Biomedical subjects

Satoshi B Sato

Publications and source records attributed to Satoshi B Sato.

8 recordsLinked to original sources

PubMed2006

Increased lipid rafts and accelerated lipopolysaccharide-induced tumor necrosis factor-alpha secretion in Abca1-deficient macrophages.

Lipid rafts on the cell surface are believed to be very important as platforms for various cellular functions. The aim of this study was to know whether defective lipid efflux may influence lipid rafts on the cell surface and their related cellular functions. We investigated macrophages with defective lipid efflux from ATP binding cassette transporter A1-deficient (Abca1-KO) mice. Lipid rafts were evaluated by the following two novel probes: a biotinylated and protease (subtilisin Carlsberg)-nicked derivative of theta-toxin and a fluorescein ester of polyethylene glycol-derived cholesterol. Lipid rafts in Abca1-KO macrophages were increased, as demonstrated by both probes. Moreover, activities of nuclear factor kappaB, mRNA and intracellular distribution, and secretion of tumor necrosis factor-alpha (TNF-alpha) were examined after stimulation by lipopolysaccharides (LPSs). LPS-induced responses of the activation of nuclear factor kappaB and TNF-alpha were more prompt and accelerated in the Abca1-KO macrophages compared with wild-type macrophages. Modification of lipid rafts by cyclodextrin and nystatin corrected the abnormal response, suggesting an association between the increased lipid rafts and abnormal TNF-alpha secretion. We report here that Abca1-KO macrophages with defective lipid efflux exhibited increased lipid rafts on the cell surface and accelerated TNF-alpha secretion.

ATP Binding Cassette Transporter 1↗

PubMed2005

Role of membrane sphingomyelin and ceramide in platform formation for Fas-mediated apoptosis.

Engagement of the Fas receptor (CD95) initiates multiple signaling pathways that lead to apoptosis, such as the formation of death-inducing signaling complex (DISC), activation of caspase cascades, and the generation of the lipid messenger, ceramide. Sphingomyelin (SM) is a major component of lipid rafts, which are specialized structures that enhance the efficiency of membrane receptor signaling and are a main source of ceramide. However, the functions of SM in Fas-mediated apoptosis have yet to be clearly defined, as the responsible genes have not been identified. After cloning a gene responsible for SM synthesis, SMS1, we established SM synthase-defective WR19L cells transfected with the human Fas gene (WR/Fas-SM(-)), and cells that have been functionally restored by transfection with SMS1 (WR/Fas-SMS1). We show that expression of membrane SM enhances Fas-mediated apoptosis through increasing DISC formation, activation of caspases, efficient translocation of Fas into lipid rafts, and subsequent Fas clustering. Furthermore, WR/Fas-SMS1 cells, but not WR/Fas-SM(-) cells, showed a considerable increase in ceramide generation within lipid rafts upon Fas stimulation. These data suggest that a membrane SM is important for Fas clustering through aggregation of lipid rafts, leading to Fas-mediated apoptosis.

Animals↗

PubMed2005

Imaging lipid rafts.

Lipid rafts are plasma membrane microdomains enriched in sphingolipids and cholesterol. These domains have been suggested to serve as platforms for various cellular events, such as signaling and membrane trafficking. However, little is known about the distribution and dynamics of lipids in these microdomains. Here we report investigations carried out using recently developed probes for the lipid components of lipid rafts: lysenin, a sphingomyelin-binding protein obtained from the coelomic fluid of the earthworm Eisenia foetida; and the fluorescein ester of poly(ethyleneglycol) cholesteryl ether (fPEG-Chol), which partitions into cholesterol-rich membranes. Lysenin reveals that the organization of sphingomyelin differs between different cell types and even between different membrane domains within the same cell. When added to live cells, fPEG-Chol is distributed exclusively on the outer leaflet of the plasma membrane and is clustered dynamically upon activation of receptor signaling. The surface-bound fPEG-Chol is slowly internalized via a clathrin-independent pathway into endosomes with lipid raft markers.

Cholesterol↗

PubMed2004

Ultrastructural study of echinocytes induced by poly (ethylene glycol)-cholesterol.

Poly (ethylene glycol)-cholesterol (PEG-Chol) consists of a hydrophilic PEG and hydrophobic cholesterol moiety. When PEG-Chol was applied to erythrocytes, the reagent quantitatively induced protrusions by exclusively distributing in the outer monolayer of the membrane. This kind of response has been regarded as a general response that reduces the stress of expansion of the outer monolayer. However, the relationship between the membrane architecture and the distribution of such molecules is unknown. In this study, we examined the distribution of tagged PEG-Chol along the shape change pathway. The echinocytic shape was initiated by the initial formation of bumps on the rim of the discoid, which subsequently elongated as protrusions. These protrusions contained aggregates of granular structures, which appeared to accommodate the increase in the outer monolayer area. At higher concentrations, PEG-Chol further induced sphero-echinocytosis that resulted in numerous branched protrusion processes. We found that PEG-Chol was exclusively distributed in these protrusions and, in particular, accumulated at the tips. These results suggested that externally intercalated PEG-Chol was sequestrated from erythrocytes as membrane protrusions through an as-yet-unknown mechanism.

