Makoto Seo

A possible role of lysophospholipids produced by calcium-independent phospholipase A2 in membrane-raft budding and fission

Phospholipase A(2) (PLA(2)) not only plays a role in the membrane vesiculation system but also mediates membrane-raft budding and fission in artificial giant liposomes. This study aimed to demonstrate the same effects in living cells. Differentiated Caco-2 cells were cultured on filter membranes. MDCK cells were challenged with Influenza virus. The MDCK cultures were harvested for virus titration with a plaque assay. Alkaline phosphatase (ALP), a membrane-raft associated glycosylphosphatidylinositol (GPI)-anchored protein, was 70% released by adding 0.2 mmol/l lysophosphatidylcholine, which was abolished by treatment with a membrane-raft disrupter, methyl-beta-cyclodextrin. Activation of calcium-independent PLA(2) (iPLA(2)) by brefeldin A increased the apical release of ALP by approximately 1.5-fold (p<0.01), which was blocked by PLA(2) inhibitor bromoenol lactone (BEL). BEL also reduced Influenza virus production into the media (<10%) in the MDCK culture. These results suggest that cells utilize inverted corn-shaped lysophospholipids generated by PLA(2) to modulate plasma membrane structure and assist the budding of raft-associated plasma membrane particles, which virus utilizes for its budding. Brush borders are enriched with membrane-rafts and undergo rapid turnover; thus, PLA(2) may be involved in the regulatory mechanism in membrane dynamism. Further, iPLA(2) may provide a therapeutic target for viral infections.

Statins Activate Human PPARα Promoter and Increase PPARα mRNA Expression and Activation in HepG2 Cells

Statins increase peroxisome proliferator-activated receptor α (PPARα) mRNA expression, but the mechanism of this increased PPARα production remains elusive. To examine the regulation of PPARα production, we examined the effect of 7 statins (atorvastatin, cerivastatin, fluvastatin, pitavastatin, pravastatin, rosuvastatin, and simvastatin) on human PPARα promoter activity, mRNA expression, nuclear protein levels, and transcriptional activity. The main results are as follows. (1) Majority of statins enhanced PPARα promoter activity in a dose-dependent manner in HepG2 cells transfected with the human PPARα promoter. This enhancement may be mediated by statin-induced HNF-4α. (2) PPARα mRNA expression was increased by statin treatment. (3) The PPARα levels in nuclear fractions were increased by statin treatment. (4) Simvastatin, pravastatin, and cerivastatin markedly enhanced transcriptional activity in 293T cells cotransfected with acyl-coenzyme A oxidase promoter and PPARα/RXRα expression vectors. In summary, these data demonstrate that PPARα production and activation are upregulated through the PPARα promoter activity by statin treatment.

The Fibronectin RGD Motif Is Required for Multiple Angiogenic Events During Early Embryonic Development

In the present study we demonstrated that inactivation of the RGD motif in mice results in a severe circulation defect that differs from the defect seen in mice lacking FN. Our findings indicate that FN is essential for both vasculogenesis and angiogenesis, and the RGD motif plays specific roles in angiogenesis.

Direct and simple fluorescence detection method for oxidized lipoproteins.

The quantification of low-density lipoprotein (LDL) and high-density lipoprotein (HDL) is currently one of the most important clinical measurements for characterizing metabolic syndrome. However, recent studies have revealed additional factors that may be more strongly associated with the coronary heart disease than simple measurement of LDL or HDL levels, such as small dense (sd) LDL particles and oxidized LDL or HDL particles. Although several methods using enzyme-antibody detection systems or fluorescent probes have been devised to characterize these factors, such methods are expensive to implement for clinical measurements. Here, we present a straightforward analytical method for direct quantitation of oxidized lipoproteins by fluorescence spectrometry, with excitation in the UV (365 +/- 10 nm) or visible (470 +/- 10 nm) range and emission detected at 450 +/- 30 nm or 535 +/- 15 nm. This method can be readily applied for clinical measurement in patients with dyslipidemia using only 1 microL of 1 mg/mL of lipoprotein and without the need for any expensive detection antibodies. Using this new technique, biological samples from patients with dyslipidemia showed higher fluorescence intensities than samples from normal subjects when detecting oxidized LDL and light HDL (d = 1.063-1.125 g/mL), whereas samples from patients with dyslipidemia showed lower fluorescence intensities than samples from normal subjects when measuring oxidized heavy HDL (d = 1.125-1.210 g/mL) levels.

Acarbose attenuates postprandial hyperlipidemia: investigation in an intestinal absorptive cell model

Acarbose attenuates postprandial hyperlipidemia: investigation in an intestinal absorptive cell model Takanari Nakano⁎, Ikuo Inoue, Makoto Seo, Seiichiro Takahashi, Tsugikazu Komoda, Shigehiro Katayama Department of Biochemistry, Faculty of Medicine, Saitama Medical University, 350-0495 Japan CURE/UCLA & BBRI, West Los Angeles VA Medical Center, Los Angeles, CA 90073, USA Department of Diabetes and Endocrinology, Faculty of Medicine, Saitama Medical University, 350-0495 Japan Received 13 January 2009; accepted 3 February 2009

Geometrical Separation Method for Lipoproteins Using Bioformulated-Fiber Matrix Electrophoresis: Size of High-Density Lipoprotein Does Not Reflect Its Density

The increasing number of patients with metabolic syndrome is a critical global problem. In this study, we describe a novel geometrical electrophoretic separation method using a bioformulated-fiber matrix to analyze high-density lipoprotein (HDL) particles. HDL particles are generally considered to be a beneficial component of the cholesterol fraction. Conventional electrophoresis is widely used but is not necessarily suitable for analyzing HDL particles. Furthermore, a higher HDL density is generally believed to correlate with a smaller particle size. Here, we use a novel geometrical separation technique incorporating recently developed nanotechnology (Nata de Coco) to contradict this belief. A dyslipidemia patient given a 1-month treatment of fenofibrate showed an inverse relationship between HDL density and size. Direct microscopic observation and morphological observation of fractionated HDL particles confirmed a lack of relationship between particle density and size. This new technique may improve diagnostic accuracy and medical treatment for lipid related diseases.

Determination of Circulating Native and Denaturated HDL Concentrations and its Clinical Implications

Publisher Summary The high susceptibility of high density lipoprotein (HDL) to oxidation is associated with its antiatherogenic effects. By acting as a sacrificial target for oxidation, HDL can prevent low density lipoprotein (LDL) from oxidation, which plays a key role in the progression of atherosclerosis. Oxidized HDL increases in the early stage of the development of atherosclerosis. The high oxidative susceptibility of HDL not only contributes to its antiatherogenic effects but also may serve as an early marker for oxidative stress as HDL acts as a sacrificial target for oxidation. HDL can reduce the level of oxidized LDL by accepting the lipid hydroperoxides of oxidized LDL. The oxidation is mediated by a variety of radicals or enzymes. Assays for oxidized HDL have been developed and applied for the measurement of clinical specimens. Oxidized HDL is decreased in patients who are supposed to have increased oxidative stress. Oxidized HDL may be removed from the circulation rapidly considering the more rapid clearance of oxidized HDL than native HDL. Moreover, decreased oxidized HDL in patients suggests that the pathway to clear or scavenge oxidized HDL is accelerated in some disease conditions.