Expression of liver plasma membrane transporters in gallstone-susceptible and gallstone-resistant mice

2002 ◽  
Vol 361 (3) ◽  
pp. 673-679 ◽  
Author(s):  
Oliver MÜLLER ◽  
Carmen SCHALLA ◽  
Jürgen SCHEIBNER ◽  
Eduard F. STANGE ◽  
Michael FUCHS
Keyword(s):  
Gene Expression ◽  
Plasma Membrane ◽  
Protein Expression ◽  
Bile Salt ◽  
Membrane Transporters ◽  
Biliary Lipid ◽  
Liver Plasma Membrane ◽  
Lithogenic Diet ◽  
Secretion Rates ◽  
Salt Secretion

We tested the hypothesis that differential expression of liver plasma membrane transporters might account for variations in biliary lipid secretion rates between gallstone-susceptible C57L/J and gallstone-resistant AKR/J mice. Plasma membrane fractions and total RNA isolated from livers of mice fed with a control or lithogenic (15% fat/1.25% cholesterol/0.5% cholic acid) diet were used for measurements of steady-state gene expression of hepatobiliary transport systems for bile salts (Ntcp1/Slc10a1, Oatp1/Slc21a1 and Bsep/Abcb11), phospholipids (Mdr2/Abcb4), organic anions (Mrp2/Abcc2) and organic cations (Oct1/Slc22a1). Irrespective of the diet, the steady-state gene expression of hepatobiliary transporters did not differ significantly between the two strains. Despite a higher basal bile flow and bile-salt secretion in C57L mice, Mrp2 (Abcc2) and Bsep (Abcb11) expression did not differ between the two strains. Elevated biliary phospholipid secretion in response to the lithogenic diet was linked to increased Mdr2 (Abcb4) protein expression, whereas the induction of Oct1 (Slc22a1) might reflect an enhanced uptake of choline for augmented phospholipid synthesis. In response to the lithogenic diet, Bsep (Abcb11) protein expression was up-regulated only marginally and bile salt secretion did not increase. The down-regulation of Ntcp1 (Slc10a1) protein expression might protect hepatocytes from high intracellular bile-salt loads. We conclude that variations in protein function rather than in the gene expression of liver plasma membrane transporters might account for variations in biliary lipid secretion rates. Our findings support the concept that the formation of lithogenic bile is caused by the hypersecretion of bile salts as a result of augmented availability of canalicular membrane cholesterol, possibly amplified by bile-salt—phospholipid uncoupling due to the increased bile flow.

scholarly journals Selective biliary lipid secretion at low bile-salt-output rates in the isolated perfused rat liver. Effects of phalloidin

1986 ◽  
Vol 237 (1) ◽  
pp. 301-304 ◽  
Author(s):  
K Rahman ◽  
R Coleman
Keyword(s):  
Bile Salt ◽  
Bile Salts ◽  
Isolated Perfused Rat Liver ◽  
Perfused Rat Liver ◽  
Biliary Lipid ◽  
Perfusion Fluid ◽  
Lipid Secretion ◽  
Rat Livers ◽  
Secretion Rates ◽  
Salt Secretion

At high bile-salt-secretion rates the biliary secretion of phospholipids and cholesterol is dependent on that of the bile salts. However, at low bile-salt outputs some secretion remains. Isolated perfused rat livers were used in these experiments in order to study the bile-salt-independent secretion of biliary lipids. The livers were isolated and saline (0.9% NaCl), or phalloidin dissolved in saline, was added to the perfusion fluid after 1 h of liver isolation. The concentration and output of cholesterol was significantly decreased in phalloidin-treated livers compared with the controls, whereas there was no significant decrease in phospholipids; the secretion of cholesterol and phospholipids can thus be uncoupled from each other by the action of phalloidin. These experiments suggest that a proportion of cholesterol gets into bile independently of bile salts and phospholipids. These findings are discussed in relation to the supersaturation of some biles with cholesterol and its relationship to the bile-salt-independent fraction of cholesterol.