Cell Surface Extensions↗

PubMed2004

Distribution and transport of cholesterol-rich membrane domains monitored by a membrane-impermeant fluorescent polyethylene glycol-derivatized cholesterol.

Cholesterol-rich membrane domains function in various membrane events as diverse as signal transduction and membrane traffic. We studied the interaction of a fluorescein ester of polyethylene glycol-derivatized cholesterol (fPEG-Chol) with cholesterol-rich membranes both in cells and in model membranes. Unlike filipin and other cholesterol probes, this molecule could be applied as an aqueous dispersion to various samples. When added to live cells, fPEG-Chol distributed exclusively in the outer plasma membrane leaflet and was enriched in microdomains that dynamically clustered by the activation of receptor signaling. The surface-bound fPEG-Chol was slowly internalized via clathrin-independent pathway into endosomes together with lipid raft markers. Noteworthy, fPEG-Chol could be microinjected in the living cells in which we found Golgi apparatus as the sole major organelle to be labeled. PEG-Chol, thus, provides a novel, sensitive probe for unraveling the dynamics of cholesterol-rich microdomains in living cells.

Biological Transport↗

PubMed2003

Oligomerization and pore formation of a sphingomyelin-specific toxin, lysenin.

Lysenin is a novel protein derived from coelomic fluid of the earthworm Eisenia foetida, which specifically recognizes sphingomyelin and induces cytolysis. The mechanism underlying lysenin-induced cell lysis has not been clarified. In this report we studied the interaction of lysenin with red blood cells as well as artificial liposomes. Our results showed that lysenin bound membranes and assembled to SDS-resistant oligomers in a sphingomyelin-dependent manner, leading to the formation of pores with a hydrodynamic diameter of approximately 3 nm. Antibody scanning analysis suggested that the C-terminal region of lysenin was exposed, whereas the N-terminal was hidden in the isolated oligomer complex. Differential scanning calorimetry revealed that lysenin interacted with both hydrophilic head group and hydrophobic hydrocarbon tails of sphingomyelin. Oligomerization but not binding was affected by the amide-linked fatty acid composition of sphingomyelin, suggesting the role of membrane fluidity in the oligomerization step.

Amino Acid Sequence↗

PubMed2003

Actin-rich spherical extrusion induced in okadaic acid-treated K562 cells by crosslinking of membrane microdomains.

Interconnection between surface microdomains and the actin cytoskeleton is vital to various cellular activities. We studied the responses of okadaic acid (OKA)-treated K562 leukemia cells to crosslinking of membrane microdomains. Although OKA alone induced clustering of surface-bound F-actin, addition of a biotinylated poly(ethylene glycol) derivative of cholesterol (bPEG-Chol) and subsequent binding of streptavidin (SA) further induced accumulation of the clusters, resulting in the formation of a spherical cell extrusion. This extrusion was also induced by direct crosslinking of a raft marker, CD59, and ganglioside GM1. In addition, we found that knockout of the gene encoding Fyn kinase inhibited formation of the spherical extrusion in murine T-cells. In bPEG-Chol/SA-treated cells, CD59, ganglioside GM1, and clathrin/AP-2 were all accumulated on the surface of the actin-rich extrusion, whereas dynamin and transferrin receptors were unaffected. Intermediate filaments, mitochondria, and other vesicles also accumulated. These results suggest that crosslinking of membrane domains exaggerates the linkage between actin and a defined set of membrane proteins in OKA-treated cells.

Actins↗

PubMed2002

Cinnamycin (Ro 09-0198) promotes cell binding and toxicity by inducing transbilayer lipid movement.

Cinnamycin is a unique toxin in that its receptor, phosphatidylethanolamine (PE), resides in the inner layer of the plasma membrane. Little is known about how the toxin recognizes PE and causes cytotoxicity. We showed that cinnamycin induced transbilayer phospholipid movement in target cells that leads to the exposure of inner leaflet PE to the toxin. Model membrane studies revealed that cinnamycin induced transbilayer lipid movement in a PE concentration-dependent manner. Re-orientation of phospholipids was accompanied by an increase in the incidence of beta-sheet structure in cinnamycin. When the surface concentration of PE was high, cinnamycin induced membrane re-organization such as membrane fusion and the alteration of membrane gross morphology. These results suggest that cinnamycin promotes its own binding to the cell and causes toxicity by inducing transbilayer lipid movement.

Anti-Bacterial Agents↗