bile acids in humans. We have previously demonstrated that lithocholate acts as an FXR antagonist, and here we show that the other four bile acids, chenodeoxycholate (CDCA), deoxycholate (DCA), cholate (CA), and ursodeoxycholate (UDCA), act as selective FXR agonists in a gene-specific fashion. In an in vitro coactivator association assay, CDCA fully activated FXR, whereas CA partially activated FXR and DCA and UDCA had negligible activities. Similar results were also obtained from a glutathione S-transferase pull-down assay in which only CDCA and the synthetic FXR agonist GW4064 significantly increased the interaction of SRC-1 with FXR. In FXR transactivation assays with a bile salt export pump (BSEP) promoter-driven luciferase construct, bile acids showed distinct abilities to activate the BSEP promoter: CDCA, DCA, CA, and UDCA increased luciferase activity by 25-, 20-, 18-, and 8-fold, respectively. Consistently, CDCA increased BSEP mRNA by 750-fold in HepG2 cells, whereas DCA, CA, and UDCA induced BSEP mRNA by 250-, 75-, and 15-fold, respectively. Despite the partial induction of BSEP mRNA, CA, DCA, and UDCA effectively repressed expression of cholesterol 7alpha-hydroxylase, another FXR target. We further showed that all four bile acids significantly increased FXR protein, suggesting the existence of an auto-regulatory loop in FXR signaling pathways. In conclusion, these results suggest that the binding of each bile acid results in a different FXR conformations, which in turn differentially regulates expression of individual FXR targets.  (+info)