Journal: Microbiome

Article Title: Engineering a dysbiotic biofilm model for testing root caries interventions through microbial modulation

doi: 10.1186/s40168-024-01862-5

Figure Lengend Snippet: Collagenase and gelatinase activity of the biofilms with and without pre-treatments or post-treatments with the anacardic acid LDT11 and the cranberry extract. A Pre-treatment gelatinase and collagenase activity (EnzChek Gelatinase/Collagenase assay kit using the DQ collagen type I and DQ gelatine as substrates). Statistical analysis: 2-way ANOVA, Dunn’s multiple comparisons test. B Pre-treatment strategy: gelatine hydrolysis, assessed at two time points (48 h and 7 days). The bar graph indicates the number of replicates that were positive for hydrolysis. The image below the graph displays two tubes containing gelatine: the top tube shows positive hydrolysis with the gelatine in a liquid state, while the bottom tube shows negative hydrolysis with the gelatine remaining solid. C Post-treatment gelatinase activity (EnzChek Gelatinase/Collagenase assay kit using the DQ gelatine as substrate). Fluorescence was measured at three time points 2 h, 7 h and 24 h. Statistical analysis: 2-way ANOVA, Dunnett’s multiple comparisons test

Article Snippet: To determine collagenase/gelatinase activity, the dysbiotic biofilms from both strategies were assayed in the EnzChek® Gelatinase/Collagenase Assay Kit using the DQ collagen type I (from bovine skin, fluorescein conjugate) and DQ gelatine as substrates (Molecular Probes, Inc.), following the supplier protocol.

Techniques: Activity Assay, Fluorescence

Journal: Neuron

Article Title: Loss of adaptive myelination contributes to methotrexate chemotherapy-related cognitive impairment

doi: 10.1016/j.neuron.2019.04.032

Figure Lengend Snippet: A) Representative image demonstrating RNAscope visualization of frontal cortex deep layer neurons (NeuN, white), astrocytes (Glast, green), and Bdnf mRNA (red). DAPI, blue. Scale bar = 20 μm

Article Snippet: RNAscope was performed according to Fluorescent Multiplex Reagent Kit protocol using RNAscope probes against BDNF (Advanced Cell Diagnostics, 424821), RbFox3 (NeuN; Advanced Cell Diagnostics 313311-C2) and Slc1a3 (Glast; Advanced Cell Diagnostics, 320851-C3).

Techniques:

Journal: Science Advances

Article Title: IL4-driven microglia modulate stress resilience through BDNF-dependent neurogenesis

doi: 10.1126/sciadv.abb9888

Figure Lengend Snippet: ( A ) Behavioral screening of LS and HS subpopulations of mice exposed to 3-week CMS. ( B ) Quantitative PCR to examine mRNA expression of cytokines, chemokines, growth factors, and trophic factors in the hippocampus and prefrontal cortex of HS and LS mice. Data are standardized to control group ( n = 4 to 6, each sample in triplicate). ( C ) Enzyme-linked immunosorbent assay to assay the protein level of IL4 in prefrontal cortex, cerebellum, hippocampus, amygdala, and olfactory bulb of control, HS, and LS mice. Each circle represents one mouse ( n = 6, each sample in triplicate). ( D ) Western blotting shows the levels of IL4, IL4 receptor α chain (IL4Rα), STAT6, and pSTAT6 in the hippocampus of control, HS, and LS mice. IL4 and IL4Rα are normalized to β-actin, and the pSTAT6 is normalized to STAT6. ( n = 4 to 6, each sample in triplicate). ( E ) Correlation between sucrose preference or immobility duration in TST, hippocampal Arg1 levels, and hippocampal IL4 levels in CMS-exposed mice. Each circle represents one mouse ( n = 6). ( F ) Morphological characters of hippocampal microglia in control, HS, and LS mice. Scale bars, 50 μm (top) and 10 μm (bottom). The histogram represents the quantification of number and branch length of Iba1 + cells in hippocampus. Each dot in the bar graph represents the average of all micrograph (five to six micrographs for each mouse) in each mouse ( n = 5). ( G ) Immunofluorescence micrographs and quantification of Arg1 + microglia in hippocampus of control, LS, and HS mice ( n = 5). Scale bars, 50 μm. Data are presented as means ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005, one-way ANOVA with Tukey test (B, C, D, F, and G). DAPI, 4′,6-diamidino-2-phenylindole.

Article Snippet: In situ hybridization was performed according to the protocol of the Bdnf -RNAscope Multiplex Fluorescent Probe Reagent Kit v2 and Mm- Il4 -RNAscope 2.5 VS Fluorescent Probe Reagent Kit (Advanced Cell Diagnostics, California, USA).

