Journal: International Journal of Molecular Sciences

Article Title: Mesenchymal Stem Cells Restore Endothelial Integrity and Alleviate Emotional Impairments in a Diabetic Mouse Model via Inhibition of MMP-9 Activity

doi: 10.3390/ijms26073355

Figure Lengend Snippet: Figure 2. DM mice exhibit synaptic deficits and increased neuroinflammation in the HIP. (A) Repre- sentative WB images showing the expression levels of the presynaptic protein synapsin I (SYN1) and the postsynaptic protein postsynaptic density protein 95 (PSD95) in the HIP of CTL and DM mice. β-actin was used as an internal control. (B) Semi-quantitative analysis of PSD95 and SYN1 expression levels from immunoblot experiments. n = 6. (C) Representative images of Iba1 immunostaining in the HIP of CTL and DM mice. The area within the dashed box is magnified and displayed in the insets. Scale bars: 100 µm (main images) and 20 µm (insets). (D) Quantitative analysis of Iba1-positive microglia in the HIP of CTL and DM mice. n = 5. (E) qPCR analysis of iNOS and Arg1 mRNA levels in the HIP of CTL and DM mice. n = 6. (F) qPCR analysis of IL-6, IL-1β, and TNF-α mRNA levels in the HIP of CTL and DM mice. n = 6. (G) ELISA measuring IL-6, IL-1β, and TNF-α levels in the serum of CTL and DM mice. n = 3. Data are presented as mean ± SEM. Statistical significance was determined using a one-tailed unpaired t-test. * p < 0.05, ** p < 0.01, *** p < 0.001.

Article Snippet: The membranes were blocked and incubated overnight with the following antibodies: PSD95 (SYSN, Toronto, ON, Canada, 124002, 1:1000); SYN1 (SYSN, 106011, 1:1000); Cldn5 (Invitrogen, 35-2500, 1:1000); Ocln (Invitrogen, 71-1500, 1:1000); and MMP9 (Boster, Shanghai, China; PB9669, 1:1000).

Techniques: Expressing, Control, Western Blot, Immunostaining, Enzyme-linked Immunosorbent Assay, One-tailed Test

Journal: International Journal of Molecular Sciences

Article Title: Mesenchymal Stem Cells Restore Endothelial Integrity and Alleviate Emotional Impairments in a Diabetic Mouse Model via Inhibition of MMP-9 Activity

doi: 10.3390/ijms26073355

Figure Lengend Snippet: Figure 6. MSC treatment attenuated DM-induced neuroinflammation and synaptic deficits. (A) Representative immunofluorescence images of Iba1 staining in the HIP of mice from the CTL, MSC, DM, and DM+MSC groups. The dashed box indicates the magnified region shown in the insets. Scale bars: 100 µm (main images) and 20 µm (insets). (B) Quantitative analysis of Iba1+ microglia in the HIP. n = 4. (C) qPCR analysis of iNOS and Arg1 mRNA levels in the HIP of mice from the CTL, MSC, DM, and DM+MSC groups. n = 9. (D) qPCR analysis of mRNA levels of IL-6, IL-1β, and TNF-α in the HIP of mice from the CTL, MSC, DM, and DM+MSC groups. n = 9. (E) ELISA results showing serum levels of IL-6, IL-1β, and TNF-α in CTL, MSC, DM, and DM+MSC mice. n = 4. (F) Representative WB images of PSD95 and SYN1 expression in the HIP. β-actin was used as a loading control. (G) Semi-quantitative analysis of PSD95 and SYN1 protein levels. n = 6. Data are presented as mean ± SEM. Two-way ANOVA followed by Tukey’s post hoc test. ** p < 0.01, *** p < 0.001.

Article Snippet: The membranes were blocked and incubated overnight with the following antibodies: PSD95 (SYSN, Toronto, ON, Canada, 124002, 1:1000); SYN1 (SYSN, 106011, 1:1000); Cldn5 (Invitrogen, 35-2500, 1:1000); Ocln (Invitrogen, 71-1500, 1:1000); and MMP9 (Boster, Shanghai, China; PB9669, 1:1000).

Techniques: Immunofluorescence, Staining, Enzyme-linked Immunosorbent Assay, Expressing, Control

Journal: Chinese Journal of Lung Cancer

Article Title: 神经内分泌分化不是非小细胞肺癌高恶性度的指标

doi: 10.3779/j.issn.1009-3419.2011.08.03

Figure Lengend Snippet: 主要试剂、来源及工作浓度 Main reagents, sources and working concentrations

Article Snippet: Rabbit anti-human synaptophysin (Syn) monoclonal antibody , ZSGB-BIO , SP11 , 1:100.

