Journal: Scientific Reports

Article Title: Immobilization stress-induced Escherichia coli causes anxiety by inducing NF-κB activation through gut microbiota disturbance

doi: 10.1038/s41598-018-31764-0

Figure Lengend Snippet: Immobilization stress (IS) induced anxiety-like behaviors and colitis in mice. Experimental time ( a ). Anxiety-like behaviors of mice were measured in the EPM ( b ) (time spent in open arms [OT] and open arm entries [OE]), MB (marble buried, c ) and LDT ( d , time spent in light area [TL] and number of transitions [NT]). Control mice (NC) were not treated with IS. NF-κB (p-p65 and p65), BDNF, claudin-5, and β-actin were measured in the hippocampus by immunoblotting ( e ). Brain TNF-α ( f ) IL-1β ( g ) and IL-6 levels ( h ) were analyzed by ELISA kit. Iba-1 + /CD11b + and CD11b + /CD11b + populations were assayed in CA1 and CA3 regions of the hippocampus by a confocal microscope ( i ). Blood corticosterone ( j ) TNF-α (k), IL-6 ( l ) and LPS levels ( m ) were analyzed by ELISA or LAL kit. Colitis markers colon length ( n ) and myeloperoxidase (MPO) activity ( o ) were measured in the colon. Colonic iNOS and COX-2 expression, NF-κB activation ( p ) and tight junction proteins (occludin and claudin-1), and β-actin ( q ) were analyzed by immunoblotting. Fecal LPS level ( r ) and CD11b + /CD11b + population ( s ) were analyzed by ELISA kit and confocal microscope ( i ) respectively. All data except immunoblotting data (n = 4) were expressed as mean ± SD (n = 8). # p < 0.05 vs. NC group.

Article Snippet: Antibodies for p65, p-p65, BDNF, claudin-5, occludin, caludin-1, iNOS, COX-2, and β-actin were purchased from Cell Signaling Technology (Beverly, MA).

Techniques: Control, Western Blot, Enzyme-linked Immunosorbent Assay, Microscopy, Activity Assay, Expressing, Activation Assay

Journal: Scientific Reports

Article Title: Immobilization stress-induced Escherichia coli causes anxiety by inducing NF-κB activation through gut microbiota disturbance

doi: 10.1038/s41598-018-31764-0

Figure Lengend Snippet: The fecal microbiota transplantation of IS-treated mice (FIS) induced anxiety-like behaviors and colitis in mice. FNC (fecal microbiota of control mice not treated with IS [NC]) and FIS were orally gavaged. Experimental time ( a ). Anxiety-like behaviors of mice were measured in the EPM ( b , time spent in open arms [OT] and open arm entries [OE]), MB (marble buried, c ) and LDT ( d , time spent in light area [TL] and number of transitions [NT]) tasks. NF-κB (p-p65 and p65), BDNF, claudin-5, and β-actin were measured in the hippocampus by immunoblotting ( e ). Brain TNF-α ( f ) IL-1β ( g ) and IL-6 levels ( h ) were analyzed by ELISA kit. Blood corticosterone ( f ) IL-6 ( g ) and LPS levels ( h ) were analyzed by ELISA or LAL kit. Iba-1 + /CD11b + and CD11b + /CD11b + populations were assayed by a confocal microscope ( i ). Blood corticosterone ( j ) TNF-α ( k ) IL-6 ( l ) and LPS levels ( m ) were analyzed by ELISA or LAL kit. Colitis markers colon length ( n ) and myeloperoxidase (MPO) activity ( o ) were measured in the colon. Colonic iNOS, COX-2, NF-κB ( p ) and tight junction proteins (occludin and claudin-1), and β-actin ( q ) were analyzed by immunoblotting. Fecal LPS level ( r ) and CD11b + /CD11b + population ( s ) were analyzed by ELISA and confocal microscopy ( i ) respectively. FNC and FIS in figures indicate groups treated with fecal microbiota of control and IS-treated mice (suspended in 1% dextrose), respectively. All data except immunoblotting data (n = 4) were expressed as mean ± SD (n = 8). # p < 0.05 vs. FNC group.

