By expression cloning using fluorescent-labeled high density lipoprotein (HDL), we isolated two clones that conferred the cell surface binding of HDL. Nucleotide sequence of the two clones revealed that one corresponds to scavenger receptor class B, type 1 (SRBI) and the other encoded a novel protein with 228 amino acids. The primary structure of the newly identified HDLbinding protein resembles GPI-anchored proteins consisting of an N-terminal signal sequence, an acidic region with a cluster of aspartate and glutamate residues, an Ly-6 motif highly conserved among the lymphocyte antigen family, and a C-terminal hydrophobic region. This newly identified HDL-binding protein designated GPI-anchored HDL-binding protein 1 (GPI-HBP1), was susceptible to phosphatidylinositol-specific phospholipase C treatment and binds HDL with high affinity (calculated K d ‫؍‬ 2-3 g/ml). Similar to SRBI, GPI-HBP1 mediates selective lipid uptake but not the protein component of HDL. Among various ligands for SRBI, HDL was most preferentially bound to GPI-HBP1. In contrast to SRBI, GPI-HBP1 lacked HDL-dependent cholesterol efflux. The GPI-HBP1 transcripts were detected with the highest levels in heart and, to a much lesser extent, in lung and liver. In situ hybridization revealed the accumulation of GPI-HBP1 transcripts in cardiac muscle cells, hepatic Kupffer cells and sinusoidal endothelium, and bronchial epithelium and alveolar macrophages in the lung.High density lipoprotein (HDL) 1 plays a key role in the transportation of cholesterol to extrahepatic tissues including steroidogenic tissues and in the reverse transportation of cholesterol from extrahepatic tissues to the liver (1). Unlike the low density lipoprotein (LDL) receptor pathway, the delivery of cholesterol from HDL to cells is mediated by selective lipid uptake from HDL particles and is independent of internalization of HDL. Reverse cholesterol transportation requires the extraction of cholesterol from extrahepatic cells by HDL and the subsequent delivery of cholesterol to hepatocytes.Several HDL-binding proteins have been identified including class B type I scavenger receptor (SRBI) (2, 3), two candidate hepatic HDL receptors designated HDL-binding proteins 1 and 2 (4, 5), 80-and 130-kDa GPI-anchored HDL-binding proteins expressed in human macrophages (6), 110-kDa GPI-anchored HDL-binding protein expressed in HepG2 cells (7), and recently characterized 95-kDa HDL-binding protein (8). To date, only SRBI appears to be a physiological HDL receptor based on the selective uptake of cholesterol esters into cells and the efflux of cholesterol from cells to HDL mediated by SRBI (1). Consistent with the postulated physiological role, SRBI is highly expressed in tissues that selectively take up cholesterol esters from HDL including liver, adrenal gland, testis, and ovary (3). Although hepatic overexpression of SRBI mediated by an adenovirus encoding SRBI resulted in a dramatic reduction of plasma cholesterol (9), the targeted disruption of the murine SRBI gene led to a modest inc...

