AIMTo study the expressions of scavenger receptor class B type I(SR-BI) and peroxisome proliferator-activated receptor gamma (PPARgamma) in atherosclerotic mini swine and provide a new mechanism for investigating the pathogenesis of atherosclerosis.

METHODSChinese mini swine were fed by a normal control diet or a high fat/high cholesterol diet for 12 months after common carotid artery injury induced by balloon denudation. Plasma total cholesterol(TC), high-density lipoprotein cholesterol (HDL-C) and triglycerides (TG) were determined by commercially enzymatic methods every two months. The sections, which were taken from liver and abdominal aorta, were stained with hematoxylin eosin. The expressions of SR-BI and PPARgamma mRNA and protein in liver and aorta tissue were detected by reverse transcriptase-polymerase chain reaction (RT-PCR), Western blot and immunohistochemistry respectively.

RESULTSAt the end of 12 months, plasma TC, HDL-C and TG in HFHC mini swine were increased. There were fatty liver and atherosclerotic plaque in mini swine live and aorta respectively. The expression of SR-BI was upregulated in HFHC mini swine liver and aorta tissue.

CONCLUSIONHFHC may induce atherosclerosis and the expression of SR-BI and PPARgamma. Upregulating SR-BI expression may inhibit atherosclerosis. Increasing SR-BI expression in liver and aorta may accelerate SR-BI-mediated reverse cholesterol transport and develop a new anti-atherogenic strategy.

Animals ; Arteriosclerosis ; pathology ; Atherosclerosis ; metabolism ; PPAR gamma ; metabolism ; Receptors, Scavenger ; metabolism ; Swine

OBJECTIVETo investigate the effect of FXR on scavenger receptor class B type I (SR-BI) expression.

METHODSHuman vascular endothelium Eahy926 cells were treated with FXR agonist androsterone, and the specific target gene of FXR SHP mRNA was detected by RT-PCR. SR-BI mRNA and protein were determined using RT-PCR, real-time PCR and Western blotting.

RESULTSThe level of SHP mRNA in Eahy926 cells increased after androsterone treatment at different concentrations for 24 h, demonstrating FXR activation in the cells. RT-PCR, real-time PCR and Western blotting detected increased SR-BI expression at both mRNA and protein levels after FXR activation.

CONCLUSIONFXR increases the expression of SR-BI in human vascular endothelium cells.

Androsterone ; pharmacology ; Cell Line ; Endothelial Cells ; drug effects ; metabolism ; Humans ; Receptors, Cytoplasmic and Nuclear ; agonists ; metabolism ; Scavenger Receptors, Class B ; metabolism

Animals ; Enterovirus A, Human ; metabolism ; Humans ; Lysosome-Associated Membrane Glycoproteins ; metabolism ; Receptors, Scavenger ; metabolism ; Virion ; metabolism ; Virus Attachment

AIMTo develop a fluorescence polarization-based high throughput screening and identify ligands for human Lectin-like oxidized low-density lipoprotein receptor-1 (hLOX-1).

METHODSSequential ultracentrifugation at 4 degrees C from normolipidemic fasting volunteers to obtain low density lipoprotein (LDL), which was modified by CuSO4 (5 micromol x L(-1)) at 37 degrees C for 24 h. The assay was based on the interaction between receptor and ligand, and hLOX-1 was labeled by FITC and bound to its specific ligand, oxLDL. Different reaction time and DMSO concentration were optimized to determine the stability and tolerance of fluorescence polarization (FP) assay. 3 200 compounds were screened in black 384-well microplate by FP-based competitive displacement assay, at excitation filter of 485 nm and emission filter of 530 nm. Z' was used to assess the assay quality.

RESULTSThe FP-based HTS was formatted in a 384-well microplate with a Z' factor of 0. 75, and three active compounds for hLOX-1 were identified with IC50 below 40 micromol x L(-1) from total 3 200 compounds.

CONCLUSIONThe results indicated that the fluorescence polarization assay is stable, sensitive, reproducible and well suited for high throughput screening efforts.