Article Snippet: Antibodies for p65, p-p65, BDNF, claudin-5, occludin, caludin-1, iNOS, COX-2, and β-actin were purchased from Cell Signaling Technology (Beverly, MA).

Techniques: Transplantation Assay, Control, Western Blot, Enzyme-linked Immunosorbent Assay, Microscopy, Activity Assay, Confocal Microscopy

Journal: Scientific Reports

Article Title: Immobilization stress-induced Escherichia coli causes anxiety by inducing NF-κB activation through gut microbiota disturbance

doi: 10.1038/s41598-018-31764-0

Figure Lengend Snippet: Escherichia coli caused anxiety and colitis in mice. E . coli (1 × 10 9 CFU/0.2 mL/mouse) was orally gavaged. Experimental time ( a ). Anxiety-like behaviors of mice were measured in the EPM ( b , time spent in open arms [OT] and open arm entries [OE]), MB (marble buried, c ) and LDT ( d , time spent in light area [TL] and number of transitions [NT]) tasks. NF-κB (p-p65 and p65), BDNF, claudin-5, and β-actin were measured in the hippocampus by immunoblotting ( e ). Brain TNF-α ( f ) IL-1β ( g ) and IL-6 levels ( h ) were analyzed by ELISA kit. Blood corticosterone ( f ) IL-6 ( g ) and LPS levels ( h ) were analyzed by ELISA or LAL kit. Iba-1 + /CD11b + and CD11b + /CD11b + populations were assayed by a confocal microscope ( i ). Blood corticosterone ( j ) TNF-α ( k ) IL-6 ( l ) and LPS levels ( m ) were analyzed by ELISA or LAL kit. Colitis markers colon length ( n ) and myeloperoxidase (MPO) activity ( o ) were measured in the colon. Colonic iNOS, COX-2, NF-κB ( p ) and tight junction proteins (occludin and claudin-1), and β-actin ( q ) were analyzed by immunoblotting. Fecal LPS level ( r ) and CD11b + /CD11b + population ( s ) were analyzed by ELISA and confocal microscopy ( i ) respectively. NC and EC in figures indicate groups treated with vehicle alone (1% dextrose) and E . coli , respectively. All data except immunoblotting data (n = 4) were expressed as mean ± SD (n = 8). # p < 0.05 vs. NC group.

Article Snippet: Antibodies for p65, p-p65, BDNF, claudin-5, occludin, caludin-1, iNOS, COX-2, and β-actin were purchased from Cell Signaling Technology (Beverly, MA).

Techniques: Western Blot, Enzyme-linked Immunosorbent Assay, Microscopy, Activity Assay, Confocal Microscopy

Journal: Scientific Reports

Article Title: Immobilization stress-induced Escherichia coli causes anxiety by inducing NF-κB activation through gut microbiota disturbance

doi: 10.1038/s41598-018-31764-0

Figure Lengend Snippet: LPS purified from Escherichia coli (EL) caused anxiety and colitis in mice. Experimental schedule ( a ). Anxiety-like behaviors were measured on the 10 th day after the initial treatment with LPS in the EPM ( b ) MB ( d ) and LDT ( d ) tasks. LPS (10 μg/kg) was intraperitoneally treated for 5 days. NF-κB activation and BDNF and claudin-5 expression were measured in the hippocampus ( e ). Brain TNF-α ( f ) IL-1β ( g ) and IL-6 ( h ) levels were analyzed by ELISA kits. Blool corticosterone ( i ) IL-6 ( f ) TNF-α ( j ) IL-1β ( k ) IL-6 ( l ) and LPS ( m ) levels were by ELISA kits. Colitis markers body weight gain ( n ) macroscopic score ( o ) colon shortening ( p ) and MPO activity ( q ) IL-6 ( r ) TNF-α ( s ) IL-1β ( t ) and IL-10 ( u ) expression were measured in the colon. NC and EL in figures indicate groups treated with vehicle alone (1% dextrose) and E . coli LPS, respectively. All data except immunoblotting data (n = 4) were expressed as mean ± SD (n = 8). # p < 0.05 vs. NC group.