LDL receptor-related protein 5 (LRP5) plays multiple roles, including embryonic development and bone accrual development. Recently, we demonstrated that LRP5 is also required for normal cholesterol metabolism and glucose-induced insulin secretion. To further define the role of LRP5 in the lipoprotein metabolism, we compared plasma lipoproteins in mice lacking LRP5, apolipoprotein E (apoE), or both (apoE;LRP5 double knockout). On a normal chow diet, the apoE;LRP5 double knockout mice (older than 4 months of age) had ϳ60% higher plasma cholesterol levels compared with the agematched apoE knockout mice. In contrast, LRP5 deficiency alone had no significant effects on the plasma cholesterol levels. High performance liquid chromatography analysis of plasma lipoproteins revealed that cholesterol levels in the very low density lipoprotein and low density lipoprotein fractions were markedly increased in the apoE;LRP5 double knockout mice. There were no apparent differences in the pattern of apoproteins between the apoE knockout mice and the apoE; LRP5 double knockout mice. The plasma clearance of intragastrically loaded triglyceride was markedly impaired by LRP5 deficiency. The atherosclerotic lesions of the apoE;LRP5 double knockout mice aged 6 months were ϳ3-fold greater than those in the age-matched apoE-knockout mice. Furthermore, histological examination revealed highly advanced arthrosclerosis, with remarkable accumulation of foam cells and destruction of the internal elastic lamina in the apoE;LRP5 double knockout mice. These data suggest that LRP5 mediates both apoE-dependent and apoE-independent catabolism of plasma lipoproteins.Genetic defects in the catabolism of plasma lipoproteins are important causes of hypercholesterolemia and atherosclerosis in humans. The prototypic diseases are familial hypercholesterolemia, caused by a defect in the LDL 1 receptor (LDLR) (1), and familial type III hyperlipoproteinemia, caused by a defect in one of the ligands for LDLR, apolipoprotein E (apoE) (2).ApoE is hypothesized to mediate lipoprotein clearance by binding two receptors: (i) LDLR and (ii) a hepatic chylomicron remnant receptor. ApoE-deficient mice (3-5) and LDLR-deficient mice (6) exhibit hypercholesterolemia, but the severity and manifestations differ markedly. On a normal laboratory chow diet, the apoE knockout mice have much more profound hypercholesterolemia and develop spontaneous atherosclerosis (4).LDL receptor-related protein 5 (LRP5) is a member of the LDL receptor family that are characterized by the presence of cysteine-rich complement type ligand binding domains. LRP5 binds apoE-containing lipoproteins in vitro and is widely expressed in many tissues including hepatocytes, adrenal gland, and pancreas (7).LRP5 and its homologue, LRP6, are postulated to play as co-receptors for Wnt receptors, Frizzled (8 -13). The Wnt signaling pathway plays an essential role in embryonic development (14, 15) and oncogenesis (16) through various signaling molecules including Frizzled receptors (17), , and Dickkopf protei...

Monoacylglycerol lipase (MAGL) is a major serine hydrolase that hydrolyzes 2-arachidonoylglycerol (2-AG) to arachidonic acid (AA) and glycerol in the brain. Because 2-AG and AA are endogenous biologically active ligands in the brain, inhibition of MAGL is an attractive therapeutic target for CNS disorders, particularly neurodegenerative diseases. In this study, we report the structure-based drug design of novel piperazinyl pyrrolidin-2-ones starting from our hit compounds 2a and 2b. By enhancing the interaction of the piperazinyl pyrrolidin-2-one core and its substituents with the MAGL enzyme via design modifications, we identified a potent and reversible MAGL inhibitor, compound (R)-3t. Oral administration of compound (R)-3t to mice decreased AA levels and elevated 2-AG levels in the brain.

We investigated the effects of extracts of Benifuuki (a tea cultivar that contains methylated catechins such as epigallocatechin-3-O-(3-O-methyl) gallate (EGCG3”Me)) in mice fed a high-fat/high-sucrose (HF/HS) diet. This tea cultivar was then compared with an extract of Yabukita (a popular tea cultivar that lacks methylated catechins). For 6 weeks, C57BL/6J mice were fed either HF/HS diet with or without tea extracts from tea cultivars, which contained almost identical ingredients except for methylated catechins (i.e., Yabukita (0.2% and 1%) or Benifuuki (0.2% and 1%) extract powders). Supplementation with Benifuuki 0.2% markedly lowered plasma levels of TG and NEFAs compared with mice supplemented with Yabukita 0.2%. The diet containing Benifuuki 1% decreased adipose tissue weights, liver TG, and expression of lipogenic genes in the liver. These results suggested that Benifuuki had much greater lipid-lowering effects than Yabukita. Taken together, these data suggest that methylated catechins direct the strong lipid-lowering activity of Benifuuki.

Although understanding the high-resolution spatial distribution of bioactive small molecules is indispensable for elucidating their biological or pharmacological effects, there has been no analytical technique that can easily detect the naïve molecular localization in mammalian tissues. We herein present a novel in situ label-free imaging technique for visualizing bioactive small molecules, using a polyphenol. We established a 1,5-diaminonaphthalene (1,5-DAN)-based matrix-assisted laser desorption/ionization-mass spectrometry imaging (MALDI-MSI) technique for visualizing epigallocatechin-3-O-gallate (EGCG), the major bioactive green tea polyphenol, within mammalian tissue micro-regions after oral dosing. Furthermore, the combination of this label-free MALDI-MSI method and a standard-independent metabolite identification method, an isotopic fine structure analysis using ultrahigh-resolution mass spectrometer, allows for the visualization of spatially-resolved biotransformation based on simultaneous mapping of EGCG and its phase II metabolites. Although this approach has limitations of the detection sensitivity, it will overcome the drawbacks associated with conventional molecular imaging techniques, and could contribute to biological discovery.