Journal: The Journal of Biological Chemistry

Article Title: In vivo localization of human acetylcholinesterase-derived species in a β-sheet conformation at the core of senile plaques in Alzheimer's disease

doi: 10.1074/jbc.RA118.006230

Figure Lengend Snippet: Reactivity of the antibodies in brains of age-matched control and hAChE-S single transgenic mice. A, absence of reactivity of ThS and the antibodies against Aβ, hAChE-S, and AChE peptides in the brain of age-matched control mice. B, in the brain of hAChE-S single transgenic mice, the only reactivity detected is that of the antibodies recognizing globular hAChE-S. Frozen brain sections, from one animal (12 μm), were labeled with the Elite ABC M.O.M. kit or Vectastain Elite ABC rabbit IgG kit using a biotinylated anti-mouse or anti-rabbit secondary antibody and FITC-conjugated avidin (Vector Laboratories). The scale bar represents 10 μm. Shown is one of the z slices from a z stack.

Article Snippet: ABC mouse-on-mouse (M.O.M.) or Vectastain Elite ABC rabbit IgG kits were used, following the manufacturer's instructions (Vector Laboratories, Burlingame, CA).

Techniques: Transgenic Assay, Labeling, Avidin-Biotin Assay, Plasmid Preparation

Journal: The Journal of Biological Chemistry

Article Title: In vivo localization of human acetylcholinesterase-derived species in a β-sheet conformation at the core of senile plaques in Alzheimer's disease

doi: 10.1074/jbc.RA118.006230

Figure Lengend Snippet: Localization and recognition of mature plaques in the brain of hAPPswe single transgenic (A, left column) and hAChE-S/hAPPswe double transgenic mice (A, right column, and B). A, frozen brain sections (12 μm) from single and double transgenic mice were single-labeled with the Elite ABC M.O.M. kit or Vectastain Elite ABC rabbit IgG kit using a biotinylated anti-mouse or anti-rabbit secondary antibody, followed by avidin-conjugated peroxidase and then the peroxidase substrate (DAB, Vector Laboratories). The scale bar represents 200 μm (in the top left brain section). Black arrowheads indicate examples of mature plaques labeled with both anti-Aβ and anti-intact or anti-domains of hAChE-S, and the blue arrowheads indicate examples of mature plaques labeled only with anti-intact or anti-domains of hAChE-S. The insets show z projections of the same mature plaque (identified by a black circle in the full brain section) labeled with each of the following antibodies: HR2, Bam10, and 105A, and the scale bar represents 50 μm. One representative whole brain section per condition was collected as a montage of automatically tiled images. B, frozen brain sections (12 μm) from double hAChE-S/hAPPswe transgenic mice were single-labeled with the Elite ABC M.O.M. kit or Vectastain Elite ABC rabbit IgG kit using a biotinylated anti-mouse or anti-rabbit secondary antibody and FITC-conjugated avidin (Vector Laboratories). Shown are fluorescent labeling of whole brain sections (left, one representative whole brain section per condition was collected as a montage of automatically tiled images) and z projections of mature plaques (fluorescent labeling and merged phase, right). C, mouse brain map at the rostral level, indicating the main brain regions with their sub-areas. D, shown are quantitations of the area occupied overall by the fluorescent labeling from a specific antibody/reagent in the whole brain section of hAPPswe/hAChE double transgenic (i.e. the whole brain section shown in B) (top graph), the overall intensity of the fluorescent labeling in the whole brain section of hAPPswe/hAChE double transgenic (middle graph), and the fold ratio when compared with ThS of each fluorescent labeling in term of area occupied and intensity (bottom graph).

Article Snippet: ABC mouse-on-mouse (M.O.M.) or Vectastain Elite ABC rabbit IgG kits were used, following the manufacturer's instructions (Vector Laboratories, Burlingame, CA).

Techniques: Transgenic Assay, Labeling, Avidin-Biotin Assay, Plasmid Preparation

Journal: The Journal of Biological Chemistry

Article Title: In vivo localization of human acetylcholinesterase-derived species in a β-sheet conformation at the core of senile plaques in Alzheimer's disease

doi: 10.1074/jbc.RA118.006230

Figure Lengend Snippet: Localization of hAChE-S, T40 (A) and β-sheet derived hAChE peptides (B) in the brain of hAPPswe single transgenic mice. Frozen brain sections (12 μm) from double transgenic mice were double-labeled with the Elite ABC M.O.M. kit or Vectastain Elite ABC rabbit IgG kit using a biotinylated anti-mouse or anti-rabbit secondary antibody and FITC or Texas red–conjugated avidin (Vector Laboratories). The scale bar represents 10 μm. Shown is one of the z slices from a z stack. The right panels show quantitation of the percentage of overlap between the two fluorophores within the plaques (e.g. HR2 labeling overlapping with KD69 labeling), with plaques examined from at least two different sections per condition (with the order of the antibody/reagents having been switched for the staining). The brains from two different mice were examined. Each value is derived from one individual plaque. Also indicated is the mean, and error bars are S.E.

