rabbit anti-gfap Search Results


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  • 98
    Thermo Fisher rabbit anti glial fibrillary acidic protein
    Up-regulation of astrocyte differentiation of hippocampal neural stem cells derived from both newborn and adult rats after exposure to Pb 2+ . Through the fluorescence microscopy using double labeling of 4’,6-diamidino-2-phenylindole (blue) and <t>glial</t> <t>fibrillary</t> <t>acidic</t> <t>protein</t> (red), the astrocyte differentiation of newborn rat hippocampal neural stem cells after exposure to Pb 2+ is shown in (A, 0 μM Pb 2+ ) and (C, 100 μM Pb 2+ ) respectively. Furthermore, the astrocyte commitment of adult rat hippocampal neural stem cells after exposure to Pb 2+ is shown in (B, 0 μM Pb 2+ ) and (D, 100 μM Pb 2+ ) respectively. Scale bars: 50 μm for A-D. The newborn rat hippocampal neural stem cells were more sensitive to Pb 2+ exposure in astrocyte differentiation indicated by a significant increase in the percentage of mature astrocytes even at a relatively low concentration (10 μM) of lead acetate ( b P
    Rabbit Anti Glial Fibrillary Acidic Protein, supplied by Thermo Fisher, used in various techniques. Bioz Stars score: 98/100, based on 32 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Millipore rabbit anti gfap
    Minocycline has anti-inflammatory effects in ts 1-infected mice brainstem and cultured C1 astrocytes. (A). Cell lysates or nuclear extracts from uninfected, ts 1-only and ts 1-mino C1 astrocytes are probed with anti-lκBα, NF-κB p65 and <t>COX-2.</t> (B). Mice from two groups were sacrificed at 30 dpi. Paraffin sections of the brainstem used for immunohistochemistry (IHC) assay. These sections were incubated with <t>anti-GFAP</t> (brown) and anti-COX-2 antibodies as specified in Experimental Procedures. Strong COX-2 immunoreactivity (red) is evident in astrocyte (arrow) of ts 1-only mouse (left panel), but weak in ts 1-mino mouse (right panel). Magnifications: 40X. (C). Expression of CD68, which marks activated microglia and macrophages, was examined by immunofluorescence staining using brainstem sections from uninfected, ts 1-only and ts 1-mino mice. Magnifications: 20X.
    Rabbit Anti Gfap, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 2194 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Agilent technologies rabbit anti gfap
    In vitro phenotypes of hNPCs differentiated in the neuroblast assay (NBA), followed by enrichment of the immature neuron subpopulation using MACS. ( A ) Overall experimental design. hNPC neurospheres were dissociated into single cells and plated in the NBA, generating a mixed population of neurons (N) and astrocytes ( A ). The neurons were then isolated from the astrocytes via MACS. Lastly, phenotyping was performed on the cells during the three major phases of the experiment. ( B–D ) Fluorescent images of immunocytochemical staining show hNPCs significantly lose expression of the proliferation marker <t>Ki67</t> during the NBA. Enrichment of the neuron subgroup after the NBA, using MACS, reduces Ki67 expression still further (scale bar = 20 μm) (49,6-diamidino-2-phenylindole (DAPI), blue). ( E–G ) hNPCs significantly gain expression of the immature neuron marker DCX during the NBA. Enrichment of the neuron subgroup after the NBA yields an even higher percentage of DCX+ cells (scale bar = 20 μm) (49,6-diamidino-2-phenylindole (DAPI), blue). ( H–J ) Expression of the astrocyte marker <t>GFAP</t> increases slightly during differentiation, but MACS purification removes virtually all the percentage of GFAP+ cells (scale bar = 20 μm) (49,6-diamidino-2-phenylindole (DAPI), blue). ( K ) Quantification of immunolabeled cells indicates the differentiation of hNPCs into immature neurons during the NBA, and the subsequent purification of neurons made possible by MACS. Means + s.e.m. * P
    Rabbit Anti Gfap, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 94/100, based on 6688 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    94
    Abcam rabbit anti gfap
    Fibrinogen-induced SVZ astrocytes add to cortical scar. a Representation of the different lesion regions quantified by immunoreactivity. b Fibrinogen deposition (red) and <t>GFAP</t> + astrocytes in the brain of fibrinogen-depleted mice (Ancrod) compared to controls (NaCl) at 6 days after PT (left). Yellow dotted line delineates the lesion area. The white boxes indicate the enlargement of the lesion region quantified for GFAP (right, top) and CSPGs (right, bottom) immunoreactivities. Quantifications of GFAP and <t>CSPG</t> immunoreactivities in the different lesion regions ( n = 4 mice, mean ± s.e.m, one-way ANOVA and Bonferroni’s multiple comparisons test, * P
    Rabbit Anti Gfap, supplied by Abcam, used in various techniques. Bioz Stars score: 94/100, based on 1338 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    99
    Millipore anti glial fibrillary acidic protein antibody
    Single neurons were grown on microislands consisting of purified astrocytes. Neurons were identified by labeling with the fluorescein-conjugated C fragment of tetanus toxin (CFITC; Left ), and <t>anti-glial</t> <t>fibrillary</t> <t>acidic</t> <t>protein</t> (GFAP) <t>antibody</t> was used to identify astrocytes ( Right ).
