Procyanidin dimer B2 [epicatechin-(4beta-8)-epicatechin] suppresses the expression of cyclooxygenase-2 in endotoxin-treated monocytic cells

The anti-inflammatory activity of the predominant procyanidin dimer in cocoa, dimer B2, was investigated in this study. Pretreatment of the procyanidin dimer B2 reduced COX-2 expression induced by the endotoxin lipopolysaccharide (LPS) in differentiated human monocytic cells (THP-1) in culture. To further elucidate the underlying mechanism of COX-2 inhibition by procyanidin, we examined their effects on the activation of extracellular signal-regulated protein kinase (ERK), Jun-terminal kinase (JNK), and p38 mitogen-activated protein kinase (MAPK), which are upstream enzymes known to regulate COX-2 expression in many cell types. Pretreatment with procyanidin dimer B2 decreased the activation of ERK, JNK, and p38 MAPK. In addition, procyanidin dimer B2 suppressed the NF-kappaB activation through stabilization of IkappaB proteins, suggesting that these signal-transducing enzymes could be potential targets for procyanidin dimer B2. By affecting the expression rather than the activity of COX-2, these in vitro data reported herein give further evidence on the anti-inflammatory protection by procyanidins.     

Identification of a nuclear localization signal in the retinitis pigmentosa-mutated RP26 protein, ceramide kinase-like protein

Retinitis pigmentosa (RP) is a genetically heterogeneous disease characterized by degeneration of the retina. A mutation in a new ceramide kinase (CERK) homologous gene, named CERK-like protein (CERKL), was found to cause autosomal recessive retinitis pigmentosa (RP26). Here, we show a point mutation of one of two putative nuclear localization signal (NLS) sequences inhibited the nuclear localization of the protein. Furthermore, the tetra-GFP-tagged NLS, which cannot passively enter the nucleus, was observed not only in the nucleus but also in the nucleolus. Our results provide the first evidence of the active nuclear import of CERKL and suggest that the identified NLS might be responsible for nucleolar retention of the protein. As recent studies have shown other RP-related proteins are localized in the nucleus or the nucleolus, our identification of NLS in CERKL suggests that CERKL likely plays important roles for retinal functions in the nucleus and the nucleolus.     

Cell density-dependent nuclear/cytoplasmic localization of NORPEG (RAI14) protein

NORPEG (RAI14), a developmentally regulated gene induced by retinoic acid, encodes a 980 amino acid (aa) residue protein containing six ankyrin repeats and a long coiled-coil domain [Kutty et al., J. Biol. Chem. 276 (2001), pp. 2831-2840]. We have expressed aa residues 1-287 of NORPEG and used the recombinant protein to produce an anti-NORPEG polyclonal antibody. Confocal immunofluorescence analysis showed that the subcellular localization of NORPEG in retinal pigment epithelial (ARPE-19) cells varies with cell density, with predominantly nuclear localization in nonconfluent cells, but a cytoplasmic localization, reminiscent of cytoskeleton, in confluent cultures. Interestingly, an evolutionarily conserved putative monopartite nuclear localization signal (P(270)KKRKAP(276)) was identified by analyzing the sequences of NORPEG and its orthologs. GFP-NORPEG (2-287 aa), a fusion protein containing this signal, was indeed localized to nuclei when expressed in ARPE-19 or COS-7 cells. Deletion and mutation analysis indicated that the identified nuclear localization sequence is indispensable for nuclear targeting.     

Mineralocorticoid receptor is involved in the regulation of genes responsible for hepatic glucose production

The mineralocorticoid receptor (MR) is expressed in kidney and plays a central role in the control of sodium, homeostatic fluid, and blood pressure. It has also been implicated in other functions in cardiovascular system, central nervous system, and adipose tissue. This study revealed a novel role of MR in the gene regulation related to hepatic glucose production. RNAi-mediated MR silencing led to a decrease in the expression of glucose-6-phosphatase (G6Pase), phosphoenolpyruvate carboxykinase, and fructose-1,6-bisphosphatase 1, the enzymes known to be involved in glucose production in liver. The MR-specific antagonists also down-regulated the expression of G6Pase, while the specific agonist enhanced G6Pase expression. These observations, for the first time, revealed a novel role for MR and its ligands in the regulation of de novo glucose synthesis in hepatocytes. It also suggests the potential of liver-specific MR modulation for the treatment of hyperglycemia.     

The human myeloperoxidase gene is regulated by LXR and PPARalpha ligands

Myeloperoxidase (MPO) is an oxidant-generating enzyme expressed in macrophages and implicated in atherosclerosis and cholesterol homeostasis. LXRalpha and PPARalpha regulate genes involved in cholesterol metabolism and the inflammatory response in macrophages. Here, we examine the effect of LXR and PPARalpha ligands on MPO expression. LXR and PPARalpha, as heterodimers with RXR, are shown to bind overlapping sites in an Alu receptor response element (AluRRE) in the MPO promoter. The LXR ligand T0901317 suppresses MPO mRNA expression in primary human macrophages, and in bone marrow cells and macrophages from huMPO transgenic mice. The PPARalpha ligand GW9578 downregulates MPO expression in GMCSF-macrophages, while upregulating in MCSF-macrophages. In contrast, the mouse MPO gene, which lacks the primate-specific AluRRE, is not regulated by LXR or PPARalpha ligands. These findings identify human MPO as a novel LXR and PPARalpha target gene, consistent with the role of these receptors in regulation of proinflammatory genes in macrophages.