Article Snippet: ABC mouse-on-mouse (M.O.M.) or Vectastain Elite ABC rabbit IgG kits were used, following the manufacturer's instructions (Vector Laboratories, Burlingame, CA).

Techniques: Derivative Assay, Transgenic Assay, Labeling, Avidin-Biotin Assay, Plasmid Preparation, Quantitation Assay, Staining

Journal: The Journal of Biological Chemistry

Article Title: In vivo localization of human acetylcholinesterase-derived species in a β-sheet conformation at the core of senile plaques in Alzheimer's disease

doi: 10.1074/jbc.RA118.006230

Figure Lengend Snippet: Localization of AChE-derived peptides, as recognized by antisera 3313 (A) and 3314 (B), in the brain of hAPPswe single transgenic mice. Frozen brain sections (12 μm) from single transgenic mice were double-labeled with the Elite ABC M.O.M. kit or Vectastain Elite ABC rabbit IgG kit using a biotinylated anti-mouse or anti-rabbit secondary antibody and FITC or Texas red–conjugated avidin (Vector Laboratories). The scale bar represents 10 μm. Shown is one of the z slices from a z stack. The right panels show quantitation of the percentage of overlap between the two fluorophores within the plaques (e.g. ThS labeling overlapping with 3313 labeling), with plaques examined from at least two different sections per conditions (with the order of the antibody/reagents having been switched for the staining). The brains from two different mice were examined. Each value is derived from one individual plaque. Also indicated is the mean, and error bars are S.E.

Article Snippet: ABC mouse-on-mouse (M.O.M.) or Vectastain Elite ABC rabbit IgG kits were used, following the manufacturer's instructions (Vector Laboratories, Burlingame, CA).

Techniques: Derivative Assay, Transgenic Assay, Labeling, Avidin-Biotin Assay, Plasmid Preparation, Quantitation Assay, Staining

Journal: The Journal of Biological Chemistry

Article Title: In vivo localization of human acetylcholinesterase-derived species in a β-sheet conformation at the core of senile plaques in Alzheimer's disease

doi: 10.1074/jbc.RA118.006230

Figure Lengend Snippet: Localization of hAChE-S, T40 (A), and β-sheet derived hAChE peptides (B) in the brain of hAChE-S/hAPPswe double transgenic mice. Frozen brain sections (12 μm) from double transgenic mice were double-labeled with the Elite ABC M.O.M. kit or Vectastain Elite ABC rabbit IgG kit using a biotinylated anti-mouse or anti-rabbit secondary antibody and FITC or Texas red–conjugated avidin (Vector Laboratories). The scale bar represents 10 μm. Shown is one of the z slices from a z stack, with the inset showing a z projection. The right panels show quantitation of the percentage of overlap between the two fluorophores within the plaques (e.g. HR2 labeling overlapping with Bam10 labeling), with plaques examined from at least two different sections per conditions (with the order of the antibody/reagents have been switched for the staining). The brains from two different mice were examined. Each value is derived from one individual plaque. Also indicated is the mean, and error bars are S.E.

Article Snippet: ABC mouse-on-mouse (M.O.M.) or Vectastain Elite ABC rabbit IgG kits were used, following the manufacturer's instructions (Vector Laboratories, Burlingame, CA).

Techniques: Derivative Assay, Transgenic Assay, Labeling, Avidin-Biotin Assay, Plasmid Preparation, Quantitation Assay, Staining

Journal: The Journal of Biological Chemistry

Article Title: In vivo localization of human acetylcholinesterase-derived species in a β-sheet conformation at the core of senile plaques in Alzheimer's disease

doi: 10.1074/jbc.RA118.006230

Figure Lengend Snippet: Localization of AChE-derived peptides, as recognized by antisera 3313 (A) and 3314 (B), in the brain of hAChE-S/hAPPswe double transgenic mice. Frozen brain sections (12 μm) from double transgenic mice were double-labeled with the Elite ABC M.O.M. kit or Vectastain Elite ABC rabbit IgG kit using a biotinylated anti-mouse or anti-rabbit secondary antibody and FITC or Texas red–conjugated avidin (Vector Laboratories). The scale bar represents 10 μm. Shown is one of the z slices from a z stack, with the insets showing z projections. B, arrowheads indicate hAChE-S species in discrete and distinct areas from those associated with fibrillar amyloids. The right panels show quantitation of the percentage of overlap between the two fluorophores within the plaques (e.g. ThS labeling overlapping with 3313 labeling), with plaques examined from at least two different sections per conditions (with the order of the antibody/reagents having been switched for the staining). The brains from two different mice were examined. Each value is derived from one individual plaque. Also indicated is the mean, and error bars are S.E.

Article Snippet: ABC mouse-on-mouse (M.O.M.) or Vectastain Elite ABC rabbit IgG kits were used, following the manufacturer's instructions (Vector Laboratories, Burlingame, CA).

Techniques: Derivative Assay, Transgenic Assay, Labeling, Avidin-Biotin Assay, Plasmid Preparation, Quantitation Assay, Staining

Journal: International Journal of Molecular Medicine

Article Title: Autophagy relieves the function inhibition and apoptosis-promoting effects on osteoblast induced by glucocorticoid

doi: 10.3892/ijmm.2017.3270

Figure Lengend Snippet: Apoptosis is induced during Dex treatment. (A) Annexin V-fluorescein isothiocyanate/PI staining was analyzed by flow cytometry. Representative dot plots of MC3T3-E1 cells subjected to 10 −8 , 10 −6 and 10 −4 mol/l Dex or control are presented with the percentage of Annexin V-positive cells indicated. (B) Population of apoptotic cells was increased in a dose-dependent manner. There was significant difference between 10 −6 mol/l and control group (P<0.05), 10 −4 mol/l and control group (P<0.05) at all time-points. Dex, dexamethasone; PI, propidium iodide.

Article Snippet: Following treatment with Dex, 3-MA and rapamycin, apoptosis incidence in OBs was detected by using the Annexin V-fluorescein isothiocyanate (FITC) staining kit (BD Pharmingen, Franklin Lakes, NJ, USA).

Techniques: Staining, Flow Cytometry

Journal: International Journal of Molecular Medicine

Article Title: Autophagy relieves the function inhibition and apoptosis-promoting effects on osteoblast induced by glucocorticoid

doi: 10.3892/ijmm.2017.3270

Figure Lengend Snippet: Regulation of autophagy interferes in the negative effects of Dex on MC3T3-E1 cells. (A) MC3T3-E1 cells were treated with Dex, in the presence or absence of chemical inhibitors of autophagy 3-MA. Annexin V-fluorescein isothiocyanate/PI staining was analyzed by flow cytometry. Representative dot plots of MC3T3-E1 cells subjected to Dex or 3-MA alone are presented. (B) Data expressed as percentage of Annexin V-positive cells (n=3). * P<0.05, vs. corresponding control and # P<0.05 other groups vs. 3-MA alone. Cells were treated with 10 −8 mol/l Dex and 3-MA (400 μ mol/l) or RAPA (500 nmol/l), and the expression of (C) Bax and (D) Bcl-2 were detected by western blotting. The results were representative of three independent experiments. β-actin was used as a loading control. * P<0.05; ** P<0.05. 3-MA, 3-methyladenine; PI, propidium iodide; Dex, dexamethasone; RAPA, rapamycin; Bax, BAX apoptosis regulator; Bcl-2, Bcl-2 apoptosis regulator.

Article Snippet: Following treatment with Dex, 3-MA and rapamycin, apoptosis incidence in OBs was detected by using the Annexin V-fluorescein isothiocyanate (FITC) staining kit (BD Pharmingen, Franklin Lakes, NJ, USA).

Techniques: Staining, Flow Cytometry, Expressing, Western Blot

Journal: International Journal of Oncology

Article Title: miR-3664-5P suppresses the proliferation and metastasis of gastric cancer by attenuating the NF-κB signaling pathway through targeting MTDH

doi: 10.3892/ijo.2019.4680

Figure Lengend Snippet: Overexpression of miR-3664-5P increases apoptosis and cell cycle arrest. Following treatment with miR-3664-5P inhibitors and mimics for 48 h, GC cells were stained and analyzed by flow cytometry. (A) The cell cycle distribution of MKN45 and MGC803 cells was detected using flow cytometry and the comparative analysis of cell numbers in the G1 and S + G2 phases are presented. (B) Total apoptosis of MKN45 and MGC803 cells was determined by flow cytometry and the comparative analysis of the number of apoptotic cells is presented. Data are presented as the mean ± standard error of the mean, from three independent experiments. ** P<0.01 and *** P<0.001 vs. the corresponding NC. NC, negative control; miR-inhibitor-NC, cells transfected with the negative control of the miR-3664-5P inhibitor; miR-inhibitors, cells transfected with miR-3664-5P inhibitors; miR-mimics-NC, cells transfected with the negative control of the miR-3664-5P mimics; miR-mimics, cells transfected with miR-3664-5P mimics; miR, microRNA; GC, gastric cancer.

Article Snippet: Cells (6×10 5 ) were cultured in 6-well plates and treated with 50 µ M H 2 O 2 for 2 h at 37°C to induce apoptosis and were then analyzed using an Annexin V-fluorescein isothiocyanate/propidium iodide Apoptosis Detection kit (Vazyme).

Techniques: Over Expression, Staining, Flow Cytometry, Negative Control, Transfection