Article Snippet: Antibodies for p65, p-p65, BDNF, claudin-5, occludin, caludin-1, iNOS, COX-2, and β-actin were purchased from Cell Signaling Technology (Beverly, MA).

Techniques: Purification, Activation Assay, Expressing, Enzyme-linked Immunosorbent Assay, Activity Assay, Western Blot

Journal: Scientific Reports

Article Title: Immobilization stress-induced Escherichia coli causes anxiety by inducing NF-κB activation through gut microbiota disturbance

doi: 10.1038/s41598-018-31764-0

Figure Lengend Snippet: Oral administration of Lactobacillus johnsonii alleviated immobilization stress (IS)-induced anxiety-like behaviors and colitis in mice. Experimental schedule ( a ). Anxiety-like behaviors were measured on the 10 th day after the initial treatment with FIS in the EPM ( b ) MB ( c ) and LDT ( d ) tasks. IS was treated for 10 days. NF-κB activation and BDNF and claudin-5 expression were measured in the hippocampus ( e ). Brain TNF-α ( f ) IL-1β ( g ) and IL-6 ( h ) levels were analyzed by ELISA kits. Iba-1 + /CD11b + and CD11b + /CD11b + populations were assayed in CA1 and CA3 regions of the hippocampus by a confocal microscope ( i ). Blood corticosterone ( j ) TNF-α ( k ) IL-1β ( l ) IL-6 ( m ) and LPS ( n ) levels in the blood. Colitis markers body weight gain ( o ) macroscopic score ( p ) colon shortening ( q ) and MPO activity ( r ) TNF-α ( s ) IL-1β ( t ) IL-6 ( u ) IL-10 ( v ), and NF-κB activation, iNOS, and COX-2 expression ( w ) were measured in the colon. Fecal LPS level ( x ) and gut microbiota ( y ) were measured in the feces. CD11b + /CD11b + populations were assayed in the colon by a confocal microscope ( z ). NC, IS, and IJ in figures indicate groups treated with vehicle alone (1% dextrose) in normal control mice, vehicle in IS-treated mice, and L . johnsonii in IS-treated mice, respectively. The bacterial abundance was indicated as % of NC. All data except immunoblotting data (n = 4) were expressed as mean ± SD (n = 8). # p < 0.05 vs. NC group. * p < 0.05 vs. IS-treated control (IS) group.

Article Snippet: Antibodies for p65, p-p65, BDNF, claudin-5, occludin, caludin-1, iNOS, COX-2, and β-actin were purchased from Cell Signaling Technology (Beverly, MA).

Techniques: Activation Assay, Expressing, Enzyme-linked Immunosorbent Assay, Microscopy, Activity Assay, Control, Western Blot

Journal: Scientific Reports

Article Title: Immobilization stress-induced Escherichia coli causes anxiety by inducing NF-κB activation through gut microbiota disturbance