    Anti Glial Fibrillary Acidic Protein Antibody, supplied by Millipore, used in various techniques. Bioz Stars score: 99/100, based on 1177 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    92
    Agilent technologies rabbit polyclonal anti gfap
    Intraperitoneal injection of curcumin (200 μg/g BW) in P7 mice immediately post-HI has no effect on the levels of proliferation at 48 h. (A) Assessment of the number of BrdU+ cells in P7 naïve untreated animals or naïve and HI animals treated with curcumin or DMSO did not show differences between the groups in any of the regions. (naïve n = 4, naïve + curcumin n = 5, HI + DMSO n = 7, HI + curcumin n = 8). (B) Curcumin treatment in naïve or HI animals immediately after insult did not affect the percentage of IBA1 and BrdU double positive over BrdU+ cells per region and group. The percentage of proliferating IBA1+ cells was low (10–20% of the BrdU+ cells). (C) The percentage of <t>GFAP</t> and BrdU double positive over BrdU+ cells per region and group was not affected by curcumin treatment in naïve or HI animals. The percentage of proliferating GFAP+ cells was low (10–20% of the BrdU+ cells). (D) The percentage of NG2 and BrdU double positive over BrdU+ cells per region and group was not affected by curcumin treatment in naïve or HI animals. The percentage of proliferating NG2+ cells was low (10–20% of the BrdU+ cells). (E) The percentage of NeuN and BrdU double positive over BrdU+ cells per region and group was not affected by curcumin treatment in naïve or HI animals. The percentage of proliferating NeuN+ cells comprised 80–90% of the BrdU+ cells and was the highest of all four studied cell types. (F,J,N) Immunofluorescence for rat <t>polyclonal</t> anti-BrdU (F,N) in green superimposed on the rabbit polyclonal anti-IBA1 (J,N) in red, and nuclear DAPI fluorescence in blue (N) . Note the co-localization of BrdU and IBA1 (white arrows) and the lack of such co- localization (empty arrows). (G,K,O) Immunofluorescence for rat polyclonal anti-BrdU (G,O) in green superimposed on the rabbit polyclonal anti-GFAP (K,O) in red, and nuclear DAPI fluorescence in blue (O) . Note the co-localization of BrdU and GFAP (full arrows) and the lack of such co-localization (empty arrows). (H,L,P) Immunofluorescence for rat polyclonal anti-BrdU (H,P) in green superimposed on the guinea pig polyclonal anti-NG2 (L,P) in red, and nuclear DAPI fluorescence in blue (P) . Note the co-localization of BrdU and NG2 (full arrows) and the lack of such co-localization (empty arrows). (I,M,Q) Immunofluorescence for rat polyclonal anti-BrdU (I,Q) in green superimposed on the mouse monoclonal anti-NeuN (M,Q) in red, and nuclear DAPI fluorescence in blue (Q) . Note the high level of co-localization of BrdU and NeuN (full arrows) and the lack of such co-localization (empty arrows). Abbreviations: CTX, cerebral cortex; EC – external capsule; HIP – hippocampus; PYR, pyriform cortex; STR, striatum; THAL, thalamus. Scale bar: 126 μm.
