单唾液酸四己糖神经节苷脂早期治疗中重度缺氧缺血性脑病新生儿的疗效

目的 观察单唾液酸四己糖神经节苷脂(GM1)治疗中重度缺氧缺血性脑病(HIE)新生儿的疗效.方法 中重度HIE新生儿86例随机分为2组:常规治疗组(对照组)42例;GM1干预组44例.二组患儿均予常规治疗.干预组在常规治疗的基础上早期(生后6 h内)应用GM1(20 mg)静脉滴注治疗.1次/d,10～14 d为1个疗程.治疗前治疗1个疗程和28 d 检查2组头颅CT、行为神经评分(NBNA),12个月时采用首都儿科研究所儿心量表(CDCC)评定智力/运动发育指数.采用Statistic View 5.0软件进行t检验和χ2检验.结果 GM1干预组头颅CT影像学脑结构的恢复及NBNA评分均明显优于对照组(Pa<0.05),MDI、PDI等各项指标与对照组比较均有明显改善(Pa<0.01).结论 早期应用GM1治疗中重度HIE有明显疗效.     

单唾液酸四己糖神经节苷脂钠致严重过敏反应2例

2例患者静脉滴注单唾液酸四己糖神经节苷脂钠引致严重过敏反应。第1例为91岁男性帕金森病患者,静脉滴注单唾液酸四己糖神经节苷脂钠40mg,滴注10min后出现寒战、心悸、全身冷汗,BP由入院时的155/70mmHg降至130/60mmHg。第2例为84岁男性慢性肾功能衰竭合并脑缺血患者静脉滴注60mg,滴注结束后5min出现胸闷、心悸、头晕、寒战、四肢厥冷、全身冷汗、神志淡漠,BP由入院时的150/80mmHg降至90/60mmHg。2例患者经过抗过敏治疗和对症治疗后症状消失。停用该药后未再出现类似症状。     

Characterization of the binding of cholera toxin to ganglioside GM1 immobilized onto microtitre plates

Ganglioside GM1 is the receptor for cholera toxin on cell surfaces, and the binding of cholera toxin to GM1 immobilized on microtitre plates has been reported previously by several authors as an assay for the toxin (GM1-ELISA). This assay has been examined in detail. Results were independent of the adsorption solvent for GM1 (methanol or phosphate-buffered saline), the pH of aqueous solvents (7.4-10.2) and the temperature (4-37 degrees C). High and near-maximal rates of absorbance change in the assay were found for lower concentrations of GM1 (100 ng ml(-1)) and for shorter incubation times (a few hours) than reported in the literature. A method was devised to provide a semi-quantitative estimate of the amount of GM1 bound to the plate; this was found to be in the low nanogram range. Binding of cholera toxin to the immobilized GM1 required > or =1.5 h for maximal assay results. The failure of free GM1 in solution to displace cholera toxin once bound to immobilized GM1 indicated that binding to immobilized GM1 is irreversible in the time frame of the experiment. Data from the literature support the very slow dissociation rates of the toxin-GM1 complex.     

Determination of structural and functional overlap/divergence of five proto-type galectins by analysis of the growth-regulatory interaction with ganglioside GM1 in silico and in vitro on human neuroblastoma cells

The growth-regulatory interplay between ganglioside GM1 on human SK-N-MC neuroblastoma cells and an endogenous lectin provides a telling example for glycan (polysaccharide) functionality. Galectin-1 is the essential link between the sugar signal and the intracellular response. The emerging intrafamily complexity of galectins raises the question on defining extent of their structural and functional overlap/divergence. We address this problem for proto-type galectins in this system: ganglioside GM1 as ligand, neuroblastoma cells as target. Using the way human galectin-1 interacts with this complex natural ligand as template, we first defined equivalent positioning for distinct substitutions in the other tested proto-type galectins, e.g., Lys63 vs. Leu60/Gln72 in galectins-2 and -5. As predicted from our in silico work, the tested proto-type galectins have affinity for the pentasaccharide of ganglioside GM1. In contrast to solid-phase assays, cell surface presentation of the ganglioside did not support binding of galectin-5, revealing the first level of regulation. Next, a monomeric proto-type galectin (CG-14) can impair galectin-1-dependent negative growth control by competitively blocking access to the shared ligand without acting as effector. Thus, the quaternary structure of proto-type galectins is an efficient means to give rise to functional divergence. The identification of this second level of regulation is relevant for diagnostic monitoring. It might be exploited therapeutically by producing galectin variants tailored to interfere with galectin activities associated with the malignant phenotype. Moreover, the given strategy for comparative computational analysis of extended binding sites has implications for the rational design of galectin-type-specific ligands.     