Binding, Competitive ; Drug Evaluation, Preclinical ; methods ; Fluorescence Polarization ; methods ; Humans ; Ligands ; Lipoproteins, LDL ; metabolism ; Scavenger Receptors, Class E ; metabolism

Acute-Phase Proteins ; physiology ; Carrier Proteins ; physiology ; Humans ; Membrane Glycoproteins ; physiology ; Receptors, Cell Surface ; physiology ; Receptors, Immunologic ; physiology ; Receptors, Scavenger ; Sepsis ; metabolism ; physiopathology ; Signal Transduction ; Toll-Like Receptors

OBJECTIVETo explore the regulational effect of oxidized low-density lipoprotein (Ox-LDL) on expression of type A scavenger receptor (SR-A) in human mesangial cells (HMC).

METHODSHMC line (HMCL) with high expression of SR-A (HMCL-SRA) was established after stable transfection of expressive vector with cDNA encoding SR-A. Uptake of Ox-LDL by HMCL was evaluated using Oil Red "O" staining. SR-A mRNA expression was examined using reverse transcription-polymerase chain reaction (RT-PCR).

RESULTSMore uptake of Ox-LDL was observed in the HMCL-SRA than that in the untransfected HMCL. Ox-LDL could induce SR-A mRNA expression in HMC in a dose-dependent manner, and reached a peak level after 24 h of stimulation. After 24 h of stimulation with Ox-LDL at the dose of 10, 50 and 100 micrograms/ml, SR-A mRNA expression was up-regulated to 1.35, 1.83 and 2.30-fold of controls, respectively. However, LDL had no effect on the expression of SR-A.

CONCLUSIONSIt suggests that SR-A be a major binding receptor to uptake Ox-LDL in HMC. Ox-LDL may promote the progression of chronic renal diseases through up-regulation of SR-A.

Cells, Cultured ; DNA, Complementary ; Glomerular Mesangium ; cytology ; metabolism ; Humans ; Lipoproteins, LDL ; pharmacology ; RNA, Messenger ; biosynthesis ; genetics ; Receptors, Immunologic ; biosynthesis ; genetics ; Receptors, Scavenger ; Reverse Transcriptase Polymerase Chain Reaction ; Scavenger Receptors, Class A ; Transfection ; Up-Regulation

To explore the anti-atherosclerotic mechanism of estrogen and especially observe the effect of estradiol on the content of cholesterol in J774a.1 mouse mononuclear/macrophage-derived foam cells which were incubated with oxidized low-density lipoproteins (ox-LDL). J774a.1 mouse mononuclear/macrophages were incubated with ox-LDL or with both ox-LDL and estradiol (1, 0.1 or 0.01 micromol x L(-1)). Oil red O staining was used to observe the formation of foam cells, and cholesterol oxidase fluorometric was used to determine the content of cellular cholesterol content. Western blotting and RTFQ-PCR were used to observe the expressions of scavenger receptor class B type I (SR-B I ) in J774a.1 foam cells. Compared with the control cells, J774a.1 mouse mononuclear/macrophage-derived foam cells showed significantly increased contents of total cholesterol and cholesterol ester (P < 0.001) and decreased SR-B I mRNA expression (P < 0.01). Estradiol treatment significantly lowered the contents of total cholesterol and cholesterol ester (P < 0.05), and increased SR-B I protein and mRNA expression (P < 0.01) in the foam cells in a dose-dependent manner. Estradiol can inhibit the formation of mononuclear/macrophage-derived foam cells by decreasing the contents of total cholesterol and cholesterol ester and up-regulating the expression of SR-B I in the foam cells.
Animals ; Cell Line ; Cholesterol ; metabolism ; Cholesterol Esters ; metabolism ; Estradiol ; pharmacology ; Foam Cells ; cytology ; metabolism ; Lipoproteins, LDL ; metabolism ; Macrophages ; drug effects ; metabolism ; Mice ; Scavenger Receptors, Class B ; metabolism