Identification of ferrichrome- and ferrioxamine B-mediated iron uptake by Aspergillus fumigatus

2016 ◽  
Vol 473 (9) ◽  
pp. 1203-1213 ◽  
Author(s):  
Yong-Sung Park ◽  
Ju-Yeon Kim ◽  
Cheol-Won Yun
Keyword(s):  
Gene Expression ◽  
Plasma Membrane ◽  
Aspergillus Fumigatus ◽  
Virulence Factors ◽  
Membrane Transporters ◽  
Yeast Cells ◽  
Double Deletion ◽  
Ferrioxamine B ◽  
Uptake Activity ◽  
Opportunistic Fungal Pathogen

Aspergillus fumigatus is an opportunistic fungal pathogen for immunocompromised patients, and genes involved in siderophore metabolism have been identified as virulence factors. Recently, we identified the membrane transporters sit1 and sit2, which are putative virulence factors of A. fumigatus; sit1 and sit2 are homologous to yeast Sit1, and sit1 and sit2 gene expression was up-regulated after iron depletion. When expressed heterologously in Saccharomyces cerevisiae, sit1 and sit2 were localized to the plasma membrane; sit1 efficiently complemented ferrichrome (FC) and ferrioxamine B (FOB) uptake in yeast cells, whereas sit2 complemented only FC uptake. Deletion of sit1 resulted in a decrease in FOB and FC uptake, and deletion of sit2 resulted in a decrease in FC uptake in A. fumigatus. It is of interest that a sit1 and sit2 double-deletion mutant resulted in a synergistic decrease in FC uptake activity. Both sit1 and sit2 were localized to the plasma membrane in A. fumigatus. The expression levels of the sit1 and sit2 genes were dependent on hapX under low-but not high-iron conditions. Furthermore, mirB, and sidA gene expression was up-regulated and sreA expression down-regulated when sit1 and sit2 were deleted. Although sit1 and sit2 failed to affect mouse survival rate, these genes affected conidial killing activity. Taken together, our results suggest that sit1 and sit2 are siderophore transporters and putative virulence factors localized to the plasma membrane.

scholarly journals Expression of liver plasma membrane transporters in gallstone-susceptible and gallstone-resistant mice

2002 ◽  
Vol 361 (3) ◽  
pp. 673 ◽  
Author(s):  
Oliver MÜLLER ◽  
Carmen SCHALLA ◽  
Jürgen SCHEIBNER ◽  
Eduard F. STANGE ◽  
Michael FUCHS
Keyword(s):  
Plasma Membrane ◽  
Membrane Transporters ◽  
Liver Plasma Membrane

scholarly journals Control of biliary phospholipid secretion. Effect of continuous and discontinuous infusion of taurocholate on biliary phospholipid secretion

1986 ◽  
Vol 234 (2) ◽  
pp. 421-427 ◽  
Author(s):  
K Rahman ◽  
T G Hammond ◽  
P J Lowe ◽  
S G Barnwell ◽  
B Clark ◽  
...  
Keyword(s):  
Animal Model ◽  
Bile Salt ◽  
Biliary Fistula ◽  
Perfused Liver ◽  
Biliary Lipid ◽  
Short Pulses ◽  
Lipid Secretion ◽  
Canalicular Membrane ◽  
Infusion Period ◽  
Salt Secretion

A major determinant of biliary lipid secretion is bile-salt secretion. Taurocholate (TC), a micelle-forming bile salt, was infused continuously at different rates in both isolated perfused livers and biliary-fistula rats. In both of these systems, infusion of TC brought about an elevated secretion of phosphatidylcholine for the duration of the TC infusion period. Initial phospholipid/bile-salt ratios in the bile were higher in the whole-animal model than in isolated livers, but at the higher infusion rates both secreted approx. 6 mol of phospholipid for every 100 mol of bile salt. The secretion of phospholipid, which was maintained even at high rates of bile-salt infusion, suggest a continuous and regulated phospholipid supply and secretion mechanism. In contrast, however, multiple short pulses of TC to the perfused liver, which brought about relatively equal biliary bile-salt output pulses, did not bring about equal phospholipid outputs, since the phospholipid peak size declined with each bile-salt pulse. These experiments taken together suggest either that a threshold (intracellular) bile-salt concentration may be required to ‘switch-on’ the phospholipid supply and that it may need to be maintained for continuous biliary phospholipid supply to the canalicular membrane.