The effects of

Antibodies against hydrophobic antigens are common in several autoimmune diseases. However, detection of such antibodies by standard immune-assays, such as ELISA, is problematic, in part because of the problems with coating hydrophobic molecules onto polystyrene multi-well plates. We describe a novel method of stably associating hydrophobic antigens to ELISA plates. By mixing the antigen with a hydrophobic molecule containing a hydrophilic anchor, we generate mixed lipid aggregates that can attach to ELISA plates, and are resistant to detergent wash. Using the ganglioside GM-1 and phosphatidylethanolamine conjugated to the hapten DNP (dinitrophenyl) as model antigens, we show that hydrophobic antigens incorporated into mixed lipid aggregates expose their antigenic determinants in a correct configuration. The detection limit of both GM-1 and DNP-PE was considerably improved compared to when these antigens were coated on ELISA plates using organic solvents. Furthermore GM-1 incorporated into mixed lipid aggregates can be detected by specific antibodies in patient serum. The method of incorporating hydrophobic antigens into mixed lipid aggregates for stable association to ELISA plates can presumably be applied to a vast array of hydrophobic antigens, and may well be developed into a large scale screening system for serum reactivity towards different hydrophobic antigens.     

Sensitive detection of hydrophobic antigens using a novel lipid-aggregate based ELISA

Antibodies against hydrophobic antigens are common in several autoimmune diseases. However, detection of such antibodies by standard immune-assays, such as ELISA, is problematic, in part because of the problems with coating hydrophobic molecules onto polystyrene multi-well plates. We describe a novel method of stably associating hydrophobic antigens to ELISA plates. By mixing the antigen with a hydrophobic molecule containing a hydrophilic anchor, we generate mixed lipid aggregates that can attach to ELISA plates, and are resistant to detergent wash. Using the ganglioside GM-1 and phosphatidylethanolamine conjugated to the hapten DNP (dinitrophenyl) as model antigens, we show that hydrophobic antigens incorporated into mixed lipid aggregates expose their antigenic determinants in a correct configuration. The detection limit of both GM-1 and DNP-PE was considerably improved compared to when these antigens were coated on ELISA plates using organic solvents. Furthermore GM-1 incorporated into mixed lipid aggregates can be detected by specific antibodies in patient serum. The method of incorporating hydrophobic antigens into mixed lipid aggregates for stable association to ELISA plates can presumably be applied to a vast array of hydrophobic antigens, and may well be developed into a large scale screening system for serum reactivity towards different hydrophobic antigens.     

Fucosyl-GM1 expression and amyloid-beta protein accumulation in PC12 cells

Gangliosides, sialic acid-containing glycosphingolipids, are ubiquitously expressed in all eukaryotic cells and are localized primarily in the plasma membrane. For a rat pheochromocytoma cell line, PC12, which has been used frequently as a model for investigating events leading to neuronal differentiation, it is generally thought that GM1 is a major ganglioside, based on reactivity with the probe cholera toxin B subunit (Ctxb). From a series of biochemical studies, however, it has been reported that no GM1 is expressed in PC12 cells. In this study, we have reevaluated GM1 expression and Ctxb reactivity in PC12 cells and a subcloned line, PC12D cells. Flow cytometric analysis with Ctxb revealed that about 30-50% of PC12 cells were reactive with Ctxb. However, a detailed biochemical analysis showed that PC12 cells express abundantly a different ganglioside, fucosyl-GM1, instead of GM1, and the reactivity of Ctxb in the PC12 cells actually arose from its interaction with fucosyl-GM1, which also interacts with this ligand. Because it has been claimed that amyloid-beta protein (Abeta) interacts with GM1 in PC12 cells to provide "seeding" for amyloid to accumulate, we further evaluated this possibility and found that Abeta is mostly likely interacting with fucosyl-GM1 in this cell line. Our data thus suggest that a specific interaction may occur between Abeta and fucosyl-GM1 for the accumulation of amyloid in PC12 cells.     