Bile is concentrated in the gallbladder, and is often supersaturated in terms of cholesterol concentration. Such high levels of cholesterol in gallbladder bile has clinical implications with respect to cholesterol gallstone formation. Gallbladder epithelial cells (GBEC) are exposed to high cholesterol concentrations on their apical surface. Therefore, GBEC are uniquely positioned to play an important role in modulating biliary cholesterol concentration. Recently, it has been documented that the key-transporter for polarized cholesterol and phospholipid efflux in GBEC is ATP-binding cassette transporter A1 (ABCA1) and liver X receptor alpha(LXR alpha)/retinoid X receptor (RXR) in the nucleus of GBEC which regulates ABCA1 expression. In addition, it also has been demonstrated that ligands of peroxisome proliferator-activated receptor alpha(PPARalpha) and PPARgamma modulate inflammation and affect ABCA1 expression in GBEC. This evidence proves that GBEC has a perfect system for cholesterol transport. We herein introduce the roles and mechanisms of ABCA1, ABCG5/ABCG8, scavenger receptor class B-I (SR-BI), LXRalpha/RXR, farnesoid X receptor (FXR), and PPARs related to cholesterol transport in GBEC with a review of our study experience and related literature.
Bile ; Cholesterol* ; Epithelial Cells* ; Gallbladder* ; Gallstones ; Inflammation ; Ligands ; Liver ; Metabolism* ; Peroxisome Proliferator-Activated Receptors ; Peroxisomes ; PPAR gamma ; Receptors, Cytoplasmic and Nuclear* ; Receptors, Scavenger

Fucoidan is a natural polysaccharide extracted from brown seaweeds, with a wide variety of biological features, especially the anti-inflammatory and anti-oxidative effects. Studies indicated that the anti-inflammatory effect of fucoidan related to its capacity to interact with the selectin or scavenger receptor on the cell membrane. Fucoidan can also inhibit the synthesis and release of reactive oxygen species (ROS), as well as promote its clearance, showing the anti-oxidative activity.
Animals ; Anti-Inflammatory Agents ; isolation & purification ; metabolism ; pharmacology ; Antioxidants ; isolation & purification ; pharmacology ; Polysaccharides ; isolation & purification ; metabolism ; pharmacology ; Reactive Oxygen Species ; metabolism ; Receptors, Scavenger ; metabolism ; Seaweed ; chemistry ; Selectins ; metabolism

Both hydropathy plot and in vitro translation results predict the topology of SR-BI; the receptor is an integral membrane protein of 509 amino acids, consisting of a short cytoplasmic N-terminus of 9 amino acids followed by a first transmembrane domain of 22 amino acids, the extracellular domain of 408 amino acids, the second transmembrane domain of 22 amino acids, and the cytoplasmic C-terminus of 47 amino acids. The immunoblot of rBBMV in the presence or absence of pAb589 peptide antigen (the C-terminal 22 amino acid residues of SR-BI) confirmed that the bands at apparent molecular weight of 140 and 210 kDa are SR-BI related protein which might be multimeric forms of SR-BI. 125I apo A-I overlay analysis showed that SR-BI can bind to its ligand, apo A-I, only when it is thoroughly matured - glycosylated and dimerized. The antibody which was generated against extracellular domain of SR-BI (pAb230) not only prevented 125I-labeled apo A-I from binding to 140 kDa band but also inhibited the esterified cholesterol uptake of rabbit BBMV with its IC50 value of 40 microgram/ml of IgG. In contrast, the antibody generated against the C-terminal domain of SR-BI (pAb589) did not show any effect either on cholesterol uptake of rabbit BBMV or 125I-labeled apo A-I binding to 140 kDa band. Overall results show that the ligand binding site of SR-BI in rabbit BBMV is located in extracellular domain, and SR-BI is only functional when it is part of dimeric forms which rationalize the previously found cooperative nature of the binding interaction and maybe a fundamental finding towards the so far poorly understood mechanism of SR-BI function.
Amino Acid Sequence ; Animals ; Antigens, CD36/*metabolism ; Apolipoprotein A-I/metabolism ; Binding Sites/physiology ; Blotting, Western ; Caco-2 Cells ; Cholesterol Esters/metabolism ; Humans ; Intestinal Mucosa/metabolism ; Intestine, Small/*metabolism/ultrastructure ; Iodine Radioisotopes ; Membrane Proteins/*metabolism ; Microvilli/metabolism ; Molecular Sequence Data ; Rabbits ; *Receptors, Immunologic ; Receptors, Lipoprotein/*metabolism ; Receptors, Scavenger ; Scavenger Receptors, Class B ; Surface Properties