鞘糖脂研究进展

鞘糖脂位于细胞膜脂质双分子层,是哺乳动物细胞膜上的必需组成成分之一,参与细胞的多种生物学活动,其生物学功能非常复杂。在免疫应答、细胞发育、细胞识别及分化中都发挥重要的作用。组织器官中鞘糖脂的异常表达往往与各种疾病具有明显的相关性。因此鞘糖脂的结构表征、构效关系研究以及与之相关的鞘糖脂生物合成及分解代谢途径的研究已成为近年来的热点问题。最近研究也表明鞘糖脂在疾病的发生发展及治疗过程中都具有至关重要的作用。对鞘糖脂的结构与功能的关系、糖脂的合成代谢以及糖脂的分离分析方法作一综述。     

真菌鞘糖脂的生物学功能及应用研究进展

丝状真菌作为一类重要的微生物，被广泛应用于发酵食品、工业酶和次生代谢物等工业生产中。真菌鞘糖脂主要由鞘氨醇、脂肪酸链和特殊的极性基团组成，根据极性基团的不同，分为中性鞘糖脂和酸性鞘糖脂两大类。鞘糖脂不仅参与真菌生长、细胞分化、增殖、细胞凋亡、逆境胁迫等重要生理活动，中性鞘糖脂还可作为功能性医药用品、化妆品和保健食品的重要活性组分。本文论述了真菌鞘糖脂的主要种类、结构、生物合成途径和及其参与丝状真菌生长、分化和响应逆境胁迫的生物学功能；探讨了真菌中性鞘糖脂作为抗菌肽的靶点和酸性鞘糖脂在开发抗真菌药物中的应用；同时还综述了中性鞘糖脂作为化妆品的保湿成分或保健食品的功能成分，在改善皮肤屏障功能和预防特应性皮炎中的重要作用的相关研究进展，尤其是来源于曲霉的中性鞘糖脂，可显著增强皮肤屏障功能，并可作为益生元预防肠道损伤；另外还探讨了曲霉尤其是米曲霉作为开发中性鞘糖脂生物资源的优势。     

妊娠期家兔子宫内膜的鞘糖脂表达

 妊娠期家兎子宫内膜的神经节苷脂(Gls)的含量明显低于动情期的,而中性鞘糖脂(NGSL)的含量则以妊娠中、晚期的最高,动情期最低。鞘糖脂组成变化最显著的是妊娠早期,由动情期到早孕GM_3从28.0％增加到52.7％,CMH.CDH由未测出分别增加到29.2％和21.9％,而糖链复杂的组分GD_3,GTlb和CPH的百分含量则明显减少,到妊娠中、晚期、短糖链组分逐渐减少,而复杂糖链组分渐增。中期妊娠内膜的(GIs)以GD_3为主要组分,占45％,明显高于其它各期。NGSL在妊娠中、晚期CPH增高达70％,与动情期水平相当。结果提示,妊娠期间子宫内膜的鞘糖脂含量与组成均发生明显变化,这些变化可能与子宫功能密切相关。特别是早孕对的变化,推测与子宫内膜和胚泡的识别,粘连特性的获得有关。     

糖苷合酶应用于糖蛋白和鞘糖脂的合成

糖苷合成酶是糖苷酶的亲核体氨基酸突变酶,催化寡糖的高效合成,可应用于寡糖的大规模生产。最近,糖苷合成酶被成功地应用于两类重要的生物分子——糖蛋白和鞘糖脂的高效合成,这必将对糖生物学和制药业的发展起到重要的推动作用。     

Ceramide glycosylation and fatty acid hydroxylation influence serological reactivity in Trypanosoma cruzi glycosphingolipids

Ceramide mono (CMH) or dihexoside (CDH) fractions from Trypanosoma cruzi (Dm28c clone) were identified as glucosyl and lactosylceramides containing non-hydroxylated fatty acids. The di-glycosylated form was much more efficiently recognized by sera from T. cruzi-immunized rabbits, indicating that glycosylation influences antigenicity. Fatty acid hydroxylation was also a determinant of serological reactivity, since an alpha-hydroxylated CMH, only present at the Y clone, was recognized by the hyperimmune sera. In summary, these data indicate that T. cruzi CMHs with non-hydroxylated fatty acids are unable to induce antibody responses in animal hosts, which is reverted by the addition of a sugar residue or an alpha-hydroxyl group.     