Techniques: Real-time Polymerase Chain Reaction, Expressing, Control, Enzyme-linked Immunosorbent Assay, Western Blot, Immunofluorescence

Journal: Science Advances

Article Title: IL4-driven microglia modulate stress resilience through BDNF-dependent neurogenesis

doi: 10.1126/sciadv.abb9888

Figure Lengend Snippet: ( A ) Experimental timeline of IL4 overexpression in the hippocampus and CMS protocol. ( B ) Volcano maps indicate differentially expressed genes (DEGs) in the brains of CMS-exposed and/or AAV-IL4–treated mice. ( C ) Quantitative PCR examination of the expression of several DEGs in the hippocampus of mice ( n = 4 to 6). ( D ) Western blotting shows the levels of IL4, Arg1, and iNOS in the hippocampus of CMS-exposed and/or AAV-IL4–treated mice ( n = 5 to 6). ( E ) Morphological characters of Arg1 + microglia in hippocampus. Scale bar, 100 μm. The histogram represents the quantification of number and branch length of Arg1 + -Iba1 + cells in hippocampus. Each dot in the bar graph represents the average of five to six micrographs in each mouse ( n = 6). ( F ) Flow cytometry of single-cell suspension of the whole hippocampus for microglia (CD45 int -CD11b + cells), activated microglia (CD86 + cells), and anti-inflammatory microglia (CD86 + -CD206 + cells). The histogram represents the quantification of CD45 int -CD11b + cells, CD86 + cells, and CD86 + -CD206 + cells ( n = 5 to 6). Data are presented as means ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005 versus AAV + Ctrl group, # P < 0.05, ## P < 0.01, ### P < 0.005 versus AAV + CMS group, two-way ANOVA with Tukey test (C); * P < 0.05, ** P < 0.01, *** P < 0.005, two-way ANOVA with Tukey test (D, E, and F). FITC A, fluorescein isothiocyanate area (FITC-A).

Article Snippet: In situ hybridization was performed according to the protocol of the Bdnf -RNAscope Multiplex Fluorescent Probe Reagent Kit v2 and Mm- Il4 -RNAscope 2.5 VS Fluorescent Probe Reagent Kit (Advanced Cell Diagnostics, California, USA).

Techniques: Over Expression, Real-time Polymerase Chain Reaction, Expressing, Western Blot, Flow Cytometry, Suspension

Journal: Science Advances

Article Title: IL4-driven microglia modulate stress resilience through BDNF-dependent neurogenesis

doi: 10.1126/sciadv.abb9888

Figure Lengend Snippet: ( A ) Immunofluorescence micrographs and quantification show the hippocampal neurogenesis of LS and HS mice ( n = 5). ( B ) GO enrichment analysis for DEGs in the hippocampus of CMS-exposed mice. ER, endoplasmic reticulum; FDR, false discovery rate. ( C ) Effects of IL4 overexpression and/or microglial IL4Rα down-regulation on NSPC proliferation and differentiation in the hippocampus of CX 3 CR1 Cre/ERT2 mice. Representative fluorescence micrographs showing DCX expression and BrdU incorporation in the subgranular zone (SGZ). Scale bar, 100 μm. Quantification of total number of BrdU + cells, total number of DCX + -BrdU + cells, and percentage of DCX + -BrdU + cells out of all BrdU + cells in the neurogenic zones ( n = 5). ( D ) Effects of IL4 overexpression and/or microglial IL4Rα down-regulation in the hippocampus (before IL4 overexpression) on NSPC survival and maturation. Representative fluorescence micrographs illustrating NeuN expression and BrdU incorporation in the SGZ. Scale bar, 100 μm. Quantification of total number of BrdU + cells, BrdU + -NeuN + cells, and DCX + -NeuN + cells in DG ( n = 5). ( E ) Effects of conditioned medium from microglia isolated from hippocampus on NSPC proliferation. Proliferation was monitored by quantifying the percentage of BrdU + cells ( n = 6). Scale bar, 50 μm. Representative micrographs of NSPCs cultured for 7 days in conditioned culture medium from microglia isolated from hippocampus. Cells were immunolabeled with glial fibrillary acidic protein (GFAP) to identify astrocytes and MAP2 to identify neurons. Percentages of GFAP + cells and MAP2 + cells are quantified ( n = 6). Scale bar, 30 μm. Data are presented as means ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005, one-way ANOVA with Tukey test (A) and two-way ANOVA with Tukey test (C, D, and E).

Article Snippet: In situ hybridization was performed according to the protocol of the Bdnf -RNAscope Multiplex Fluorescent Probe Reagent Kit v2 and Mm- Il4 -RNAscope 2.5 VS Fluorescent Probe Reagent Kit (Advanced Cell Diagnostics, California, USA).