Effects of sulfobromophthalein excretion on biliary lipid secretion in humans and dogs

1981 ◽  
Vol 240 (1) ◽  
pp. G85-G89
Author(s):  
E. A. Shafter ◽  
R. M. Preshaw
Keyword(s):  
Bile Salt ◽  
Lipid Composition ◽  
Control Period ◽  
Physical Interaction ◽  
Biliary Lipid ◽  
Lipid Secretion ◽  
Maximal Output ◽  
Biliary Lipid Composition ◽  
Molar Percent ◽  
Salt Secretion

The effect of sulfobromophthalein (BSP) on biliary lipid secretion was investigated in 12 cholecystectomized subjects, using a duodenal marker-perfusion technique. A 1-h basal period was followed by intravenous BSP infusion over 3 h, achieving the maximal excretory rate (Tm). The calculated Tm was not different from the measured maximal output. At BSP Tm, bile salt secretion was unchanged, but phospholipid, cholesterol, and bilirubin secretion were markedly reduced. Biliary lipid composition changed accordingly, higher molar percent bile salts but lower phospholipid and cholesterol. In six cholecystectomized dogs with chronic duodenal fistulas, bile was collected directly from the common duct while bile salt secretion was maintained by intravenous taurocholic acid infusion. After a 2-h control period, sufficient BSP was added to create either maximal (Tm) or submaximal conditions. BSP did not alter bile salt secretion but caused a dose-related decrease in phospholipid and cholesterol secretion. Bilirubin excretion was also reduced, whereas bile flow increased. Thus, BSP is hydrocholeretic but decreases phospholipid, cholesterol, and bilirubin secretion in both humans and dogs. The effect on biliary lipid composition is probably through a physical interaction with biliary micelles.

The mechanism of increased biliary lipid secretion in mice with genetic inactivation of bile salt export pump

2015 ◽  
Vol 308 (5) ◽  
pp. G450-G457 ◽  
Author(s):  
K. E. R. Gooijert ◽  
R. Havinga ◽  
H. Wolters ◽  
R. Wang ◽  
V. Ling ◽  
...  
Keyword(s):  
Protein Expression ◽  
Bile Salt ◽  
Biliary Secretion ◽  
Biliary Lipid ◽  
Bile Salt Export Pump ◽  
Human Bile ◽  
Lipid Secretion ◽  
Canalicular Transporters ◽  
Expression Potential ◽  
And Control

Human bile salt export pump ( BSEP) mutations underlie progressive familial intrahepatic cholestasis type 2 (PFIC2). In the PFIC2 animal model, Bsep−/−mice, biliary secretion of bile salts (BS) is decreased, but that of phospholipids (PL) and cholesterol (CH) is increased. Under physiological conditions, the biliary secretion of PL and CH is positively related (“coupled”) to that of BS. We aimed to elucidate the mechanism of increased biliary lipid secretion in Bsep−/−mice. The secretion of the BS tauro-β-muricholic acid (TβMCA) is relatively preserved in Bsep−/−mice. We infused Bsep−/−and Bsep+/+(control) mice with TβMCA in stepwise increasing dosages (150–600 nmol/min) and determined biliary bile flow, BS, PL, and CH secretion. mRNA and protein expression of relevant canalicular transporters was analyzed in livers from noninfused Bsep−/−and control mice. TβMCA infusion increased BS secretion in both Bsep−/−and control mice. The secreted PL or CH amount per BS, i.e., the “coupling,” was continuously two- to threefold higher in Bsep−/−mice ( P < 0.05). Hepatic mRNA expression of canalicular lipid transporters Mdr2, Abcg5, and Abcg8 was 45–55% higher in Bsep−/−mice (Abcg5; P < 0.05), as was canalicular Mdr2 and Abcg5 protein expression. Potential other explanations for the increased coupling of the biliary secretion of PL and CH to that of BS in Bsep−/−mice could be excluded. We conclude that the mechanism of increased biliary lipid secretion in Bsep−/−mice is based on increased expression of the responsible canalicular transporter proteins.