Covalent attachment of immunoglobulins to liposomes via glycosphingolipids

We describe a method for covalent binding of proteins to large unilamellar liposomes which involves the periodate oxidation of glycosphingolipids in the vesicle membrane. Proteins such as IgG and F(ab′)₂ may then be attached to the aldehyde groups on the glycolipid by Schiff-base formation at pH 9.5 and reduction with NaBH₄, or by reductive amination with NaBH₃CN at pH 8.4. Exposure of the vesicles to periodate, protein coupling and separation from unbound protein by a novel method of flotation in discontinuous dextran gradients does not release the vesicle contents when performed at pH 8.4. Studies on the oxidation of neutral glycolipid-containing vesicles, and on the oxidation of encapsulated glycerol 1-phosphate show that periodate influx into neutral vesicles during a 4 h exposure is appreciable at pH 5.5 but not at pH 8.4. Under optimal conditions, approx. 20% of the protein may be coupled to vesicles, and a ratio of 100–200 μg of protein/μmol of lipid is readily achieved. This method will be of great importance for the antibody-mediated targeting of vesicles to cells.     

First synthesis of two deoxy Lewisx pentaosyl glycosphingolipids

The Lewis^x–Lewis^x interaction has been increasingly studied, using a variety of techniques including nuclear magnetic resonance spectroscopy, mass spectrometry, vesicle adhesion, atomic force microscopy, and surface plasmon resonance spectroscopy. However, the detailed molecular mechanism of these weak, divalent cation dependent interactions remains unclear, and new models are needed to probe the nature of this phenomenon in term of key roles of the different hydroxyl groups on Lewis^x trisaccharide determinant involved in the Lewis^x–Lewis^x interaction. An interesting solution is to synthesize a series of Lewis^x pentaosyl glycosphingolipid derivatives in which one of the eight hydroxyl groups of Lewis^x trisaccharide is replaced by a hydrogen atom, and to test the adhesion induced by interaction of these derivatives, in order to gain insight into the functions played by the hydroxyl groups of the Lewis^x trisaccharide. This article describes the synthesis of 3d-deoxy and 4d-deoxy Lewis^x pentaosyl glycosphingolipids, to be used for study of the Lewis^x–Lewis^x interaction. Botao Fan: Deceased October 22, 2006     

The retarded rate of acid-catalyzed solvolysis of glycoside bonds between reducing-end glucose residue and ceramide in glycosphingolipids compared with that of glycoside bonds between hexopyranosides

Rates of acid-catalyzed solvolysis of glycoside bonds in glycosphingolipids were compared to establish a basis for conducting saccharide analysis. Permethylated globotetraosylceramide and asialogangliotriaosylceramide as model compounds for methylation and sugar composition analysis, respectively, were solvolyzed under acidic conditions and the sugar components thus obtained were determined at specified times by gas liquid chromatography, after they had been derivatized. Reducing-end glucose residues in both compounds were liberated more slowly than other sugar residues. Glycoside bonds between reducing-end glucose and ceramide in glycosphingolipids would thus appear to be more resistant towards acid-catalysed solvolysis than other glycoside bonds between hexopyranosides.     

Structural studies on the neutral glycosphingolipids of Manduca sexta

Glycosphingolipids (GSLs) have been implicated as playing major roles in cellular interactions and control of cell proliferation in muticellular organisms. Moreover GSLs and other sphingolipids such as sphingomyelins, ceramides and sphingosines serve a variety of roles in signal transduction. Hence, identification of structures of GSLs in different biota will shed light in understanding their physiological role. During this study, the major glycosphingolipid component present in the extracts of stage-12 and stage-17/18 metamorphosing adults of Manduca sexta was identified as mactosyl ceramide. We report the isolation of several ceramide disaccharides, a ceramide trisaccharide and a ceramide tetrasaccharide. The GSL structures were confirmed by high-resolution mass spectrometry and tandem mass spectrometry. The identity of the monosaccharides was proved using exoglycosidases. The predominant sphingosine chain-length varied from C-14 (tetradecasphing-4-enine) to C-16 (hexadecasphing-4-enine) in these GSLs. Sphingosines of both chain lengths were accompanied by their doubly unsaturated counterparts tetradecasphinga-4,6-diene and hexadecasphinga-4,6-diene. It is also interesting to note the presence of tetradecasphinganine and hexadecasphinganine in minute amounts in the form of a GSL in the extracts of M. sexta. The varying degrees of unsaturation in the sphingosine moiety of GSLs in M. sexta may be biologically significant in insect metamorphosis. The ceramide trisaccharides and ceramide tetrasaccharide belong to the arthro-series, The observation of fucose in the M. sexta GSLs is the first report of the presence of fucose in an arthroseries GSL.     

