Titers of anti-glycosphingolipid, GM1 and GQ1bα antibodies in sera of demented patients, and age-matched controls.

<p>AD, Alzheimer's disease; VD, vascular dementia; C, controls without dementia.</p

HPTLC-immunostaining of brain gangliosides, such as GM1, GD1a, GD1b, GT1b, and GQ1bα (plate A) and GM1, GD3, GQ1b, and SGPG (plate B).

<p>Lane 1, 1 µg each of GM1, GD1a, GD1b, and GT1b; 2, 1 µg of GM1 and GQ1bα; 3, 1 µg each of GM1, GD3, and GQ1b; and 4, 1 µg of SGPG. a, orcinol sulfuric acid staining; b, c, and d, immunostaining; b-1 and b-2, control sera; c-1 and c-2, AD sera (Alzheimer’s disease); d-1 and d-2, VD sera (vascular dementia). Plates A and B were developed with CH₃Cl₃:CH₃OH:0.2%CaC₂•2H₂O (55∶45:10, v/v).</p

Characteristic of all cases.

<p>AD, Alzheimer's disease; VD, vascular dementia; C, controls without dementia.</p><p>MMSE, mini-mental state test;</p>*<p>,p<0.05 vs C.</p

Structures of glycosphingolipids cited in this study.

<p>Symbols used for gangliosides and other glycosphingolipids are based on the system of Svennerholm and the nomenclature recommended by IUPAC.</p

Effect of PR-ASG treatment on cellular ROS level and HTase activity in STZ-exposed cells.

<p>A) Initially, cells were incubated under one of these sets of experimental parameters: STZ-negative, PR-ASG-negative (control); 1 mM STZ (STZ group); or 1 mM STZ supplemented with 4 µg/mL of PR-ASG (STZ+PR-ASG). After 1 hr of incubation, the cells were pre-washed with fresh medium, stained with DCF-DA (1.0 µM), and incubated in darkness for 10 min. DCF-DA green fluorescence was viewed under a fluorescence microscope. B) DCF-DA green fluorescing cells were also quantitated by a Perkin-Elmer Victor V multilabel plate reader with excitation/emission filters set at 490 and 535 nm, and expressed as the percent of the total number of cells that exhibit DCA-DA green fluorescence. C) Cells (2.5×10⁵ in 1 mL of 10% serum- containing RPMI 1640 medium well) were cultured in12-well plates. One day later, the cells were divided into 3 groups fro 1 hour of exposure to one of the following in RPMI medium containing 1.0 µg/mL rat LDL; no STZ with no PR-ASG (control); 1 mM STZ (STZ); or 1 mM STZ with 4 µg/mL of PR-ASG (STZ+PR-ASG). Subsequently, cells were pre-washed; cultured for 24 hours in serum-free RPMI-1640 medium with 0 or 4 µg/mL of PR-ASG; harvested; and eventually used for enzyme preparation. The cellular supernatant produced during enzyme preparation was assessed for activity using an Alfresa auto HTLase assay kit (Alfresa Pharma Corp., Osaka, Japan). One unit of HTase activity is defined as 1 nmol of HT hydrolyzed in 1 min by 1 mg of protein. In B) and C), values were expressed as means ± SEM, n = 6 individual experiments, and analyzed by one-way ANOVA followed by Tukey's multiples comparison test and then Dunnet's test. The single asterisk denotes a statistically significant difference between STZ and STZ+PR-ASG (*p<0.01).</p

Final body weights, blood glucose concentrations, and serum parameters in non-diabetic and diabetic rats (3 weeks).

<p>One-way ANOVA was performed to analyze the variation between 3 groups of non-diabetic or diabetic treatment. The data were then analyzed using Tukey's multiples comparison test (if parametric). Values were expressed as means±SEM. Differences between the PR-treated group and other diet groups were analyzed by Dunnet's multiple comparison test (*p<0.05, **p<0.01, ***p<0.001).</p

Suggested mechanism for PR-ASG's protective action against oxidative stress in INS-1 cells.

<p>Reactive oxgen species (ROS) generated from STZ exposure are metabolized and inactivated by SOD, to produce H₂O₂. It has been shown that an imbalance in the coordinated expression/activity of glutathione peroxidase (GSHPx) and glutathione reductase (GR) can cause excessive generation of ROS, leading to oxidative stress. GSHPx converts H₂O₂ to water using glutathione (GSH). Maintenance of the redox state in cells' redox state is controlled by intracellular regulators such as reduced glutathione (GSH) and NADPH. Both GR and glucose-6-phosphate dehydrogenase (G6PD) are enzymes suspected to have protective activity against conditions of oxidative stress. On the other hand, 2-deoxy-glucose (2-DG, an inhibitor of glycolytic pathway) and 6-aminonicotinamide (6-AN, an inhibitor of pentose phosphate pathway) both decrease the cellular levels of pyruvate and NADPH. This leads to an accumulation of H₂O₂, the building of which acts to induce apoptosis. PR-ASG's defensive action is exerted via an increase in two targets: increasing homocysteine-thiolactonase (HTase) enhances homocysteine (Hcy) metabolism; and increasing pyruvate enhances protection against cell via IGF-1-related glucose metabolism.</p

GM1 complex antigens.

<div><p>(<b>A</b>) Individual value plots for selected antigens.</p> <p>(<b>B</b>) Association between sulfatide:GM1 and GM1 binding intensities in individual sera. Points on the y-axis are GM1 negative, sulfatide:GM1 positive. Sera close to the diagonal blue line (joining points of identical binding intensity for both antigens) bind both GM1 and sulfatide:GM1 complex with similar intensity. </p> <p>(<b>C</b>) Association between phosphatidylserine:GM1 and GM1 binding intensities in individual sera. Points on the y-axis are GM1 negative, PS:GM1 positive. Sera on the x-axis are GM1 positive, PS:GM1 negative. </p> <p>(<b>D</b>) Association between phosphatidylserine:GM1 and sulfatide:GM1 binding intensities in individual sera.</p></div

Heatmap of glycolipid complex binding in GBS and control sera.

<p>Clustered heatmap for the 35 antigens significantly associated with GBS (arranged on the x-axis), comparing the 181 GBS sera and 161 control sera (on the y-axis). Each row represents a single serum. The log transformed arbitrary binding intensity for each antigen is represented by the colour scale, from red (log transformed arbitrary intensity of 18, most intense) to the green background (log transformed arbitrary intensity of 0, no binding), as shown at the top of the figure. Each heatmap panel represents a separate group, as labelled. (GBS, Guillain-Barré syndrome; MS, Multiple Sclerosis; ONND, Other Non-Inflammatory Neurological Diseases; HC, Healthy Controls). Blue boxes delineate clusters of GM1 and sulfatide-complex antigens. Arrows mark sulfatide complexes which cluster separately. </p

Summary of literature search.

<p>Summary of literature search.</p