Cholic acid aids absorption, biliary secretion, and phase transitions of cholesterol in murine cholelithogenesis

1999 ◽  
Vol 276 (3) ◽  
pp. G751-G760 ◽  
Author(s):  
David Q.-H. Wang ◽  
Frank Lammert ◽  
David E. Cohen ◽  
Beverly Paigen ◽  
Martin C. Carey
Keyword(s):  
Cholic Acid ◽  
Bile Flow ◽  
Cholesterol Biosynthesis ◽  
Cholesterol Absorption ◽  
Biliary Lipid ◽  
Biliary Cholesterol ◽  
Intestinal Cholesterol Absorption ◽  
Lipid Secretion ◽  
Lithogenic Diet ◽  
Secretion Rates

Cholic acid is a critical component of the lithogenic diet in mice. To determine its pathogenetic roles, we fed chow or 1% cholesterol with or without 0.5% cholic acid to C57L/J male mice, which because of lith genes have 100% gallstone prevalence rates. After 1 yr on the diets, we measured bile flow, biliary lipid secretion rates, hepatic cholesterol and bile salt synthesis, and intestinal cholesterol absorption. After hepatic conjugation with taurine, cholate replaced most tauro-β-muricholate in bile. Dietary cholic acid plus cholesterol increased bile flow and biliary lipid secretion rates and reduced cholesterol 7α-hydroxylase activity significantly mostly via deoxycholic acid, cholate’s bacterial 7α-dehydroxylation product but did not downregulate cholesterol biosynthesis. Intestinal cholesterol absorption doubled, and biliary cholesterol crystallized as phase boundaries shifted. Feeding mice 1% cholesterol alone produced no lithogenic or homeostatic effects. We conclude that in mice cholic acid promotes biliary cholesterol hypersecretion and cholelithogenesis by enhancing intestinal absorption, hepatic bioavailability, and phase separation of cholesterol in bile.

scholarly journals SUN-267 Dissecting Type 2 CRH Receptor Signaling Characteristics in the Hypothalamic Cell Line MHYPOA-2/30

2020 ◽  
Vol 4 (Supplement_1) ◽  
Author(s):  
Viridiana Alcantara-Alonso ◽  
Patricia de Gortari ◽  
Robert Dallmann ◽  
Dimitris Grammatopoulos
Keyword(s):  
Gene Expression ◽  
Plasma Membrane ◽  
Protein Expression ◽  
Cell Line ◽  
Cellular Localization ◽  
Luciferase Reporter ◽  
Mrna Levels ◽  
Luciferase Reporter Gene