Temperature-induced modifications of glycosphingolipids in plasma membranes of Neurospora crassa

Plasma membranes isolated from a cell-wall-less mutant of Neurospora crassa grown at 37 and 15°C display large differences in lipid compositions. A free sterol-to-phospholipid ratio of 0.8 was found in 37°C membranes, while 15°C plasma membranes exhibited a ratio of nearly 2.0. Membranes formed under both growth conditions were found to contain glycosphingolipids. Cultures grown at the low temperature, however, were found to contain 6-fold higher levels of glycosphingolipids and a corresponding 2-fold reduction of phospholipid levels. The high glycosphingolipid content at 15°C compensates for the reduced levels of phospholipids in such a way that sterol/polar lipid ratios are almost the same in plasma membranes under the two growth conditions. Temperature-dependent changes in plasma-membrane phospholipid and glycosphingolipid species were also observed. Phosphatidylethanolamine levels were sharply reduced at 15°C, in addition to a moderate increase in levels of unsaturated phospholipid fatty acids. Glycosphingolipids contained high levels of long-chain hydroxy fatty acids, which constituted 75% of the total fraction at 37°C, but only 50% at 15°C. Compositional changes were also observed in the long-chain base component of glycosphingolipids with respect to growth temperature. Fluorescence polarization studies indicate that the observed lipid modifications in 15°C plasma membranes act to modulate bulk fluidity of the plasma-membrane lipids with respect to growth temperature. These studies suggest that coordinate modulation of glycosphingolipid, phospholipid and sterol content may be involved in regulation of plasma-membrane fluid properties during temperature acclimation.     

Binding specificities of the lectins from Helix pomatia, soybean and peanut against different glycosphingolipids in liposome membranes

The binding specificities of the lectins from Helix pomatia, soybean and peanut against glycosphingolipids containing β-linked terminal D-galactose and N-acetyl-D-galactosamine and the role of sialic acid as a modulator of the binding specificity were investigated. The test system used consisted of liposomes containing the glycosphingolipids and lectins coupled to gel columns. Of the investigated glycosphingolipids only ganglioside GM2 bound to soybean agglutinin while Helix pomatia agglutinin was found to bind only GA2. Peanut agglutinin showed good affinity both for ganglioside GM1 and its asialoderivative GA1.     

Possible roles of glycosphingolipids in lipid rafts

Recent studies have suggested that glycosphingolipid (GSL)-cholesterol microdomains in cell membranes may function as platforms for the attachment of lipid-modified proteins, such as glycosylphosphatidylinositol (GPI)-anchored proteins and src-family tyrosine kinases. The microdomains are proposed to be involved in membrane trafficking of GPI-anchored proteins and in signal transduction via src-family kinases. Here, the possible roles of GSLs in the physical properties of these microdomains, as well as in membrane trafficking and signal transduction, are discussed. Sphingolipid depletion inhibits the intracellular transport of GPI-anchored proteins in biosynthetic traffic and endocytosis via GPI-anchored proteins. Antibodies against GSLs as well as GPI-anchored proteins co-precipitate src-family kinases. Antibody-mediated cross-linking of GSLs, as well as that of GPI-anchored proteins, induces a transient increase in the tyrosine phosphorylation of several substrates. Thus, GSLs have important roles in lipid rafts.     

氯胺酮在体内的分布与代谢

目前,氯胺酮(ketamine,俗称K粉)的滥用现象日益严重,并有愈演愈烈之势,特别流行于舞厅等娱乐场所,对社会治安和经济建设造成极大危害。滥用毒品倾向呈现年轻化、男女平均化且女性滥用者的比例上升和用药多元化的趋势,由滥用鸦片类药物发展到多种药物的滥用。调查发现,毒品滥用中K粉比例达20%以上,短期可使语言迷糊,引起幻觉,长期可使人记忆力衰退及认知能力障碍,导致心功能损害。因吸食氯胺酮或服用含有氯胺酮的摇头丸、迷奸药而中毒死亡的案件逐年上升。本文综述了对氯胺酮在体内各个部分的分布与代谢,更多、更快、更好的为氯胺酮相关案件的检材选取,分析检测及体内过程的评价提供科学依据。