Molecular mimicry: sensitization of Lewis rats with Campylobacter jejuni lipopolysaccharides induces formation of antibody toward GD3 ganglioside

Recently we have reported cases of demyelinating inflammatory neuropathy showing elevated titers of anti-GD3 antibodies, which occurs rarely in Guillain-Barré syndrome. To examine the correlation between the anti-GD3 antibody titer and Campylobacter jejuni infection, we sensitized female Lewis rats with lipopolysaccharides (LPSs) from serotype HS19 of C. jejuni and examined changes in nerve conduction velocity and nerve conduction block (P/D ratio). After 16 weeks of sensitization, animals revealed decreases of nerve conduction velocity and conduction block (P/D ratio) and high titer of anti-GD3 antibodies. These anti-GD3 antibodies also blocked transmission in neuromuscular junctions of spinal cord-muscle cells cocultures. The GD3 epitope was verified to be located on the Schwann cell surface and nodes of Ranvier in rat sciatic nerve. To determine the target epitope for GD3 antibodies in causing nerve dysfunction, the LPS fraction containing the GD3 epitope was purified from the total LPS by using an anti-GD3 monoclonal antibody-immobilized affinity column. Subsequently, chemical analysis of the oligosaccharide portion was performed and confirmed the presence of a GD3-like epitope as having the following tetrasaccharide structure: NeuAcalpha2-8NeuAc2-3Galbeta1-4Hep. Our data thus support the possibility of a contribution of GD3 mimicry as a potential pathogenic mechanism of peripheral nerve dysfunction.     

高血压脑出血术后应用神经节苷脂的临床疗效分析

目的观察神经节苷脂治疗高血压脑出血术后的临床疗效。方法将80例高血压脑出血术后患者随机分为神经节苷脂组和对照组各40例,两组均给予常规治疗;神经节苷脂组在常规的治疗上应用神经节苷脂营养神经细胞;对照组仅给予常规治疗,比较两组患者颅内血肿+水肿体积、神经功能、日常生活能力恢复情况和临床疗效。结果与对照组比较,神经节苷脂组可以显著减少颅内水肿体积(P<0.05),改善患者神经功能的恢复情况(P<0.05)、日常生活能力(P<0.05)和临床疗效(P<0.05)。结论高血压脑出血术后应用神经节苷脂治疗,可以显著提高患者生存质量,临床疗效显著。     

单唾液酸四己糖神经节苷脂改善脑梗死患者神经功能缺损症状的疗效探讨

目的：探讨单唾液酸四己糖神经节苷脂改善脑梗死患者神经功能缺损症状的疗效,探讨其临床适用性。方法取2014年6月—2015年12月在该院就诊的124例脑梗死患者作为研究对象,随机分为对照组和实验组各62例,对照组给予常规治疗与康复训练,实验组在对照组的基础上给予单唾液酸四己糖神经节苷脂治疗,对比两组患者治疗前后NIHSS评分和治疗后的临床疗效。结果实验组患者治疗后NIHSS评分(10.42±4.33)明显低于对照组(15.11±4.78),差异有统计学意义(P<0.05)。对照组总有效(率)为43(69.35%),明显低于实验组57(91.94%),差异有统计学意义(P<0.05)。结论单唾液酸四己糖神经节苷脂能有效改善脑梗死患者神经功能缺损症状,临床疗效显著,值得临床推广应用。