Abstract The stress peptides coticotropin-releasing hormone (CRH) and urocortins (Ucns) exert anorectic effects acting mainly through the type 2 CRH receptor (CRH-R2) in the hypothalamus. Impairment of CRH-R2 signaling in chronically stressed rats has been linked with the development of hyperphagia (Alcantara-Alonso et al. Neuropeptides, 2017) however the exact mechanisms and molecular defects are unknown. In the present study we used the mHypoA-2/30, a hypothalamic immortalized cell line derived from adult mice (Belsham et al. FASEB J, 2009) to further explore the signaling molecules mediating the anorexigenic effect of the CRH-R2 cognate agonist urocortin 2 (Ucn2). Specifically, we investigated mRNA, protein expression and cellular localization of CRH-R2 in the mHypoA-2/30 neurons. Additionally, we examined the effects of Ucn2 on the phosphorylation of CREB and AMPK, as well as its transcriptional effects on genes of feeding-related peptides and molecules involved in modulation of circadian rhythms. Both CRH-R2 mRNA and protein expression were detected in mHypoA-2/30; indirect immunoflourescence experiments using a specific CRH-R2 antibody demonstrated widespread localization in the plasma membrane and cytoplasm. Moreover, the receptor sub-cellular localization was redistributed in response to activation by Ucn2 (100 nM), as the plasma membrane immunofluorescent signal was decreased after 4h of agonist treatment, suggesting CRH-R2 homologous internalization. We also observed a 50% increase in the phosphorylation of CREB associated with a concomitant decrease in AMPK phosphorylation after 30 min of Ucn2 treatment. Among the panel of hypothalamic genes analyzed, we identified after 24h of Ucn2 treatment increases in the gene expression of the anorexigenic peptides neurotensin and proopiomelanocortin. Interestingly, sustained CRH-R2 activation also led to an increase in the mRNA levels of Aryl Hydrocarbon Receptor Nuclear Translocator Like (ARNTL), a protein involved in the control of circadian rhythm. A luciferase reporter gene analysis of ARNTL showed that the mHypoA-2/30 cells also exhibit circadian patterns of expression and that the treatment with Ucn2 enhanced circadian amplitude of ARNTL reporter on these cells, which in turn may be involved in glucocorticoid release in circadian cycles and stimulating appetite during the activity phase of the animals. In conclusion, we found that the mHypoA-2/30 cell line expresses endogenous functional CRH-R2 that is linked to downstream regulation of anorexigenic gene expression. This cell line appears to be a useful in vitro tool to study hypothalamic CRH-R2 signaling machinery involved in central control of food intake and circadian cycles.

Molecular mechanisms of hepatic bile salt transport from sinusoidal blood into bile

1995 ◽  
Vol 269 (6) ◽  
pp. G801-G812 ◽  
Author(s):  
P. J. Meier
Keyword(s):  
Plasma Membrane ◽  
Bile Salt ◽  
Bile Salts ◽  
Organic Anion ◽  
Salt Transport ◽  
Salt Flux ◽  
Salt Uptake ◽  
Canalicular Bile ◽  
Salt Secretion ◽  
Bile Salt Transport

An increasingly complex picture has emerged in recent years regarding the bile salt transport polarity of hepatocytes. At the sinusoidal (or basolateral) plasma membrane two bile salt-transporting polypeptides have been cloned. The Na(+)-taurocholate-cotransporting polypeptide (Ntcp) can account for most, if not all, physiological properties of the Na(+)-dependent bile salt uptake function in mammalian hepatocytes. The cloned organic anion-transporting protein (Oatp1) can mediate Na(+)-independent transport of bile salts, sulfobromophthalein, estrogen conjugates, and a variety of other amphipathic cholephilic compounds. Hence, Oatp1 appears to correspond to the previously suggested basolateral multispecific bile sale transporter. Intracellular bile salt transport can be mediated by different pathways. Under basal bile salt flux conditions, conjugated trihydroxy bile salts bind to cytoplasmic binding proteins and reach the canalicular plasma membrane predominantly via cytoplasmic diffusion. More hydrophobic mono- and dihydroxy and high concentrations of trihydroxy bile salts associate with intracellular membrane-bound compartments, including transcytotic vesicles, endoplasmic reticulum (ER), and Golgi complex. A facilitated bile salt diffusion pathway has been demonstrated in the ER. The exact role of these and other (e.g., lysosomes, "tubulovesicular structures") organelles in overall vectorial transport of bile salts across hepatocytes is not yet known. Canalicular bile salt secretion is mediated by two ATP-dependent transport systems, one for monovalent bile salts and the second for divalent sulfated or glucuronidated bile salt conjugates. The latter is identical with the canalicular multispecific organic anion transporter, which also transports other divalent organic anions, such as glutathione S-conjugates. Potential dependent canalicular bile salt secretion has also been suggested to occur, but its exact mechanism and physiological significance remain unclear, since a potential driven bile salt uptake system has also been identified in the ER. Hypothetically, and similar to changes in cell volume, the intracellular potential could also play a role in the regulation of the number of bile salt carriers at the canalicular membrane and thereby indirectly influence the maximal canalicular bile salt transport capacity of hepatocytes.

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