doi: 10.1038/s41598-018-31764-0

Figure Lengend Snippet: Oral administration of Lactobacillus johnsonii alleviated Escherichia coli -induced anxiety-like behaviors and colitis in mice. Experimental schedule ( a ). Anxiety-like behaviors were measured on the 10 th day after the initial treatment with FIS in the EPM ( b ) MB ( c ) and LDT ( d ) tasks. IS was treated for 10 days. NF-κB activation and BDNF and claudin-5 expression were measured in the hippocampus ( e ). Brain TNF-α ( f ) IL-1β ( g ) and IL-6 ( h ) levels were analyzed by ELISA kits. Iba-1 + /CD11b + and CD11b + /CD11b + populations were assayed in CA1 and CA3 regions of the hippocampus by a confocal microscope ( i ). Blood corticosterone ( j ) TNF-α ( k ) IL-1β ( l ) IL-6 ( m ) and LPS ( n ) levels in the blood. Colitis markers body weight gain ( o ) macroscopic score ( p ) colon shortening ( q ) and MPO activity ( r ) TNF-α ( s ) IL-1β ( t ) IL-6 ( u ) IL-10 ( v ) and NF-κB activation, iNOS, and COX-2 expression ( w ) were measured in the colon. Fecal LPS level ( x ) and gut microbiota ( y ) were measured in the feces. CD11b + /CD11b + populations were assayed in the colon by a confocal microscope ( z ). NC, EC, and EJ in figures indicate groups treated with vehicle alone (1% dextrose) in control mice, vehicle in E . coli -treated mice, and L . johnsonii in E . coli -treated mice, respectively. The bacterial abundance was indicated as % of NC. All data except immunoblotting data (n = 4) were expressed as mean ± SD (n = 8). # p < 0.05 vs. NC group. * p < 0.05 vs. EC-treated control (EC) group.

Article Snippet: Antibodies for p65, p-p65, BDNF, claudin-5, occludin, caludin-1, iNOS, COX-2, and β-actin were purchased from Cell Signaling Technology (Beverly, MA).

Techniques: Activation Assay, Expressing, Enzyme-linked Immunosorbent Assay, Microscopy, Activity Assay, Control, Western Blot

Journal: Stroke

Article Title: Electrical Stimulation of the Cerebral Cortex Exerts Antiapoptotic, Angiogenic, and Anti-Inflammatory Effects in Ischemic Stroke Rats Through Phosphoinositide 3-Kinase/Akt Signaling Pathway

doi: 10.1161/strokeaha.109.563627

Figure Lengend Snippet: Figure 4. Upregulation of neurotrophic factors in the stroke rats receiving electric stimulation. The schematic diagram shows the regions in the cortex and striatum of rats with electric stimulation (A). Black box, cortex; white box, striatum. Western blots of BDNF, GDNF, and VEGF with -actin demonstrated the upregulation of all the factors both in the cortex and striatum of rats with electric stim- ulation compared with the control group (B). Quantification shown in C. Data represent mean percentagesSE. *P0.05 versus cortex of the control rats. **P0.05 versus striatum of the control rats.

Article Snippet: The membranes were probed at room temperature for 12 hours with mouse antivascular endothelial growth factor (VEGF) antibody (1:1000, #555036; BD Pharmingen), rabbit anti-glial cell line-derived neurotrophic factor (GDNF) antibody (1:500, #sc-328; Santa Cruz, CA), rabbit anti-brain-derived neurotrophic factor (BDNF) antibody (1:500, #546; Santa Cruz), rabbit antiphosphorylated Akt antibody (1:500, #9271; Cell Signaling), rabbit anti-Akt antibody (1:1000, #9272; Cell Signaling), and antimouse anti- -actin (1:2000, A5441; Sigma) as primary antibodies followed by exposure to peroxidaseconjugated secondary antibodies (1:2000, antimouse IgG; 1:2000, antirabbit IgG; Amersham Pharmacia Biotech; 1:2000) for 1 hour.