Techniques: Immunofluorescence, Over Expression, Fluorescence, Expressing, BrdU Incorporation Assay, Isolation, Cell Culture, Immunolabeling

Journal: Science Advances

Article Title: IL4-driven microglia modulate stress resilience through BDNF-dependent neurogenesis

doi: 10.1126/sciadv.abb9888

Figure Lengend Snippet: ( A ) Western blotting shows the levels of BDNF and TrkB in the hippocampus of LS and HS mice. BDNF and TrkB are normalized to β-actin ( n = 3). ( B ) Western blotting shows the levels of BDNF and TrkB in the hippocampus of CMS and/or AAV-IL4–treated mice. BDNF and TrkB are normalized to β-actin ( n = 4). ( C ) BDNF was detected in hippocampal microglia (Iba1 + cells) using Bdnf -RNAscope. ( D ) The mRNA expression of BDNF was observed in hippocampal microglia of AAV-IL4–treated mice ( n = 4). ( E ) Double immunohistochemical staining of BDNF and Arg1 in the hippocampus of AAV-IL4 mice. Scale bar, 20 μm. ( F ) Effects of CMS, knockdown of microglial IL4Rα, and overexpression of IL4 on BDNF protein levels in the hippocampus ( n = 5). ( G ) Effects of k252a treatment to block the BDNF/TrkB pathway on hippocampal neurogenesis in AAV-IL4 mice under CMS exposure. Representative fluorescence micrographs showing DCX expression and BrdU incorporation in the SGZ. Scale bar, 100 μm. Quantification of the number of DCX + cells and total number of DCX + -BrdU + cells in the neurogenic zones ( n = 5). DMSO, dimethyl sulfoxide. ( H ) The expression and secretion of BDNF in primary microglia after IL4 treatment in vitro ( n = 6). ( I ) Effects of anti-BDNF antibody or K252a on NSPC proliferation (BrdU + cells) in the presence of the conditioned culture medium from IL4-treated microglia ( n = 5). Scale bar, 20 μm. Data are presented as means ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005, one-way ANOVA with Tukey test (A, G, and I), two-way ANOVA with Tukey test (B, D, and F), and two-tailed t test (H).

Article Snippet: In situ hybridization was performed according to the protocol of the Bdnf -RNAscope Multiplex Fluorescent Probe Reagent Kit v2 and Mm- Il4 -RNAscope 2.5 VS Fluorescent Probe Reagent Kit (Advanced Cell Diagnostics, California, USA).

Techniques: Western Blot, RNAscope, Expressing, Immunohistochemical staining, Staining, Knockdown, Over Expression, Blocking Assay, Fluorescence, BrdU Incorporation Assay, In Vitro, Two Tailed Test

Journal: Science Advances

Article Title: IL4-driven microglia modulate stress resilience through BDNF-dependent neurogenesis

doi: 10.1126/sciadv.abb9888

Figure Lengend Snippet: ( A ) Effects of overexpression of IL4 in the hippocampus on stress-induced depressive-like behaviors. Mice were injected stereotactically with AAV-IL4 or AAV, allowed to recover for 2 weeks, and then subjected to a CMS protocol consisting of exposure to three random stressors daily for 4 weeks. The mice were assessed by SPT and FST ( n = 8). ( B ) Correlations between sucrose preference and percentage of Arg1 + microglia in hippocampus, immobility time in FST and percentage of Arg1 + microglia in hippocampus, sucrose preference and concentration of BDNF in hippocampus, immobility time in FST and concentration of BDNF in hippocampus, sucrose preference and number of BrdU + -DCX + cells in DG, and immobility time in FST and number of BrdU + -DCX + cells in DG. Each circle represents one mouse ( n = 6). ( C ) Effects of microglial IL4Rα down-regulation (before IL4 overexpression) on stress-induced depressive-like behaviors, as assessed by SPT and FST ( n = 8 to 10). ( D ) Assessment of stress-induced depressive-like behaviors in WT mice when treated with K252a or TMZ with sCMS exposure ( n = 8). ( E ) Assessment of stress-induced depressive-like behaviors in AAV-IL4 mice when treated with k252a or TMZ ( n = 8 to 10). Data are presented as means ± SEM. * P < 0.05, ** P < 0.01, *** P < 0.005, two-way ANOVA with Bonferroni test (A and C) and one-way ANOVA with Bonferroni test (D and E).

Article Snippet: In situ hybridization was performed according to the protocol of the Bdnf -RNAscope Multiplex Fluorescent Probe Reagent Kit v2 and Mm- Il4 -RNAscope 2.5 VS Fluorescent Probe Reagent Kit (Advanced Cell Diagnostics, California, USA).

Techniques: Over Expression, Injection, Concentration Assay

Journal: Cancer investigation

Article Title: In situ protein expression of RRM1, ERCC1 and BRCA1 in metastatic breast cancer patients treated with gemcitabine-based chemotherapy

doi: 10.3109/07357900903095722

Figure Lengend Snippet: Patients characteristics

Article Snippet: Patients with HER2 positive tumors [3+ by immunohistochemistry (IHC) on the HercepTest (Dako A/S) or 2+ by IHC and dual color fluorescence in situ hybridization (FISH; PathVision HER2 DNA Probe Kit, Vysis, Inc.) positivity] were also eligible, provided they had progressed after at least one trastuzumab-based therapy for metastatic disease.

Techniques: