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  • 99
    Thermo Fisher anti map 2
    Entry of P85 in primary rat cortical neurons visualized by confocal microscopy (A) Cells exposed to TRITC-P85 (red, 0.001% w/v) for 1 hr and then stained by DRAQ-5 (20 nM, 30 min.) to visualize nucleus. (B) Cells exposed to TRITC-P85 as in panel (A) and then fixed and stained by <t>anti-MAP-2</t> antibody (10 μg/ml, 4° C overnight) and Alexa 488- labeled secondary antibody. (C) Cells exposed to TRITC-P85 (0.001% w/v) in the presence of Alexa 488-labeled CTB (5 μg/ml) for 60 min, washed and subjected to time lapse live cell imaging after 5 and 30 min.
    Anti Map 2, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 99/100, based on 41 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore anti map2
    Axin directs axon formation in cultured hippocampal neurons. A–F , Axin is required for axon formation. Dissociated hippocampal neurons were transfected with shAxin construct, together with GFP expression construct, and fixed at 3 DIV ( A , B ), 4 DIV ( C , D ), or 5 DIV ( E , F ) after transfection. Neurons were costained with antibodies against GFP, axonal markers Tau-1 (green), smi-312 (green), or synapsin I (Syn I; green), and dendritic marker <t>MAP2</t> (red) as indicated. The axon was defined as Tau-1, smi-312, or synapsin I positive, longer than 100 μm, and at least twice as long as the other processes. Arrows indicated the axons; arrowheads indicated the non-axon neurites. Scale bar, 20 μm. B , D , F , Quantitative analyses of axonal phenotypes. The axonal phenotypes of transfected hippocampal neurons were classified into three groups: NA (no-axon; negative for Tau-1, smi-312, or Syn I but positive for MAP2), SA (single-axon), and MA (multiple-axon) neurons. More than 300 neurons were examined and quantified for each condition. Data were presented as mean ± SEM; ** p
    Anti Map2, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2079 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Millipore map2
    Microglial expression of TREM2 is variable among HAND donors. Vibratome sections of frontal cortex were double immunolabeled for TREM2 (red) and cell-specific markers (green) for microglia (IBA1), neurons <t>(MAP2),</t> and astroglia (GFAP). ( A ) To identify TREM2 expression on microglia, vibratome sections of frontal cortex tissues were immunolabeled for TREM2 (red) and IBA1 (green), and then visualized using confocal microscopy. ( B ) To identify TREM2 expression on neurons, vibratome sections of frontal cortex tissues were immunolabeled for TREM2 (red) and MAP2 (green), and then visualized using confocal microscopy. ( C ) To identify TREM2 expression on astroglia, vibratome sections of frontal cortex tissues were immunolabeled for TREM2 (red) and GFAP (green), and then visualized using confocal microscopy. Bar = 10μm. Analyzed by two-way ANOVA; **p
    Map2, supplied by Millipore, used in various techniques. Bioz Stars score: 92/100, based on 4066 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Cell Signaling Technology Inc anti map 2
    The effects of Ly-NSC-CM treatment on neuronal damage. (a) Neocortical neuron identification: analysis of <t>MAP-2</t> (green) protein immunofluorescence in mouse primary neocortical neuron cultures. (b) Neurons were treated with t-BHP from 2.5 μ M to 40 μ M for 24 h, and cell viability was examined by MTT assay. (c) The morphology of neocortical neurons and their neurite extensions. (d) Neuronal cell viability according to an MTT assay. n = 3 per group. Data are described as mean ± SEM. ∗ P
    Anti Map 2, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 96/100, based on 30 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Millipore map2 a b
    The effects of Ly-NSC-CM treatment on neuronal damage. (a) Neocortical neuron identification: analysis of <t>MAP-2</t> (green) protein immunofluorescence in mouse primary neocortical neuron cultures. (b) Neurons were treated with t-BHP from 2.5 μ M to 40 μ M for 24 h, and cell viability was examined by MTT assay. (c) The morphology of neocortical neurons and their neurite extensions. (d) Neuronal cell viability according to an MTT assay. n = 3 per group. Data are described as mean ± SEM. ∗ P
    Map2 A B, supplied by Millipore, used in various techniques. Bioz Stars score: 88/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    93
    Millipore mouse anti map2
    Postsynaptic expression of SAP97 enhances expression of presynaptic proteins. A–F , Representative images of hippocampal neurons transfected with GFP–SAP97 ( a1 , c1 , e1 ) or GFP ( b1 , d1 , f1 ) and double stained for synaptophysin ( a2 , b2 ), synapsin ( c2 , d2 ), or Bassoon ( e2 , f2 ) and <t>MAP2</t> ( a3–f3 ) to visualize the dendrites. Individual channels are shown in grayscale for better resolution. Enlargements of the boxed regions are shown in the respective right panels ( a1′–f2′ ). Scale bar: (in a1 ) 10 μm. G , Quantitative analysis of changes in the average puncta intensity of the indicated presynaptic protein induced by overexpression of GFP–SAP97 (gray bars) or GFP (black bars). Bars show mean ± SEM expressed as percentage of average puncta intensity in neighboring untransfected neurons in the same microscopic field ( n = number of cells). *** p
    Mouse Anti Map2, supplied by Millipore, used in various techniques. Bioz Stars score: 93/100, based on 1751 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore map2 primary antibody
    Deletion of Apoa1 and Apoe significantly affect dendritic morphology in CA1 but not in CA2 region of hippocampus. <t>MAP2</t> and H3342 staining were used for dendritic tree reconstruction in the hippocampal CA, and CA2 regions of APP/WT, APP/Abca1 ko and APP/DKO
    Map2 Primary Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 22 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    86
    WuXi AppTec anti map 2
    Neural differentiation of BMSCs on the 6 th day of in vitro transfection. (A, B) Immunofluorescence for the neuronal markers β-III tubulin (A1 and A3; red) and <t>MAP-2</t> (B1 and B3; green) in BMSCs transfected with pLVX-EN-rno-miR124 (miR-124 + ) after 6 days of in vitro differentiation. Scale bars: 100 μm. BMSCs developed dendrites and neurites, similar to neurons (A2, A4, B2, B4). Scale bars: 100 μm. (C) Western blotting was used to detect the expression of synaptophysin (38 kDa). BMSCs: Bone marrow-derived mesenchymal stem cells; MAP-2: microtubule-associated protein-2.
    Anti Map 2, supplied by WuXi AppTec, used in various techniques. Bioz Stars score: 86/100, based on 7 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore monoclonal anti map2 antibody
    CD47 is the presynaptic receptor for SIRPα-mediated presynaptic maturation (a,b) Hippocampal neurons were stained for CD47 together with neurofilament, <t>MAP2</t> or SIRPα. CD47 puncta are abundant in neurofilament-positive axons and not in MAP2-positve dendrites (a) and co-localized with SIRPα (b). Reproduced five times. (c) Hippocampal neurons prepared from CD47KO mice and control littermates were treated with sSIRPα and stained for VGLUT1. sSIRPα does not increase the number and size of VGLUT1 puncta in CD47KO neurons. n = 80/93/102/108 fields from 4 cultures. (d) HEK cells expressing SIRPα, neuroligin1, or control HEK cells (labeled with GFP) were co-cultured with hippocampal neurons prepared from WT or CD47KO mice for 2 days and stained for synapsin. HEK cells expressing SIRPα induce presynaptic differentiation in co-cultured WT hippocampal neurons, but fail to do so in co-cultured CD47KO neurons. HEK cells expressing neuroligin1 induce presynaptic differentiation in both WT and CD47KO neurons. Graphs show quantification of synapsin puncta number and size. Data are from 38/34/34/35/42/40 fields from 5 cultures. (e) Presynaptic expression of CD47 restores responsiveness to sSIRPα in CD47KO neurons. Hippocampal neurons prepared from WT or CD47KO mice were transfected with the synaptophysin-YFP plasmid or with the synaptophysin-YFP and CD47 plasmids. Cultures were treated with sSIRPα, and presynaptic differentiation of transfected neurons was assessed by synaptophysin-YFP clustering. sSIRPα does not increase the number and size of synaptophysin-YFP puncta in CD47KO neurons, but introduction of CD47 restores responsiveness to sSIRPα. n = 33/37/33/35/37/36 neurites from 3 cultures. (f) Postsynaptic expression of SIRPα rescues presynaptic defects in SIRPαKO neurons. Hippocampal neurons prepared from conditional SIRPαKO mice were transfected with an mCherry plasmid (control), Cre and mCherry plasmids (knockout) or Cre, mCherry and SIRPα plasmids (rescue) and stained for VGLUT1. Both the number and size of VGLUT1 puncta on mCherry-expressing dendrites were decreased in SIRPαKO relative to control, but postsynaptic expression of SIRPα rescues the defects. n = 41/33/36 neurites from 3 cultures. Student’s t-test (c) or ANOVA followed by Tukey test (d–f). Scale bar, 5 μm (c), 10 μm (all others).
    Monoclonal Anti Map2 Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 946 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Abcam monoclonal map 2 antibody
    CD47 is the presynaptic receptor for SIRPα-mediated presynaptic maturation (a,b) Hippocampal neurons were stained for CD47 together with neurofilament, <t>MAP2</t> or SIRPα. CD47 puncta are abundant in neurofilament-positive axons and not in MAP2-positve dendrites (a) and co-localized with SIRPα (b). Reproduced five times. (c) Hippocampal neurons prepared from CD47KO mice and control littermates were treated with sSIRPα and stained for VGLUT1. sSIRPα does not increase the number and size of VGLUT1 puncta in CD47KO neurons. n = 80/93/102/108 fields from 4 cultures. (d) HEK cells expressing SIRPα, neuroligin1, or control HEK cells (labeled with GFP) were co-cultured with hippocampal neurons prepared from WT or CD47KO mice for 2 days and stained for synapsin. HEK cells expressing SIRPα induce presynaptic differentiation in co-cultured WT hippocampal neurons, but fail to do so in co-cultured CD47KO neurons. HEK cells expressing neuroligin1 induce presynaptic differentiation in both WT and CD47KO neurons. Graphs show quantification of synapsin puncta number and size. Data are from 38/34/34/35/42/40 fields from 5 cultures. (e) Presynaptic expression of CD47 restores responsiveness to sSIRPα in CD47KO neurons. Hippocampal neurons prepared from WT or CD47KO mice were transfected with the synaptophysin-YFP plasmid or with the synaptophysin-YFP and CD47 plasmids. Cultures were treated with sSIRPα, and presynaptic differentiation of transfected neurons was assessed by synaptophysin-YFP clustering. sSIRPα does not increase the number and size of synaptophysin-YFP puncta in CD47KO neurons, but introduction of CD47 restores responsiveness to sSIRPα. n = 33/37/33/35/37/36 neurites from 3 cultures. (f) Postsynaptic expression of SIRPα rescues presynaptic defects in SIRPαKO neurons. Hippocampal neurons prepared from conditional SIRPαKO mice were transfected with an mCherry plasmid (control), Cre and mCherry plasmids (knockout) or Cre, mCherry and SIRPα plasmids (rescue) and stained for VGLUT1. Both the number and size of VGLUT1 puncta on mCherry-expressing dendrites were decreased in SIRPαKO relative to control, but postsynaptic expression of SIRPα rescues the defects. n = 41/33/36 neurites from 3 cultures. Student’s t-test (c) or ANOVA followed by Tukey test (d–f). Scale bar, 5 μm (c), 10 μm (all others).
    Monoclonal Map 2 Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    96
    Abcam antibody against map 2
    Sodium hydrosulfide treatment (250 μM) could significantly repair OGD-induced injuries of neurite sprouting and outgrowth in the young and aged hippocampal neurons in 7 DIV. (A) Typical hippocampal neurons with extending neurites in the cultures, stained for <t>MAP-2.</t> Scale bar = 40 μm. The neurite sprouting and outgrowth were analyzed by the number of primary dendrites per cell (B) , the number of dendritic end tips (C) , and the average neurite length (D) . Data were presented by the mean ± SD. ∗ p
    Antibody Against Map 2, supplied by Abcam, used in various techniques. Bioz Stars score: 96/100, based on 5 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Abcam anti map 2 antibody
    HIV + sup ± morphine-mediated neurite damage. Cells were fixed at specific intervals after treatment and labeled for <t>MAP-2</t> (green) and TUNEL (red). ( A ) Digital images of neuronal cultures at 72 h after treatment; scale bar = 40 µm. ( B ) The ‘Sholl score’ was assessed only for TUNEL(-) neurons in the digital images and converted into neurite length in µm via a micrometer-scale calibration. The findings were reported as average total neurite length per neuron (µm) ± SEM. Significance was analyzed by one-way ANOVA and Duncan's post hoc test from n = 4 separate experiments. At all time-points and in both culture systems, all groups exposed to HIV + sup showed significantly reduced neurite length (* p
    Anti Map 2 Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 31 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
    https://www.bioz.com/result/anti map 2 antibody/product/Abcam
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    97
    Abcam anti map 2
    Characterization of human glioblastoma stem cells. Representative images of GSC (derived from GBM6) spheroids (A) and monolayer on Matrigel (B) cultured in stem cell medium supplemented with EGF and bFGF and differentiated cells (C) obtained by switching culture conditions from growth factor-deprived to FBS-containing medium. Left panels reproduce phase contrast images at different magnification (10× and 20×). Right panels depict immunofluorescence for stem cell markers OLIG2, nestin, and Oct4 (A,B) , and the glia (GFAP) and neuron <t>(MAP-2)</t> differentiation markers (C) . GFAP and MAP-2 expression is depicted from the same microscope field, showing in most cells the co-expression of both markers. Similar results were obtained analyzing all the GSC cultures that enter the study.
    Anti Map 2, supplied by Abcam, used in various techniques. Bioz Stars score: 97/100, based on 52 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Entry of P85 in primary rat cortical neurons visualized by confocal microscopy (A) Cells exposed to TRITC-P85 (red, 0.001% w/v) for 1 hr and then stained by DRAQ-5 (20 nM, 30 min.) to visualize nucleus. (B) Cells exposed to TRITC-P85 as in panel (A) and then fixed and stained by anti-MAP-2 antibody (10 μg/ml, 4° C overnight) and Alexa 488- labeled secondary antibody. (C) Cells exposed to TRITC-P85 (0.001% w/v) in the presence of Alexa 488-labeled CTB (5 μg/ml) for 60 min, washed and subjected to time lapse live cell imaging after 5 and 30 min.

    Journal: Biomaterials

    Article Title: The Utilization of Pathogen-Like Cellular Trafficking by Single Chain Block Copolymer

    doi: 10.1016/j.biomaterials.2009.11.020

    Figure Lengend Snippet: Entry of P85 in primary rat cortical neurons visualized by confocal microscopy (A) Cells exposed to TRITC-P85 (red, 0.001% w/v) for 1 hr and then stained by DRAQ-5 (20 nM, 30 min.) to visualize nucleus. (B) Cells exposed to TRITC-P85 as in panel (A) and then fixed and stained by anti-MAP-2 antibody (10 μg/ml, 4° C overnight) and Alexa 488- labeled secondary antibody. (C) Cells exposed to TRITC-P85 (0.001% w/v) in the presence of Alexa 488-labeled CTB (5 μg/ml) for 60 min, washed and subjected to time lapse live cell imaging after 5 and 30 min.

    Article Snippet: For detection of anti-MAP-2, mouse-specific IgG secondary antibody (1:250) conjugated to Alexa 488 (Invitrogen Inc, Carlsbad, CA.) was added to cells and visualized using confocal microscopy.

    Techniques: Confocal Microscopy, Staining, Labeling, CtB Assay, Live Cell Imaging

    Amino acid sequences of the MAP peptides and their physical locations on P . falciparum antigens. (A) MAP-1 vaccine. (B) MAP-2 vaccine. (C) MAP-3 vaccine.

    Journal: Infection and Immunity

    Article Title: Multiple Antigen Peptide Vaccines against Plasmodium falciparum Malaria ▿

    doi: 10.1128/IAI.00533-10

    Figure Lengend Snippet: Amino acid sequences of the MAP peptides and their physical locations on P . falciparum antigens. (A) MAP-1 vaccine. (B) MAP-2 vaccine. (C) MAP-3 vaccine.

    Article Snippet: A 100-μl aliquot of anti-MAP-2 antibody at a 1:50 dilution in PBS was added to each well of the Lab-Tek slide containing fixed liver stage parasites developing in HC-04 cells and incubated at 37°C for 1 h. After three washings, 5 min each at room temperature, a 100-μl aliquot of goat anti-mouse Alexa Fluor 488 (Invitrogen Corporation) at a 1:200 dilution in PBS containing a 1:1,000 dilution of 4′,6-diamidino-2-phenylindole (DAPI) from a 2-mg/ml stock and 0.001% Evans blue, was added to each well of the Lab-Tek slide.

    Techniques:

    IIF IgG reactivity against P . falciparum liver stage parasites in C57BL/6, A/J, BALB/c, and CD1 strains of mice that received three subcutaneous injections of MAP vaccines in Montanide ISA 51. The reactivity for liver stage parasites was tested against (A) polyclonal mouse anti-LSA-1 antibody (positive control), (B) MAP-2-immunized BALB/c mouse sera, (C) C57BL/6-immunized mouse sera, (D) CD1-immunized mouse sera, and (E) normal mouse serum. The arrow points to the developing P . falciparum parasites. The hepatocyte nucleus is stained blue with DAPI.

    Journal: Infection and Immunity

    Article Title: Multiple Antigen Peptide Vaccines against Plasmodium falciparum Malaria ▿

    doi: 10.1128/IAI.00533-10

    Figure Lengend Snippet: IIF IgG reactivity against P . falciparum liver stage parasites in C57BL/6, A/J, BALB/c, and CD1 strains of mice that received three subcutaneous injections of MAP vaccines in Montanide ISA 51. The reactivity for liver stage parasites was tested against (A) polyclonal mouse anti-LSA-1 antibody (positive control), (B) MAP-2-immunized BALB/c mouse sera, (C) C57BL/6-immunized mouse sera, (D) CD1-immunized mouse sera, and (E) normal mouse serum. The arrow points to the developing P . falciparum parasites. The hepatocyte nucleus is stained blue with DAPI.

    Article Snippet: A 100-μl aliquot of anti-MAP-2 antibody at a 1:50 dilution in PBS was added to each well of the Lab-Tek slide containing fixed liver stage parasites developing in HC-04 cells and incubated at 37°C for 1 h. After three washings, 5 min each at room temperature, a 100-μl aliquot of goat anti-mouse Alexa Fluor 488 (Invitrogen Corporation) at a 1:200 dilution in PBS containing a 1:1,000 dilution of 4′,6-diamidino-2-phenylindole (DAPI) from a 2-mg/ml stock and 0.001% Evans blue, was added to each well of the Lab-Tek slide.

    Techniques: Mouse Assay, Positive Control, Staining

    Axin directs axon formation in cultured hippocampal neurons. A–F , Axin is required for axon formation. Dissociated hippocampal neurons were transfected with shAxin construct, together with GFP expression construct, and fixed at 3 DIV ( A , B ), 4 DIV ( C , D ), or 5 DIV ( E , F ) after transfection. Neurons were costained with antibodies against GFP, axonal markers Tau-1 (green), smi-312 (green), or synapsin I (Syn I; green), and dendritic marker MAP2 (red) as indicated. The axon was defined as Tau-1, smi-312, or synapsin I positive, longer than 100 μm, and at least twice as long as the other processes. Arrows indicated the axons; arrowheads indicated the non-axon neurites. Scale bar, 20 μm. B , D , F , Quantitative analyses of axonal phenotypes. The axonal phenotypes of transfected hippocampal neurons were classified into three groups: NA (no-axon; negative for Tau-1, smi-312, or Syn I but positive for MAP2), SA (single-axon), and MA (multiple-axon) neurons. More than 300 neurons were examined and quantified for each condition. Data were presented as mean ± SEM; ** p

    Journal: The Journal of Neuroscience

    Article Title: Cdk5-Mediated Phosphorylation of Axin Directs Axon Formation during Cerebral Cortex Development

    doi: 10.1523/JNEUROSCI.3120-11.2011

    Figure Lengend Snippet: Axin directs axon formation in cultured hippocampal neurons. A–F , Axin is required for axon formation. Dissociated hippocampal neurons were transfected with shAxin construct, together with GFP expression construct, and fixed at 3 DIV ( A , B ), 4 DIV ( C , D ), or 5 DIV ( E , F ) after transfection. Neurons were costained with antibodies against GFP, axonal markers Tau-1 (green), smi-312 (green), or synapsin I (Syn I; green), and dendritic marker MAP2 (red) as indicated. The axon was defined as Tau-1, smi-312, or synapsin I positive, longer than 100 μm, and at least twice as long as the other processes. Arrows indicated the axons; arrowheads indicated the non-axon neurites. Scale bar, 20 μm. B , D , F , Quantitative analyses of axonal phenotypes. The axonal phenotypes of transfected hippocampal neurons were classified into three groups: NA (no-axon; negative for Tau-1, smi-312, or Syn I but positive for MAP2), SA (single-axon), and MA (multiple-axon) neurons. More than 300 neurons were examined and quantified for each condition. Data were presented as mean ± SEM; ** p

    Article Snippet: Monoclonal antibodies used for immunostaining or immunoblotting included the following: anti-MAP2, anti-α-tubulin, anti-β-tubulin III (TuJ1), and anti-acetylated α-tubulin (Ace-α-tub) from Sigma; anti-Tau-1, anti-MAP2, and anti-β-actin from Millipore Bioscience Research Reagents; anti-GSK-3β from BD Transduction Laboratory and from Millipore; anti-Cdk5 (DC-17) from Santa Cruz Biotechnology; anti-CRMP-2 from IBL; anti-TAG1 (4D7) from Developmental Studies Hybridoma Bank; and anti-smi-312 from Covance.

    Techniques: Cell Culture, Transfection, Construct, Expressing, Marker

    Microglial expression of TREM2 is variable among HAND donors. Vibratome sections of frontal cortex were double immunolabeled for TREM2 (red) and cell-specific markers (green) for microglia (IBA1), neurons (MAP2), and astroglia (GFAP). ( A ) To identify TREM2 expression on microglia, vibratome sections of frontal cortex tissues were immunolabeled for TREM2 (red) and IBA1 (green), and then visualized using confocal microscopy. ( B ) To identify TREM2 expression on neurons, vibratome sections of frontal cortex tissues were immunolabeled for TREM2 (red) and MAP2 (green), and then visualized using confocal microscopy. ( C ) To identify TREM2 expression on astroglia, vibratome sections of frontal cortex tissues were immunolabeled for TREM2 (red) and GFAP (green), and then visualized using confocal microscopy. Bar = 10μm. Analyzed by two-way ANOVA; **p

    Journal: Journal of neurochemistry

    Article Title: Alterations in brain TREM2 and Amyloid-β levels are associated with neurocognitive impairment in HIV-infected persons on antiretroviral therapy

    doi: 10.1111/jnc.14582

    Figure Lengend Snippet: Microglial expression of TREM2 is variable among HAND donors. Vibratome sections of frontal cortex were double immunolabeled for TREM2 (red) and cell-specific markers (green) for microglia (IBA1), neurons (MAP2), and astroglia (GFAP). ( A ) To identify TREM2 expression on microglia, vibratome sections of frontal cortex tissues were immunolabeled for TREM2 (red) and IBA1 (green), and then visualized using confocal microscopy. ( B ) To identify TREM2 expression on neurons, vibratome sections of frontal cortex tissues were immunolabeled for TREM2 (red) and MAP2 (green), and then visualized using confocal microscopy. ( C ) To identify TREM2 expression on astroglia, vibratome sections of frontal cortex tissues were immunolabeled for TREM2 (red) and GFAP (green), and then visualized using confocal microscopy. Bar = 10μm. Analyzed by two-way ANOVA; **p

    Article Snippet: The following antibodies were used in immunoblot, immunohistochemistry or both: TREM2 (1:400; R & D Systems; Cat# AF1828; RRID:AB_2208689), ACTB (1:2000; Sigma-Aldrich; Cat# A5316; RRID:AB_476743), MAP2 (1:200; Sigma-Aldrich Cat#M4403; RRID:AB_477193), GFAP (1:200; Dako Cat# Z0334; RRID:AB_10013382), and IBA1 (1:1000; WAKO Cat#019–19741; RRID:AB_839504).

    Techniques: Expressing, Immunolabeling, Confocal Microscopy

    The effects of Ly-NSC-CM treatment on neuronal damage. (a) Neocortical neuron identification: analysis of MAP-2 (green) protein immunofluorescence in mouse primary neocortical neuron cultures. (b) Neurons were treated with t-BHP from 2.5 μ M to 40 μ M for 24 h, and cell viability was examined by MTT assay. (c) The morphology of neocortical neurons and their neurite extensions. (d) Neuronal cell viability according to an MTT assay. n = 3 per group. Data are described as mean ± SEM. ∗ P

    Journal: Oxidative Medicine and Cellular Longevity

    Article Title: The Secretion from Neural Stem Cells Pretreated with Lycopene Protects against tert-Butyl Hydroperoxide-Induced Neuron Oxidative Damage

    doi: 10.1155/2018/5490218

    Figure Lengend Snippet: The effects of Ly-NSC-CM treatment on neuronal damage. (a) Neocortical neuron identification: analysis of MAP-2 (green) protein immunofluorescence in mouse primary neocortical neuron cultures. (b) Neurons were treated with t-BHP from 2.5 μ M to 40 μ M for 24 h, and cell viability was examined by MTT assay. (c) The morphology of neocortical neurons and their neurite extensions. (d) Neuronal cell viability according to an MTT assay. n = 3 per group. Data are described as mean ± SEM. ∗ P

    Article Snippet: After conditioned medium treatment, the medium was decanted and the cells were washed three times with PBS; they were then fixed with 4% paraformaldehyde for 30 min. Then NSCs and neurons were permeabilized with 0.3% Triton X-100 in PBS for 15 min. After being blocked in 3% bovine serum albumin (BSA) with PBS at room temperature for 1 h, NSCs and neurons were washed again in PBS and finally incubated overnight at 4°C with one of anti-MAP 2, anti-nestin, or anti-cleaved caspase-3 antibodies (1 : 200, Cell Signaling Technology, USA).

    Techniques: Immunofluorescence, MTT Assay

    Neural differentiation of BMSCs on the 6 th day of in vitro transfection. (A, B) Immunofluorescence for the neuronal markers β-III tubulin (A1 and A3; red) and MAP-2 (B1 and B3; green) in BMSCs transfected with pLVX-EN-rno-miR124 (miR-124 + ) after 6 days of in vitro differentiation. Scale bars: 100 μm. BMSCs developed dendrites and neurites, similar to neurons (A2, A4, B2, B4). Scale bars: 100 μm. (C) Western blotting was used to detect the expression of synaptophysin (38 kDa). BMSCs: Bone marrow-derived mesenchymal stem cells; MAP-2: microtubule-associated protein-2.

    Journal: Neural Regeneration Research

    Article Title: Overexpression of microRNA-124 promotes the neuronal differentiation of bone marrow-derived mesenchymal stem cells

    doi: 10.4103/1673-5374.135333

    Figure Lengend Snippet: Neural differentiation of BMSCs on the 6 th day of in vitro transfection. (A, B) Immunofluorescence for the neuronal markers β-III tubulin (A1 and A3; red) and MAP-2 (B1 and B3; green) in BMSCs transfected with pLVX-EN-rno-miR124 (miR-124 + ) after 6 days of in vitro differentiation. Scale bars: 100 μm. BMSCs developed dendrites and neurites, similar to neurons (A2, A4, B2, B4). Scale bars: 100 μm. (C) Western blotting was used to detect the expression of synaptophysin (38 kDa). BMSCs: Bone marrow-derived mesenchymal stem cells; MAP-2: microtubule-associated protein-2.

    Article Snippet: Sections were rinsed with PBS, and incubated with mouse anti-β-III tubulin (1:100, Santa Cruz Biotechnology, Santa Cruz, CA, USA) or mouse anti-MAP-2 (1:50, Cell Signaling, Boston, MA, USA) for 24 hours at 4°C.

    Techniques: In Vitro, Transfection, Immunofluorescence, Western Blot, Expressing, Derivative Assay

    Postsynaptic expression of SAP97 enhances expression of presynaptic proteins. A–F , Representative images of hippocampal neurons transfected with GFP–SAP97 ( a1 , c1 , e1 ) or GFP ( b1 , d1 , f1 ) and double stained for synaptophysin ( a2 , b2 ), synapsin ( c2 , d2 ), or Bassoon ( e2 , f2 ) and MAP2 ( a3–f3 ) to visualize the dendrites. Individual channels are shown in grayscale for better resolution. Enlargements of the boxed regions are shown in the respective right panels ( a1′–f2′ ). Scale bar: (in a1 ) 10 μm. G , Quantitative analysis of changes in the average puncta intensity of the indicated presynaptic protein induced by overexpression of GFP–SAP97 (gray bars) or GFP (black bars). Bars show mean ± SEM expressed as percentage of average puncta intensity in neighboring untransfected neurons in the same microscopic field ( n = number of cells). *** p

    Journal: The Journal of Neuroscience

    Article Title: Transsynaptic Signaling by Postsynaptic Synapse-Associated Protein 97

    doi: 10.1523/JNEUROSCI.5247-05.2006

    Figure Lengend Snippet: Postsynaptic expression of SAP97 enhances expression of presynaptic proteins. A–F , Representative images of hippocampal neurons transfected with GFP–SAP97 ( a1 , c1 , e1 ) or GFP ( b1 , d1 , f1 ) and double stained for synaptophysin ( a2 , b2 ), synapsin ( c2 , d2 ), or Bassoon ( e2 , f2 ) and MAP2 ( a3–f3 ) to visualize the dendrites. Individual channels are shown in grayscale for better resolution. Enlargements of the boxed regions are shown in the respective right panels ( a1′–f2′ ). Scale bar: (in a1 ) 10 μm. G , Quantitative analysis of changes in the average puncta intensity of the indicated presynaptic protein induced by overexpression of GFP–SAP97 (gray bars) or GFP (black bars). Bars show mean ± SEM expressed as percentage of average puncta intensity in neighboring untransfected neurons in the same microscopic field ( n = number of cells). *** p

    Article Snippet: The following primary antibodies and dilutions were used: rabbit anti-microtubule-associated protein 2 (anti-MAP2; 1:1500; Chemicon, Temecula, CA), mouse anti-MAP2 (1:800; Sigma), mouse anti-PSD-95 (1:100; Sigma), rat anti-Homer 1b,c (1:100; Chemicon), rabbit anti-synaptohysin (1:2000; Zymed, San Francisco, CA), mouse monoclonal anti-synapsin (1:2000; Chemicon), rabbit anti-SPAL (1:100; gift from T. Akiyama, Institute for Molecular and Cellular Biosciences, University of Tokyo, Tokyo, Japan), rabbit anti-Shank-1A (1:100; gift from M. Sheng, Massachusetts Institute of Technology, Cambridge, MA), rabbit antibody directed at the N terminus of GluR1 (1:1000; gift from R. Huganir, Johns Hopkins University School of Medicine, Baltimore, MD), rabbit anti-ProSAP2/Shank3 antibodies (1:100; gift from Tobias Boecher, Magdeburg, Germany).

    Techniques: Expressing, Transfection, Staining, Over Expression

    Specific structural domains of SAP97 are required for its presynaptic effects. A , Representative images of neurons transfected with GFP–SAP97 ( a1 ), GFP–SAP97ΔS97N ( b1 ), GFP–SAP97ΔGK ( c1 ), and GFP–SAP97ΔPDZ1–3 ( d1 ). Each neuron was double stained for Bassoon ( a2–d2 ) and MAP2 (not shown). Merged images are shown in a3–d3 . Scale bar: (in a1 ) 10 μm. B , Left, Schematic diagrams of SAP97 constructs used in this study. Different structural domains are indicated. Right, Quantification of changes in Bassoon puncta intensity induced by the different SAP97 constructs. The black bars show the average intensity of Bassoon puncta that colocalize with the GFP–SAP97 puncta in a single dendrite, and the gray bars show the average intensity of puncta that do not colocalize in the same dendrite (mean ± SEM; numbers in parentheses indicate number of cells examined). Neurons expressing GFP–SAP97ΔS97N, GFP–SAP97ΔGK, GFP–SAP97ΔPDZ3, or GFP–SAP97ΔPDZ1–3 showed a significant decrease in the average Bassoon puncta intensity when compared with neurons overexpressing wild-type GFP–SAP97 (*** p

    Journal: The Journal of Neuroscience

    Article Title: Transsynaptic Signaling by Postsynaptic Synapse-Associated Protein 97

    doi: 10.1523/JNEUROSCI.5247-05.2006

    Figure Lengend Snippet: Specific structural domains of SAP97 are required for its presynaptic effects. A , Representative images of neurons transfected with GFP–SAP97 ( a1 ), GFP–SAP97ΔS97N ( b1 ), GFP–SAP97ΔGK ( c1 ), and GFP–SAP97ΔPDZ1–3 ( d1 ). Each neuron was double stained for Bassoon ( a2–d2 ) and MAP2 (not shown). Merged images are shown in a3–d3 . Scale bar: (in a1 ) 10 μm. B , Left, Schematic diagrams of SAP97 constructs used in this study. Different structural domains are indicated. Right, Quantification of changes in Bassoon puncta intensity induced by the different SAP97 constructs. The black bars show the average intensity of Bassoon puncta that colocalize with the GFP–SAP97 puncta in a single dendrite, and the gray bars show the average intensity of puncta that do not colocalize in the same dendrite (mean ± SEM; numbers in parentheses indicate number of cells examined). Neurons expressing GFP–SAP97ΔS97N, GFP–SAP97ΔGK, GFP–SAP97ΔPDZ3, or GFP–SAP97ΔPDZ1–3 showed a significant decrease in the average Bassoon puncta intensity when compared with neurons overexpressing wild-type GFP–SAP97 (*** p

    Article Snippet: The following primary antibodies and dilutions were used: rabbit anti-microtubule-associated protein 2 (anti-MAP2; 1:1500; Chemicon, Temecula, CA), mouse anti-MAP2 (1:800; Sigma), mouse anti-PSD-95 (1:100; Sigma), rat anti-Homer 1b,c (1:100; Chemicon), rabbit anti-synaptohysin (1:2000; Zymed, San Francisco, CA), mouse monoclonal anti-synapsin (1:2000; Chemicon), rabbit anti-SPAL (1:100; gift from T. Akiyama, Institute for Molecular and Cellular Biosciences, University of Tokyo, Tokyo, Japan), rabbit anti-Shank-1A (1:100; gift from M. Sheng, Massachusetts Institute of Technology, Cambridge, MA), rabbit antibody directed at the N terminus of GluR1 (1:1000; gift from R. Huganir, Johns Hopkins University School of Medicine, Baltimore, MD), rabbit anti-ProSAP2/Shank3 antibodies (1:100; gift from Tobias Boecher, Magdeburg, Germany).

    Techniques: Transfection, Staining, Construct, Expressing

    SAP97 has larger presynaptic effect than PSD-95 and SAP102. A–C , Representative images of hippocampal neurons transfected with GFP–SAP97 ( A ), SAP102–GFP ( B ), or PSD-95–GFP ( C ) and double stained for Bassoon (middle) and MAP2 (right). Individual channels are shown in grayscale for better resolution. Scale bar: (in A ) 10 μm. D , Quantification of the effects of GFP–SAP97 and PSD-95–GFP (left part) or GFP–SAP97 and SAP102–GFP (right) on average Bassoon puncta intensity (mean ± SEM). Comparisons were made between transfected neurons from sister cultures ( n = number of cells). *** p

    Journal: The Journal of Neuroscience

    Article Title: Transsynaptic Signaling by Postsynaptic Synapse-Associated Protein 97

    doi: 10.1523/JNEUROSCI.5247-05.2006

    Figure Lengend Snippet: SAP97 has larger presynaptic effect than PSD-95 and SAP102. A–C , Representative images of hippocampal neurons transfected with GFP–SAP97 ( A ), SAP102–GFP ( B ), or PSD-95–GFP ( C ) and double stained for Bassoon (middle) and MAP2 (right). Individual channels are shown in grayscale for better resolution. Scale bar: (in A ) 10 μm. D , Quantification of the effects of GFP–SAP97 and PSD-95–GFP (left part) or GFP–SAP97 and SAP102–GFP (right) on average Bassoon puncta intensity (mean ± SEM). Comparisons were made between transfected neurons from sister cultures ( n = number of cells). *** p

    Article Snippet: The following primary antibodies and dilutions were used: rabbit anti-microtubule-associated protein 2 (anti-MAP2; 1:1500; Chemicon, Temecula, CA), mouse anti-MAP2 (1:800; Sigma), mouse anti-PSD-95 (1:100; Sigma), rat anti-Homer 1b,c (1:100; Chemicon), rabbit anti-synaptohysin (1:2000; Zymed, San Francisco, CA), mouse monoclonal anti-synapsin (1:2000; Chemicon), rabbit anti-SPAL (1:100; gift from T. Akiyama, Institute for Molecular and Cellular Biosciences, University of Tokyo, Tokyo, Japan), rabbit anti-Shank-1A (1:100; gift from M. Sheng, Massachusetts Institute of Technology, Cambridge, MA), rabbit antibody directed at the N terminus of GluR1 (1:1000; gift from R. Huganir, Johns Hopkins University School of Medicine, Baltimore, MD), rabbit anti-ProSAP2/Shank3 antibodies (1:100; gift from Tobias Boecher, Magdeburg, Germany).

    Techniques: Transfection, Staining

    SAP97 PDZ1 and PDZ2 ligand-binding pockets are critical for its presynaptic effects. A , Representative images of neurons transfected with GFP–SAP97 or GFP–SAP97 PDZ domain mutants, as indicated in the left panels. Each row of images shows GFP fluorescence (left), Bassoon labeling (middle), and merged images (right). Staining for MAP2 is not shown. The arrowheads indicate Bassoon puncta on transfected neurons. The arrows indicate Bassoon puncta on untransfected neighboring neurons. Scale bar, 10 μm. Mutants are point mutants for the ligand-binding pocket of the individual PDZ1 (PDZ1F 203 H), PDZ2 (PDZ2F 298 H), and PDZ3 (PDZ3F 470 H) domains and the three possible combinations of two mutant PDZ domains (PDZ1 2FH, PDZ1 3FH, and PDZ2 3FH). B , Quantification of the average Bassoon puncta intensity that colocalized with puncta of the GFP-tagged SAP97 mutants (mean ± SEM; n = number of cells; *** p

    Journal: The Journal of Neuroscience

    Article Title: Transsynaptic Signaling by Postsynaptic Synapse-Associated Protein 97

    doi: 10.1523/JNEUROSCI.5247-05.2006

    Figure Lengend Snippet: SAP97 PDZ1 and PDZ2 ligand-binding pockets are critical for its presynaptic effects. A , Representative images of neurons transfected with GFP–SAP97 or GFP–SAP97 PDZ domain mutants, as indicated in the left panels. Each row of images shows GFP fluorescence (left), Bassoon labeling (middle), and merged images (right). Staining for MAP2 is not shown. The arrowheads indicate Bassoon puncta on transfected neurons. The arrows indicate Bassoon puncta on untransfected neighboring neurons. Scale bar, 10 μm. Mutants are point mutants for the ligand-binding pocket of the individual PDZ1 (PDZ1F 203 H), PDZ2 (PDZ2F 298 H), and PDZ3 (PDZ3F 470 H) domains and the three possible combinations of two mutant PDZ domains (PDZ1 2FH, PDZ1 3FH, and PDZ2 3FH). B , Quantification of the average Bassoon puncta intensity that colocalized with puncta of the GFP-tagged SAP97 mutants (mean ± SEM; n = number of cells; *** p

    Article Snippet: The following primary antibodies and dilutions were used: rabbit anti-microtubule-associated protein 2 (anti-MAP2; 1:1500; Chemicon, Temecula, CA), mouse anti-MAP2 (1:800; Sigma), mouse anti-PSD-95 (1:100; Sigma), rat anti-Homer 1b,c (1:100; Chemicon), rabbit anti-synaptohysin (1:2000; Zymed, San Francisco, CA), mouse monoclonal anti-synapsin (1:2000; Chemicon), rabbit anti-SPAL (1:100; gift from T. Akiyama, Institute for Molecular and Cellular Biosciences, University of Tokyo, Tokyo, Japan), rabbit anti-Shank-1A (1:100; gift from M. Sheng, Massachusetts Institute of Technology, Cambridge, MA), rabbit antibody directed at the N terminus of GluR1 (1:1000; gift from R. Huganir, Johns Hopkins University School of Medicine, Baltimore, MD), rabbit anti-ProSAP2/Shank3 antibodies (1:100; gift from Tobias Boecher, Magdeburg, Germany).

    Techniques: Ligand Binding Assay, Transfection, Fluorescence, Labeling, Staining, Mutagenesis

    Bexarotene restores dendritic morphology of the CA1 region of the hippocampus of APOE4 mice. A , Representative images of MAP2 staining and dendritic tree reconstruction in the hippocampal CA1 region using Imaris filament tracing software (60×

    Journal: The Journal of Neuroscience

    Article Title: Bexarotene-Activated Retinoid X Receptors Regulate Neuronal Differentiation and Dendritic Complexity

    doi: 10.1523/JNEUROSCI.1001-15.2015

    Figure Lengend Snippet: Bexarotene restores dendritic morphology of the CA1 region of the hippocampus of APOE4 mice. A , Representative images of MAP2 staining and dendritic tree reconstruction in the hippocampal CA1 region using Imaris filament tracing software (60×

    Article Snippet: Primary antibodies were mouse anti-MAP2 antibody (MAB378, 1:2000; Millipore) and goat anti-DCX antibody (sc-8066, 1:1000; Santa Cruz Biotechnology).

    Techniques: Mouse Assay, Staining, Software

    Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f mice did not affect subacute infarction but reduced neuronal injury at the subacute phase postischemia. (a) Representative T2-weighted imaging (T2WI) of ex vivo oil- and Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f brains at 14 days Rp. (b) Infarct volume and brain atrophy in oil- or Tam-treated Cx3cr1-Cre ER ; NHE1 f/f brains at 14 days Rp with T2WI. Data are mean ± SEM. N = 4. (c) Representative MAP2 staining of oil- and Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f brains at 14 days Rp. (d) Infarct volume and brain atrophy in oil- or Tam-treated Cx3cr1-Cre ER ; NHE1 f/f brains at 14 days Rp with MAP2 staining. Data are mean ± SEM. N = 7. (e) Representative immunofluorescent images of MAP2, NeuN, and To-pro-3 staining in the CL, IL perilesion and IL core areas in the cortex and striatum of the oil- and Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f mice at 14 days post-tMCAO. (f) Quantitative analysis of MAP2 intensity in the CL, IL perilesion and IL core areas in the cortex and striatum of the oil- and Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f mice at 14 days post-tMCAO. N = 3. * p

    Journal: Glia

    Article Title: Selective role of Na+/H+ exchanger in Cx3cr1+ microglial activation, white matter demyelination, and post-stroke function recovery

    doi: 10.1002/glia.23456

    Figure Lengend Snippet: Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f mice did not affect subacute infarction but reduced neuronal injury at the subacute phase postischemia. (a) Representative T2-weighted imaging (T2WI) of ex vivo oil- and Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f brains at 14 days Rp. (b) Infarct volume and brain atrophy in oil- or Tam-treated Cx3cr1-Cre ER ; NHE1 f/f brains at 14 days Rp with T2WI. Data are mean ± SEM. N = 4. (c) Representative MAP2 staining of oil- and Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f brains at 14 days Rp. (d) Infarct volume and brain atrophy in oil- or Tam-treated Cx3cr1-Cre ER ; NHE1 f/f brains at 14 days Rp with MAP2 staining. Data are mean ± SEM. N = 7. (e) Representative immunofluorescent images of MAP2, NeuN, and To-pro-3 staining in the CL, IL perilesion and IL core areas in the cortex and striatum of the oil- and Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f mice at 14 days post-tMCAO. (f) Quantitative analysis of MAP2 intensity in the CL, IL perilesion and IL core areas in the cortex and striatum of the oil- and Tam-treated Cx3cr1-Cre ER ; Nhe1 f/f mice at 14 days post-tMCAO. N = 3. * p

    Article Snippet: The sections were incubated with blocking solution (10% normal goat serum and 0.3% Triton X-100 in PBS) for 1 hr at room temperature and were then incubated with mouse monoclonal anti-GFAP (1:200, Cell Signaling Technology) and rabbit polyclonal anti-IBA1 (1:200, Wako, Italy) antibodies, or mouse monoclonal anti-MAP2 (1:200, EMD Millipore) and rabbit monoclonal anti-NeuN (1:200, Abcam) antibodies, or mouse monoclonal anti-APC (1:100, EMD Millipore) and rabbit monoclonal anti-MBP (1:100, Abcam) antibodies in the blocking solution for overnight at 4°C.

    Techniques: Mouse Assay, Imaging, Ex Vivo, Staining

    Immunohistochemical and immunofluorescent double-staining of GBM tissue samples revealed strong P-AXL expression in CD31 positive endothelial cells ( A., B ., D .; arrowheads), no colocalization with aSMA ( B ., C ., D .), and partial colabeling with PDGFR-ß positive pericytes ( E .; arrowheads) in microvascular proliferation. Glioma cells adjacent to microvascular proliferation (dashed lines) showed strong immunopositivity for PDGFR-ß ( E .; arrows). P-AXL was further expressed by neoplastic glioma cells as noted by colabeling with GFAP ( F .- H .; arrowheads), MAP2 ( I .- K .; arrowheads), Nestin ( L .- N .; arrowheads), and ZEB1 O .- Q .; arrowheads). (Scale bar: 20 μm A ., 50 μm B .- Q .).

    Journal: Oncotarget

    Article Title: Phospho-AXL is widely expressed in glioblastoma and associated with significant shorter overall survival

    doi: 10.18632/oncotarget.18468

    Figure Lengend Snippet: Immunohistochemical and immunofluorescent double-staining of GBM tissue samples revealed strong P-AXL expression in CD31 positive endothelial cells ( A., B ., D .; arrowheads), no colocalization with aSMA ( B ., C ., D .), and partial colabeling with PDGFR-ß positive pericytes ( E .; arrowheads) in microvascular proliferation. Glioma cells adjacent to microvascular proliferation (dashed lines) showed strong immunopositivity for PDGFR-ß ( E .; arrows). P-AXL was further expressed by neoplastic glioma cells as noted by colabeling with GFAP ( F .- H .; arrowheads), MAP2 ( I .- K .; arrowheads), Nestin ( L .- N .; arrowheads), and ZEB1 O .- Q .; arrowheads). (Scale bar: 20 μm A ., 50 μm B .- Q .).

    Article Snippet: The following primary and secondary antibodies were used: polyclonal rabbit anti-GFAP (1:2000, Dako), monoclonal mouse anti-CD31 (1:100, clone JC70A, Dako), monoclonal mouse anti-MAP2 (1:10000, clone HM-2, Sigma Aldrich), monoclonal mouse anti-α-smooth-muscle actin (aSMA, 1:200, clone 1A4, Dako), monoclonal mouse anti-Nestin (1:200, clone 10C2, Millipore), polyclonal rabbit anti-PDGFR-ß (1:10, Santa Cruz), polyclonal rabbit anti-ZEB1 (1:300, Sigma Aldrich), polyclonal anti-Fibronectin (1:1000, Sigma Aldrich), monoclonal mouse anti-phospho-AXL (pTyr779) (1:50, clone 713610, R & D Systems), polyclonal rabbit anti-phospho-AXL (pTyr691) (1:50, Sigma Aldrich), monoclonal rabbit anti-AXL C89E7 (1:100, Cell Signaling), FITC-conjugated donkey anti-rabbit IgG (1:1000, Dianova, 711-095-152), Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1:1000, Dianova, 111-545-003), Alexa Fluor® 488-conjugated goat anti-mouse IgG (1:1000, Dianova, 115-545-003), Cy3-conjugated goat anti-mouse IgG (1:1000, Dianova, 115-165-003), Cy3-conjugated goat anti-rabbit IgG (1:1000, Dianova, 111-165-003), and Cy3-conjugated donkey anti-mouse IgG (1:1000, Dianova, 115-165-146).

    Techniques: Immunohistochemistry, Double Staining, Expressing

    Synaptotoxic effects of amyloid oligomers in primary neuronal cultures. a Primary hippocampal neuronal cultures exposed to Aβ40, Aβ42 or vehicle treated (Ctrl). The cultures were fixed and stained for synaptic marker Synapsin I (green) and neuronal marker MAP2 (red). Scale bar corresponds to 20 μm. b Mean fluorescent intensities of synapsin I staining were divided by mean fluorescent intensities of MAP2 staining for hippocampal cultures treated with Aβ40, Aβ42 or vehicle treated (Ctrl). Average results from three independent experiments are shown. Values are shown as mean ± SEM. *: p

    Journal: Molecular Neurodegeneration

    Article Title: STIM2 protects hippocampal mushroom spines from amyloid synaptotoxicity

    doi: 10.1186/s13024-015-0034-7

    Figure Lengend Snippet: Synaptotoxic effects of amyloid oligomers in primary neuronal cultures. a Primary hippocampal neuronal cultures exposed to Aβ40, Aβ42 or vehicle treated (Ctrl). The cultures were fixed and stained for synaptic marker Synapsin I (green) and neuronal marker MAP2 (red). Scale bar corresponds to 20 μm. b Mean fluorescent intensities of synapsin I staining were divided by mean fluorescent intensities of MAP2 staining for hippocampal cultures treated with Aβ40, Aβ42 or vehicle treated (Ctrl). Average results from three independent experiments are shown. Values are shown as mean ± SEM. *: p

    Article Snippet: Primary antibodies anti-map2 mAb (1:1000, Chemicon, mAB378) and anti-synapsin I (1:1000, Chemicon, Temecula, CA) were diluted in 2.5 % BSA in PBS with 0.025 % Tween-20 and incubated at room temperature for 3 h. After three times wash, the hippocampal cultures were incubated in 2.5 % BSA in PBS solution with the secondary antibody (1:1000, Alexa Fluor 488 or 594, Invitrogen) for 1.5 h at room temperature and visualized by a confocal microscope (Thorlabs, Russia).

    Techniques: Staining, Marker

    GABA immunoreactivity in cortical cultures. A , MAP2-positive neurons from cortical cultures after 7 d in vitro , viewed with fluorescein filters. B , Same field of view as in A , using rhodamine filters to show GABA-positive neurons. C , GABA-positive multipolar

    Journal: The Journal of Neuroscience

    Article Title: Brain-Derived Neurotrophic Factor Mediates the Activity-Dependent Regulation of Inhibition in Neocortical Cultures

    doi: 10.1523/JNEUROSCI.17-12-04527.1997

    Figure Lengend Snippet: GABA immunoreactivity in cortical cultures. A , MAP2-positive neurons from cortical cultures after 7 d in vitro , viewed with fluorescein filters. B , Same field of view as in A , using rhodamine filters to show GABA-positive neurons. C , GABA-positive multipolar

    Article Snippet: To double-label cultures against both GABA and MAP2, cultures were incubated concurrently in a rabbit anti-GABA polyclonal antiserum (Sigma, 1:800) and a mouse monoclonal anti-MAP2 antibody (Sigma, 1:500).

    Techniques: In Vitro

    Deletion of Apoa1 and Apoe significantly affect dendritic morphology in CA1 but not in CA2 region of hippocampus. MAP2 and H3342 staining were used for dendritic tree reconstruction in the hippocampal CA, and CA2 regions of APP/WT, APP/Abca1 ko and APP/DKO

    Journal: Brain

    Article Title: Opposing effects of Apoe/Apoa1 double deletion on amyloid-β pathology and cognitive performance in APP mice

    doi: 10.1093/brain/awv293

    Figure Lengend Snippet: Deletion of Apoa1 and Apoe significantly affect dendritic morphology in CA1 but not in CA2 region of hippocampus. MAP2 and H3342 staining were used for dendritic tree reconstruction in the hippocampal CA, and CA2 regions of APP/WT, APP/Abca1 ko and APP/DKO

    Article Snippet: The sections were incubated for 72 h at 4°C with MAP2 primary antibody (1:2000; #MAB378, Millipore) and washed with PBS.

    Techniques: Staining

    Repetitive and locomotor behavior of NOS2.KO and WT, and effect of siNOS2 in PC12 cells MAP2 expression. Reduced total activity (A) and increased percentage of repetitive behaviors (B) in NOS2KO females compared to WT ( n = 6–7/group). Effect of clomipramine (10 mg/kg in the drinking water) on total activity (C) and repetitive behaviors (D) in NOS2KO females ( n = 5–6/group). (E) the silencing of NOS2 compromises the expression of MAP2 in KCl-induced differentiation of PC12 cells ( n = 4–5/group). ∗ p

    Journal: PeerJ

    Article Title: Inducible nitric oxide synthase (NOS2) knockout mice as a model of trichotillomania

    doi: 10.7717/peerj.4635

    Figure Lengend Snippet: Repetitive and locomotor behavior of NOS2.KO and WT, and effect of siNOS2 in PC12 cells MAP2 expression. Reduced total activity (A) and increased percentage of repetitive behaviors (B) in NOS2KO females compared to WT ( n = 6–7/group). Effect of clomipramine (10 mg/kg in the drinking water) on total activity (C) and repetitive behaviors (D) in NOS2KO females ( n = 5–6/group). (E) the silencing of NOS2 compromises the expression of MAP2 in KCl-induced differentiation of PC12 cells ( n = 4–5/group). ∗ p

    Article Snippet: Following blockade with 1% BSA in PBST, antibody against MAP2 (#AB5622; Millipore, Darmstadt, Germany) was incubated overnight at 4 °C.

    Techniques: Expressing, Activity Assay

    Neural differentiation of BMSCs on the 6 th day of in vitro transfection. (A, B) Immunofluorescence for the neuronal markers β-III tubulin (A1 and A3; red) and MAP-2 (B1 and B3; green) in BMSCs transfected with pLVX-EN-rno-miR124 (miR-124 + ) after 6 days of in vitro differentiation. Scale bars: 100 μm. BMSCs developed dendrites and neurites, similar to neurons (A2, A4, B2, B4). Scale bars: 100 μm. (C) Western blotting was used to detect the expression of synaptophysin (38 kDa). BMSCs: Bone marrow-derived mesenchymal stem cells; MAP-2: microtubule-associated protein-2.

    Journal: Neural Regeneration Research

    Article Title: Overexpression of microRNA-124 promotes the neuronal differentiation of bone marrow-derived mesenchymal stem cells

    doi: 10.4103/1673-5374.135333

    Figure Lengend Snippet: Neural differentiation of BMSCs on the 6 th day of in vitro transfection. (A, B) Immunofluorescence for the neuronal markers β-III tubulin (A1 and A3; red) and MAP-2 (B1 and B3; green) in BMSCs transfected with pLVX-EN-rno-miR124 (miR-124 + ) after 6 days of in vitro differentiation. Scale bars: 100 μm. BMSCs developed dendrites and neurites, similar to neurons (A2, A4, B2, B4). Scale bars: 100 μm. (C) Western blotting was used to detect the expression of synaptophysin (38 kDa). BMSCs: Bone marrow-derived mesenchymal stem cells; MAP-2: microtubule-associated protein-2.

    Article Snippet: The membranes were blocked with 5% skimmed milk powder in TBST (10 mmol/L Tris-HCl, pH 7.5, 150 mmol/L NaCl, 0.05% Tween-20) and incubated overnight with mouse anti-β-III tubulin, anti-MAP-2 (1:1,000, Abgent Biotechnology, San Diego, CA, USA), anti-synaptophysin or anti-β-actin (1:3,000, Abgent Biotechnology) antibody at 4°C.

    Techniques: In Vitro, Transfection, Immunofluorescence, Western Blot, Expressing, Derivative Assay

    CD47 is the presynaptic receptor for SIRPα-mediated presynaptic maturation (a,b) Hippocampal neurons were stained for CD47 together with neurofilament, MAP2 or SIRPα. CD47 puncta are abundant in neurofilament-positive axons and not in MAP2-positve dendrites (a) and co-localized with SIRPα (b). Reproduced five times. (c) Hippocampal neurons prepared from CD47KO mice and control littermates were treated with sSIRPα and stained for VGLUT1. sSIRPα does not increase the number and size of VGLUT1 puncta in CD47KO neurons. n = 80/93/102/108 fields from 4 cultures. (d) HEK cells expressing SIRPα, neuroligin1, or control HEK cells (labeled with GFP) were co-cultured with hippocampal neurons prepared from WT or CD47KO mice for 2 days and stained for synapsin. HEK cells expressing SIRPα induce presynaptic differentiation in co-cultured WT hippocampal neurons, but fail to do so in co-cultured CD47KO neurons. HEK cells expressing neuroligin1 induce presynaptic differentiation in both WT and CD47KO neurons. Graphs show quantification of synapsin puncta number and size. Data are from 38/34/34/35/42/40 fields from 5 cultures. (e) Presynaptic expression of CD47 restores responsiveness to sSIRPα in CD47KO neurons. Hippocampal neurons prepared from WT or CD47KO mice were transfected with the synaptophysin-YFP plasmid or with the synaptophysin-YFP and CD47 plasmids. Cultures were treated with sSIRPα, and presynaptic differentiation of transfected neurons was assessed by synaptophysin-YFP clustering. sSIRPα does not increase the number and size of synaptophysin-YFP puncta in CD47KO neurons, but introduction of CD47 restores responsiveness to sSIRPα. n = 33/37/33/35/37/36 neurites from 3 cultures. (f) Postsynaptic expression of SIRPα rescues presynaptic defects in SIRPαKO neurons. Hippocampal neurons prepared from conditional SIRPαKO mice were transfected with an mCherry plasmid (control), Cre and mCherry plasmids (knockout) or Cre, mCherry and SIRPα plasmids (rescue) and stained for VGLUT1. Both the number and size of VGLUT1 puncta on mCherry-expressing dendrites were decreased in SIRPαKO relative to control, but postsynaptic expression of SIRPα rescues the defects. n = 41/33/36 neurites from 3 cultures. Student’s t-test (c) or ANOVA followed by Tukey test (d–f). Scale bar, 5 μm (c), 10 μm (all others).

    Journal: Nature neuroscience

    Article Title: Synapse Maturation by Activity-Dependent Ectodomain Shedding of SIRP?

    doi: 10.1038/nn.3516

    Figure Lengend Snippet: CD47 is the presynaptic receptor for SIRPα-mediated presynaptic maturation (a,b) Hippocampal neurons were stained for CD47 together with neurofilament, MAP2 or SIRPα. CD47 puncta are abundant in neurofilament-positive axons and not in MAP2-positve dendrites (a) and co-localized with SIRPα (b). Reproduced five times. (c) Hippocampal neurons prepared from CD47KO mice and control littermates were treated with sSIRPα and stained for VGLUT1. sSIRPα does not increase the number and size of VGLUT1 puncta in CD47KO neurons. n = 80/93/102/108 fields from 4 cultures. (d) HEK cells expressing SIRPα, neuroligin1, or control HEK cells (labeled with GFP) were co-cultured with hippocampal neurons prepared from WT or CD47KO mice for 2 days and stained for synapsin. HEK cells expressing SIRPα induce presynaptic differentiation in co-cultured WT hippocampal neurons, but fail to do so in co-cultured CD47KO neurons. HEK cells expressing neuroligin1 induce presynaptic differentiation in both WT and CD47KO neurons. Graphs show quantification of synapsin puncta number and size. Data are from 38/34/34/35/42/40 fields from 5 cultures. (e) Presynaptic expression of CD47 restores responsiveness to sSIRPα in CD47KO neurons. Hippocampal neurons prepared from WT or CD47KO mice were transfected with the synaptophysin-YFP plasmid or with the synaptophysin-YFP and CD47 plasmids. Cultures were treated with sSIRPα, and presynaptic differentiation of transfected neurons was assessed by synaptophysin-YFP clustering. sSIRPα does not increase the number and size of synaptophysin-YFP puncta in CD47KO neurons, but introduction of CD47 restores responsiveness to sSIRPα. n = 33/37/33/35/37/36 neurites from 3 cultures. (f) Postsynaptic expression of SIRPα rescues presynaptic defects in SIRPαKO neurons. Hippocampal neurons prepared from conditional SIRPαKO mice were transfected with an mCherry plasmid (control), Cre and mCherry plasmids (knockout) or Cre, mCherry and SIRPα plasmids (rescue) and stained for VGLUT1. Both the number and size of VGLUT1 puncta on mCherry-expressing dendrites were decreased in SIRPαKO relative to control, but postsynaptic expression of SIRPα rescues the defects. n = 41/33/36 neurites from 3 cultures. Student’s t-test (c) or ANOVA followed by Tukey test (d–f). Scale bar, 5 μm (c), 10 μm (all others).

    Article Snippet: Dilutions and sources of antibodies are: anti-VGLUT1 (1:5,000; Millipore; AB5905), anti-PSD95 (1:250; NeuroMab; 75-028), anti-VGAT (1:1,500; Synaptic Systems; 131003), anti-MAP2 (1:3,000; Sigma-Aldrich; M4403), anti-neurofilament (1:1,000; Covance; SMI-312), anti-bassoon (1:500; Enzo Life Sciences; ADI-VAM-PS003), anti-GFP (1:1,000; Millipore; AB16901), anti-GFAP (1:500; Synaptic Systems; 173002), anti-NeuN (1:500; Millipore; MAB377), anti-calbindin (1:500; Sigma; C9848), anti-synapsin (1:2000; a kind gift from P. Greengard and A. Nairn, Rockefeller University), antibody Py (1:50; a kind gift from M. Webb and P.L.

    Techniques: Staining, Mouse Assay, Expressing, Labeling, Cell Culture, Transfection, Plasmid Preparation, Knock-Out

    FGF22 and SIRPα promote the early or late stage of glutamatergic presynaptic differentiation (a) In situ hybridization for Sirpα in the hippocampus during synapse formation (positive signals in black). Sirpα mRNA is highly expressed at P21, the time for synapse maturation, but not at P8, the time for initial synapse differentiation. Reproduced three times. (b) Western blotting for the SIRPα protein (tubulin as control) in the hippocampus. The amount of SIRPα significantly increases from P8 to P21. Full-length blots are presented in Supplementary Figure 10 . (c,d) Hippocampal cultures at DIV11 were stained with the antibodies indicated. (c) SIRPα proteins are abundant on MAP2-positive dendrites but not on neurofilament (NF)-positive axons.(d) SIRPα is concentrated at VGLUT1-postive glutamatergic synapses but not at VGAT-positive GABAergic synapses. Reproduced five times. (e) HEK cells expressing SIRPα, neuroligin1 (NLGN1), or control HEK cells (labeled with GFP) were co-cultured with hippocampal neurons for 2 days and stained for synapsin. The synapsin puncta formed on HEK cells expressing SIRPα are significantly more dense and larger than those formed on control HEK cells and are comparable to the ones on HEK cells expressing NLGN1. Data are from 24/23/22 fields from 5 cultures. (f,g) Recombinant soluble SIRPα (sSIRPα) was applied to hippocampal cultures from DIV1–11.(f) sSIRPα treatment significantly increases the number (x 1,000 puncta per mm 2 ) and size of VGLUT1 puncta as compared to PBS control (n = 57 fields from 5 cultures). (g) Representative traces and summary data of whole-cell recordings of mEPSCs from control and sSIRPα-treated hippocampal neurons. mEPSC frequency, but not amplitude, increases by sSIRPα treatment. (n = 57/63 cells from 5 cultures). (h) Schematic timeline of the experiment shown in (i,j). Cultured hippocampal cells were treated with FGF22 or sSIRPα from DIV1–4 (beginning of synapse formation), DIV4–8 (middle of synapse formation), or DIV8–11 (ending of synapse formation). All cultures were fixed on DIV11. (i) Staining of hippocampal cultures for VGLUT1 after treatment with FGF22 or sSIRPα as shown in (h). (j) Numbers and sizes of VGLUT1 puncta after treatment on DIV1–4, DIV4–8, or DIV8–11. FGF22 treatment is most effective at increasing VGLUT1 clustering when incubated from DIV1–4, while sSIRPα is most effective when incubated from DIV8–11. Data are shown as percentage of PBS control and from 32/43/40/34/27/26 fields from 5 cultures. (k) sSIRPα or FGF22 was applied to hippocampal cultures prepared from WT or FGF22KO mice, and the cultures were stained for VGLUT1 and Py, which labels dendrites of CA3 pyramidal neurons. Fewer and smaller VGLUT1 puncta were on CA3 neurons in FGF22KO cultures relative to WT cultures; the defects were rescued by the application of FGF22, but not by sSIRPα. n = 19/23/25/25/17/23 neurites from 3 cultures. Bars in the graphs are mean ± SEM. *p

    Journal: Nature neuroscience

    Article Title: Synapse Maturation by Activity-Dependent Ectodomain Shedding of SIRP?

    doi: 10.1038/nn.3516

    Figure Lengend Snippet: FGF22 and SIRPα promote the early or late stage of glutamatergic presynaptic differentiation (a) In situ hybridization for Sirpα in the hippocampus during synapse formation (positive signals in black). Sirpα mRNA is highly expressed at P21, the time for synapse maturation, but not at P8, the time for initial synapse differentiation. Reproduced three times. (b) Western blotting for the SIRPα protein (tubulin as control) in the hippocampus. The amount of SIRPα significantly increases from P8 to P21. Full-length blots are presented in Supplementary Figure 10 . (c,d) Hippocampal cultures at DIV11 were stained with the antibodies indicated. (c) SIRPα proteins are abundant on MAP2-positive dendrites but not on neurofilament (NF)-positive axons.(d) SIRPα is concentrated at VGLUT1-postive glutamatergic synapses but not at VGAT-positive GABAergic synapses. Reproduced five times. (e) HEK cells expressing SIRPα, neuroligin1 (NLGN1), or control HEK cells (labeled with GFP) were co-cultured with hippocampal neurons for 2 days and stained for synapsin. The synapsin puncta formed on HEK cells expressing SIRPα are significantly more dense and larger than those formed on control HEK cells and are comparable to the ones on HEK cells expressing NLGN1. Data are from 24/23/22 fields from 5 cultures. (f,g) Recombinant soluble SIRPα (sSIRPα) was applied to hippocampal cultures from DIV1–11.(f) sSIRPα treatment significantly increases the number (x 1,000 puncta per mm 2 ) and size of VGLUT1 puncta as compared to PBS control (n = 57 fields from 5 cultures). (g) Representative traces and summary data of whole-cell recordings of mEPSCs from control and sSIRPα-treated hippocampal neurons. mEPSC frequency, but not amplitude, increases by sSIRPα treatment. (n = 57/63 cells from 5 cultures). (h) Schematic timeline of the experiment shown in (i,j). Cultured hippocampal cells were treated with FGF22 or sSIRPα from DIV1–4 (beginning of synapse formation), DIV4–8 (middle of synapse formation), or DIV8–11 (ending of synapse formation). All cultures were fixed on DIV11. (i) Staining of hippocampal cultures for VGLUT1 after treatment with FGF22 or sSIRPα as shown in (h). (j) Numbers and sizes of VGLUT1 puncta after treatment on DIV1–4, DIV4–8, or DIV8–11. FGF22 treatment is most effective at increasing VGLUT1 clustering when incubated from DIV1–4, while sSIRPα is most effective when incubated from DIV8–11. Data are shown as percentage of PBS control and from 32/43/40/34/27/26 fields from 5 cultures. (k) sSIRPα or FGF22 was applied to hippocampal cultures prepared from WT or FGF22KO mice, and the cultures were stained for VGLUT1 and Py, which labels dendrites of CA3 pyramidal neurons. Fewer and smaller VGLUT1 puncta were on CA3 neurons in FGF22KO cultures relative to WT cultures; the defects were rescued by the application of FGF22, but not by sSIRPα. n = 19/23/25/25/17/23 neurites from 3 cultures. Bars in the graphs are mean ± SEM. *p

    Article Snippet: Dilutions and sources of antibodies are: anti-VGLUT1 (1:5,000; Millipore; AB5905), anti-PSD95 (1:250; NeuroMab; 75-028), anti-VGAT (1:1,500; Synaptic Systems; 131003), anti-MAP2 (1:3,000; Sigma-Aldrich; M4403), anti-neurofilament (1:1,000; Covance; SMI-312), anti-bassoon (1:500; Enzo Life Sciences; ADI-VAM-PS003), anti-GFP (1:1,000; Millipore; AB16901), anti-GFAP (1:500; Synaptic Systems; 173002), anti-NeuN (1:500; Millipore; MAB377), anti-calbindin (1:500; Sigma; C9848), anti-synapsin (1:2000; a kind gift from P. Greengard and A. Nairn, Rockefeller University), antibody Py (1:50; a kind gift from M. Webb and P.L.

    Techniques: In Situ Hybridization, Western Blot, Staining, Expressing, Labeling, Cell Culture, Recombinant, Incubation, Mouse Assay

    Sodium hydrosulfide treatment (250 μM) could significantly repair OGD-induced injuries of neurite sprouting and outgrowth in the young and aged hippocampal neurons in 7 DIV. (A) Typical hippocampal neurons with extending neurites in the cultures, stained for MAP-2. Scale bar = 40 μm. The neurite sprouting and outgrowth were analyzed by the number of primary dendrites per cell (B) , the number of dendritic end tips (C) , and the average neurite length (D) . Data were presented by the mean ± SD. ∗ p

    Journal: Frontiers in Cellular Neuroscience

    Article Title: NaHS Protects against the Impairments Induced by Oxygen-Glucose Deprivation in Different Ages of Primary Hippocampal Neurons

    doi: 10.3389/fncel.2017.00067

    Figure Lengend Snippet: Sodium hydrosulfide treatment (250 μM) could significantly repair OGD-induced injuries of neurite sprouting and outgrowth in the young and aged hippocampal neurons in 7 DIV. (A) Typical hippocampal neurons with extending neurites in the cultures, stained for MAP-2. Scale bar = 40 μm. The neurite sprouting and outgrowth were analyzed by the number of primary dendrites per cell (B) , the number of dendritic end tips (C) , and the average neurite length (D) . Data were presented by the mean ± SD. ∗ p

    Article Snippet: Briefly, cells were fixed with 4% paraformaldehyde for 25 min and washed two times with PBS for 8 min. After permeabilization (0.1% Triton X-100, 15 min) and blocking (5% bovine serum albumin, 20 min), cells were stained with antibody against MAP-2 (Abcam, Cambridge, MA, USA) at room temperature for 2 h, then washed and incubated with the secondary antibodies for 1 h. Cells were then imaged by a scanning confocal microscope (Carl Zeiss, Germany).

    Techniques: Staining

    HIV + sup ± morphine-mediated neurite damage. Cells were fixed at specific intervals after treatment and labeled for MAP-2 (green) and TUNEL (red). ( A ) Digital images of neuronal cultures at 72 h after treatment; scale bar = 40 µm. ( B ) The ‘Sholl score’ was assessed only for TUNEL(-) neurons in the digital images and converted into neurite length in µm via a micrometer-scale calibration. The findings were reported as average total neurite length per neuron (µm) ± SEM. Significance was analyzed by one-way ANOVA and Duncan's post hoc test from n = 4 separate experiments. At all time-points and in both culture systems, all groups exposed to HIV + sup showed significantly reduced neurite length (* p

    Journal: PLoS ONE

    Article Title: Morphine Enhances HIV-1SF162-Mediated Neuron Death and Delays Recovery of Injured Neurites

    doi: 10.1371/journal.pone.0100196

    Figure Lengend Snippet: HIV + sup ± morphine-mediated neurite damage. Cells were fixed at specific intervals after treatment and labeled for MAP-2 (green) and TUNEL (red). ( A ) Digital images of neuronal cultures at 72 h after treatment; scale bar = 40 µm. ( B ) The ‘Sholl score’ was assessed only for TUNEL(-) neurons in the digital images and converted into neurite length in µm via a micrometer-scale calibration. The findings were reported as average total neurite length per neuron (µm) ± SEM. Significance was analyzed by one-way ANOVA and Duncan's post hoc test from n = 4 separate experiments. At all time-points and in both culture systems, all groups exposed to HIV + sup showed significantly reduced neurite length (* p

    Article Snippet: Culture purity was determined by immunocytochemistry using anti-MAP-2 antibody (Abcam, Cambridge, MA; ab32454) and found to be > 80% neurons.

    Techniques: Labeling, TUNEL Assay

    Characterization of human glioblastoma stem cells. Representative images of GSC (derived from GBM6) spheroids (A) and monolayer on Matrigel (B) cultured in stem cell medium supplemented with EGF and bFGF and differentiated cells (C) obtained by switching culture conditions from growth factor-deprived to FBS-containing medium. Left panels reproduce phase contrast images at different magnification (10× and 20×). Right panels depict immunofluorescence for stem cell markers OLIG2, nestin, and Oct4 (A,B) , and the glia (GFAP) and neuron (MAP-2) differentiation markers (C) . GFAP and MAP-2 expression is depicted from the same microscope field, showing in most cells the co-expression of both markers. Similar results were obtained analyzing all the GSC cultures that enter the study.

    Journal: Frontiers in Pharmacology

    Article Title: Inhibition of Chloride Intracellular Channel 1 (CLIC1) as Biguanide Class-Effect to Impair Human Glioblastoma Stem Cell Viability

    doi: 10.3389/fphar.2018.00899

    Figure Lengend Snippet: Characterization of human glioblastoma stem cells. Representative images of GSC (derived from GBM6) spheroids (A) and monolayer on Matrigel (B) cultured in stem cell medium supplemented with EGF and bFGF and differentiated cells (C) obtained by switching culture conditions from growth factor-deprived to FBS-containing medium. Left panels reproduce phase contrast images at different magnification (10× and 20×). Right panels depict immunofluorescence for stem cell markers OLIG2, nestin, and Oct4 (A,B) , and the glia (GFAP) and neuron (MAP-2) differentiation markers (C) . GFAP and MAP-2 expression is depicted from the same microscope field, showing in most cells the co-expression of both markers. Similar results were obtained analyzing all the GSC cultures that enter the study.

    Article Snippet: 356.1 (Santa Cruz Biotechnology, Dallas, TX, United States) for immunofluorescence and Western blot experiments, respectively; anti-GFAP, anti-nestin, anti-Oct4, anti-Sox2, and anti-MAP-2 (Abcam, Cambridge, United Kingdom); anti-Olig2 and anti-Sox2, (Merck Vimodrone, Italy); anti-mouse and anti-rabbit Alexa Fluor 488 and 568 secondary antibodies (Thermo Fisher Scientific, Carlsbad, CA, United States).

    Techniques: Derivative Assay, Cell Culture, Immunofluorescence, Expressing, Microscopy