Techniques:

Journal: The Journal of Biological Chemistry

Article Title: Stable α-Synuclein Oligomers Strongly Inhibit Chaperone Activity of the Hsp70 System by Weak Interactions with J-domain Co-chaperones *

doi: 10.1074/jbc.M110.127753

Figure Lengend Snippet: The initial stages in the purification of recombinant α-synuclein protofibrils. Shown is an immunoblot with an anti-α-synuclein antibody from a SDS gel (A) or a native gel (B) of cell extracts from BL21 E. coli cells expressing WT human α-synuclein. Lane 1, soluble initial crude extract. Lane 2, soluble extract after heat treatment. Lane 3, resolubilized ammonium sulfate pellet from the heat-treated soluble extract in lane 2. Numbers, estimated molecular masses from the SDS gel in kDa. Roman numbers with arrows: I, monomers; II, dimers; III, soluble oligomers; IV, large oligomers that do not enter the resolution gel.

Article Snippet: Membranes were blocked in 1% skimmed milk and incubated overnight at 4 °C with a rabbit anti-α-Syn polyclonal antibody (Stressgen, Ann Arbor, MI, catalog no. 905-565) (1/1000, v/v) and then incubated with a HRP-labeled goat anti-rabbit antibody (Bio-Rad, catalog no. 170-5046) (1/15000, v/v).

Techniques: Purification, Recombinant, Western Blot, SDS-Gel, Expressing

Journal: The Journal of Biological Chemistry

Article Title: Stable α-Synuclein Oligomers Strongly Inhibit Chaperone Activity of the Hsp70 System by Weak Interactions with J-domain Co-chaperones *

doi: 10.1074/jbc.M110.127753

Figure Lengend Snippet: Characterization of recombinant human α-synuclein (AS). A, shown is size exclusion chromatography of recombinant α-synuclein from the resolubilized amonium sulfate pellet of the heat-treated crude extract shown in lane 3 of Fig. 1, A and B. Elution profile from the Superose 6 column shows the protein concentration profile according to the absorbance at 280 nm (blue circles) and the Th-T fluorescence profile (black circles). Above the inset, Coomassie stain of the corresponding eluted fractions after separation on 12% SDS-PAGE is shown. B and C, two fractions were pooled from 15–18 ml as monomeric AS and from 7.5–11 ml as oligomeric AS, concentrated, and further separated by SDS-PAGE (B) or native-PAGE (C) and Coomassie-stained. Standard molecular masses (M) in kDa are indicated for the SDS-gel. D–F, shown are atomic force microscopy morphological studies of AS monomers and AS oligomers, as in B and C. Atomic force microscopy images (1 μm × 1 μm) of AS monomers (D) and AS oligomers (E) are shown. At higher magnitude, AS oligomers present a globular flattened-shape structure (F). G, shown is cumulative function of the diameter distribution of the AS oligomers with a mean size of 21.1 nm and a S.D. s = 2.4 nm. Inset, shown is s histogram of the distribution superimposed with a Gaussian (mean = 21.1 nm, s = 2.4 nm).

Article Snippet: Membranes were blocked in 1% skimmed milk and incubated overnight at 4 °C with a rabbit anti-α-Syn polyclonal antibody (Stressgen, Ann Arbor, MI, catalog no. 905-565) (1/1000, v/v) and then incubated with a HRP-labeled goat anti-rabbit antibody (Bio-Rad, catalog no. 170-5046) (1/15000, v/v).

Techniques: Recombinant, Size-exclusion Chromatography, Protein Concentration, Fluorescence, Staining, SDS Page, Clear Native PAGE, SDS-Gel, Microscopy

Journal: The Journal of Biological Chemistry

Article Title: Stable α-Synuclein Oligomers Strongly Inhibit Chaperone Activity of the Hsp70 System by Weak Interactions with J-domain Co-chaperones *

doi: 10.1074/jbc.M110.127753

Figure Lengend Snippet: AS oligomers are partially resistant to the Hsp70 chaperone machinery. A, AS oligomers are more resistant to the unfolding chaperones than G6PDH aggregates. Time-dependent loss of Th-T fluorescence (squares) of 1 μm heat pre-aggregated G6PDH (open squares) or 1 μm (protomers) AS oligomers (filled squares) in the presence of 5 μm DnaK, 0.75 μm DnaJ, and 1 μm GrpE (KJE) and ATP and the corresponding time-dependent reactivation of native G6PDH (filled circles) is shown. B, AS oligomers are partially dissociated by the ATP-fueled DnaK/DnaJ/GrpE chaperone system. AS oligomers were incubated as in A but for 3 h at 30 °C with or without ATP and chaperones, as specified. Samples were separated on native gel, and α-synuclein was detected by Western blot analysis as in Fig. 1.

Article Snippet: Membranes were blocked in 1% skimmed milk and incubated overnight at 4 °C with a rabbit anti-α-Syn polyclonal antibody (Stressgen, Ann Arbor, MI, catalog no. 905-565) (1/1000, v/v) and then incubated with a HRP-labeled goat anti-rabbit antibody (Bio-Rad, catalog no. 170-5046) (1/15000, v/v).

Techniques: Fluorescence, Incubation, Western Blot