Techniques: Western Blot, Control

Journal: Frontiers in Aging Neuroscience

Article Title: A Multi-Ingredient Nutritional Supplement in Combination With Resistance Exercise and High-Intensity Interval Training Improves Cognitive Function and Increases N -3 Index in Healthy Older Men: A Randomized Controlled Trial

doi: 10.3389/fnagi.2019.00107

Figure Lengend Snippet: Experimental design. Participants consumed either an experimental supplement (SUPP) or control (CON) beverage twice per day for 20 weeks (weeks 0–19, inclusive). Between weeks 7–18 (inclusive), participants completed a 12-week exercise training program. Exercise training consisted of RET twice weekly (Mondays and Fridays) and HIIT once per week (Wednesdays). At weeks –1 (baseline), 6, and 19 we assessed cognitive function and obtained a blood sample for the measurement of BDNF, 25(OH)D, and erythrocyte plasma membrane phospholipid composition. Phase 1: SUPP/CON took place between weeks 0–6, and Phase 2: SUPP/CON + EX took place between weeks 7–19. SUPP, supplement; CON, control; RET, resistance exercise training; HIIT, high-intensity interval training; BDNF, brain-derived neurotrophic factor; 25(OH)D, 25-hydroxyvitamin D.

Article Snippet: Fasting plasma 25(OH)D concentrations were measured by radioimmunoassay (DiaSorin Canada Inc.; Mississauga, ON, Canada), and fasting plasma BDNF concentrations were measured using a Quantikine Human Free BDNF ELISA kit (R&D Systems, Inc.; Minneapolis, MN, United States).

Techniques: Control, Clinical Proteomics, Membrane, Derivative Assay

Journal: Frontiers in Aging Neuroscience

Article Title: A Multi-Ingredient Nutritional Supplement in Combination With Resistance Exercise and High-Intensity Interval Training Improves Cognitive Function and Increases N -3 Index in Healthy Older Men: A Randomized Controlled Trial

doi: 10.3389/fnagi.2019.00107

Figure Lengend Snippet: Blood analyses and nutrient bioavailability.

Article Snippet: Fasting plasma 25(OH)D concentrations were measured by radioimmunoassay (DiaSorin Canada Inc.; Mississauga, ON, Canada), and fasting plasma BDNF concentrations were measured using a Quantikine Human Free BDNF ELISA kit (R&D Systems, Inc.; Minneapolis, MN, United States).

Techniques: Clinical Proteomics, Membrane

Journal: International Journal of Molecular Sciences

Article Title: Fingolimod Modulates Dendritic Architecture in a BDNF-Dependent Manner

doi: 10.3390/ijms21093079

Figure Lengend Snippet: Fingolimod-phosphate (FTY720-P) regulates neuronal architecture in a BDNF-dependent manner. ( A ) Representative Neurolucida tracings, used to perform the Sholl analysis of dendritic complexity for feGFP expressing neurons treated for 24h with: DMSO (black), 2nM FTY720-P (gray), DMSO + TrkBFc (dark blue) and 2nM FTY720-P + TrkBFc (light blue). Scale bar: 100 μm. ( B ) Sholl analysis displayed as number of dendritic intersections against distance from the cell body for DMSO (black), 2nM FTY720-P (gray), DMSO + TrkBFc (dark blue) and 2nM FTY720-P + TrkBFc (light blue) and ( C ) total dendritic complexity of all treatment groups. F value in ( B ) refers to comparison between all the 4 treatment groups. ( D ) Dendritic spine densities for DMSO (black), 2nM FTY720-P (gray), DMSO + TrkBFc (dark blue) and 2nM FTY720-P + TrkBFc (light blue) calculated using segments of secondary dendritic branches as shown in ( E ) feGFP panel. The Syn panel displays the corresponding staining of the pre-synaptic marker SynapsinI/II and the merge panel shows the images with overlapping SynapsinI/II puncta (red, pre-synapse) to its matching feGFP dendrite segment (green, post-synapse). The arrows point to coinciding puncta, indicative of mature synapse between the post and pre-synaptic compartments. Scale bar: 5μm. ( F ) The graph compares the fraction of SynapsinI/II positive feGFP labelled spines for DMSO (black), 2nM FTY720-P (gray), DMSO + TrkBFc (dark blue) and 2nM FTY720-P + TrkBFc (light blue). ( G ) Representative fields of view (FOV) of the DMSO, 2nM FTY720-P, DMSO + TrkBFc and 2nM FTY720-P + TrkBFc treated hippocampal cultures stained for c-fo s . Scale bar: 100μm. ( H ) Quantification of the proportion of c-Fos expressing neurons represented as normalized fraction for DMSO (black), 2nM FTY720-P (gray), DMSO + TrkBFc (dark blue) and 2nM FTY720-P + TrkBFc (light blue) treated cultures. All graphs represent data as mean + SEM. Numbers in the bars show either total number of neurons or of FOV analyzed, obtained from ≥3 sets of independent experiments. Two-way ANOVA followed by Bonferroni post-hoc test was used in ( B ). For ( C , D , F ) and ( H ) one-way ANOVA with Bonferroni post-hoc was used. Denotations for significance are * p < 0.05, ** p < 0.01, **** p < 0.0001.

Article Snippet: Recombinant Human BDNF protein (R&D systems, Minneapolis, MN, USA) was dissolved in sterile PBS with 0.1% BSA at a concentration of 50 ng/μL.

Techniques: Expressing, Comparison, Staining, Marker

Journal: International Journal of Molecular Sciences

Article Title: Fingolimod Modulates Dendritic Architecture in a BDNF-Dependent Manner

doi: 10.3390/ijms21093079

Figure Lengend Snippet: Treatment with the non-phosphorylated Fingolimod (FTY720) modulates neuronal architecture. ( A ) | Representative Neurolucida tracings from feGFP positive hippocampal neurons used for the Sholl analysis from cultures treated either with DMSO or 10nM FTY20 for 24 h. Scale bar: 100μm. ( B ) | Dendritic complexity shown by the number of dendritic intersections plotted against the distance from the soma for DMSO (black) and FTY720 (blue) treated neurons. The F value shows the statistical comparison between the two groups. The inset graph represents total dendritic complexity upon treatment with DMSO (black) and FTY720 (blue). ( C ) | Total dendritic length for DMSO (black) and FTY720 (blue) treated neurons. ( D ) | Representative stretches from dendrites of eGFP transfected hippocampal neurons showing dendritic spine protrusions, treated either with DMSO or FTY720 for 24h. Scale bar: 5μm. ( E ) | The graph shows dendritic spine density for DMSO (black) and FTY720 (blue) treated neurons . ( F ) | Representative images of fields of view (FOV) from primary hippocampal cultures stained with anti phospho-ERK1/2 antibody, 30 min post-application of one of the following: DMSO, 10nM FTY720, DMSO_100, 100nM FTY720, DMSO_100 + TrkB-Fc, 100nM FTY720 + TrkB-Fc or 40ng recombinant BDNF protein as a positive control. Scale bar: 100μm. ( G ) | The graph displays the fraction of pERK1/2 expressing neurons relative to the total number of MAP2 + neurons. The data is normalized to the respective controls and compared between the different treatment groups: DMSO (black), 10nM FTY720 (light blue solid), DMSO100 (gray), 100nM FTY720 (dark blue solid), 40ng recombinant BDNF (magenta), DMSO100 + TrkB-Fc (gray open), 100nM FTY720 + TrkB-Fc (dark blue open). All data is plotted as mean + SEM. Numbers in the bars show total number of neurons or FOV analyzed, obtained from ≥3 sets of independent experiments. Two-way ANOVA followed by Bonferroni post-hoc test was used in ( B ). For ( B ) total intersections, ( C ) and ( E ) unpaired Student’s t-test and for ( G ) one-way ANOVA with Bonferroni post-hoc was used. Denotations for significance are * p < 0.05, *** p < 0.001, **** p < 0.0001.

Article Snippet: Recombinant Human BDNF protein (R&D systems, Minneapolis, MN, USA) was dissolved in sterile PBS with 0.1% BSA at a concentration of 50 ng/μL.

Techniques: Comparison, Transfection, Staining, Recombinant, Positive Control, Expressing