    Rabbit Polyclonal Anti Gfap, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 92/100, based on 809 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam anti gfap antibody
    DGC activation during the course of status epilepticus. ( A ) TRAPing of DGCs at T0 + 60 to T0 + 300 min. ( B ) Co-localization of TRAPed neurons in the dentate gyrus with the neuronal marker protein <t>NeuN,</t> microglial marker protein Iba1 and astrocyte marker protein <t>GFAP.</t> The insets show boxed areas in the respective image. ( C ) The number of activated DGCs was plotted over time; n = 3 each for T0, T0 + 60, +120, +180, n = 4 for T0 + 240 and n = 2 for T0 + 300. The line depicts a linear regression fit of the data.
    Anti Gfap Antibody, supplied by Abcam, used in various techniques. Bioz Stars score: 99/100, based on 793 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    91
    Agilent technologies rabbit anti glial fibrillary acidic protein gfap
    PALS1 localizes with <t>CRB1</t> and MUPP1 to the subapical region adjacent to adherens junctions at the outer limiting membrane. Immunohistochemistry on 3-month-old retinas showing confocal images of staining for proteins associated with the Crumbs complex, PALS1, CRB1, and MUPP1, and for <t>GFAP,</t> a Müller glial cell marker. Staining for the Crumbs complex shows fragmented staining (arrows) of PALS1, CRB1, and MUPP1 in the shPals1 -Rx Cre retinas, which indicate the dislocalization of the Crumbs complex proteins at the SAR adjacent to adherens junctions at the OLM. GFAP staining shows increased reactive gliosis in the mutant retinas. Scale bar, 50 μm.
    Rabbit Anti Glial Fibrillary Acidic Protein Gfap, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 91/100, based on 493 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Abcam rabbit anti glial fibrillary acidic protein antibody
    TRPV4 activation was involved in <t>glial</t> activation and proinflammatory cytokines release. (A) Representative glial <t>fibrillary</t> <t>acidic</t> <t>protein</t> <t>antibody</t> (GFAP) and Iba-1 immunofluorescent images for the corpus callosum. Scale bar = 20 μm. (B,C) Quantitative analysis of GFAP- and Iba-1-positive cell numbers. The increased numbers of GFAP- and Iba-1-positive cells were reversed by the TRPV4 inhibitor RN-1734 ( n = 4). (D) Protein levels of tumor necrosis factor alpha (TNF-α) and interleukin 1beta (IL-1β) were detected by enzyme-linked immunosorbent assay (ELISA) ( n = 3). The data are expressed as the mean ± SEM. ** P
    Rabbit Anti Glial Fibrillary Acidic Protein Antibody, supplied by Abcam, 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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    Millipore rabbit anti glial fibrillary acidic protein gfap
    Confocal analysis of eEF1A-2/S1 during murine brain development. (A) One day after birth (P1), co-immunolocalization between <t>neurofilament</t> NF68 (red) and eEF1A-2/S1 (green) reveals absence of eEF1A-2/S1 in neurons. (B) Later in development at P14, co-immunolocalization reveals that both eEF1A-2/S1 (green) and NF68 (red) co-localize in neurons. However, eEF1A-2/S1 expression is weak. (C) The same co-localization (yellow) 20 days after birth (P20) reveals an increase in eEF1A-2/S1 abundance in the neuronal soma. (D) At P26, eEF1A-2/S1 (green) is clearly visible in the soma of neurons. (E) Twelve-month-old mouse brain shows intense co-localization (yellow) between EF1A-2/S1 and NF68 (red). (F) Co-immunolocalization of eEF1A-2/S1 (red) and <t>GFAP</t> (green) reveals that eEF1A-2/S1 is not expressed in astrocytes. Solid arrows indicate neuronal soma, open arrows indicate axons, and asterisks indicate nuclei.
    Rabbit Anti Glial Fibrillary Acidic Protein Gfap, supplied by Millipore, used in various techniques. Bioz Stars score: 94/100, based on 276 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Agilent technologies rabbit anti gfap pab
    Confocal analysis of eEF1A-2/S1 during murine brain development. (A) One day after birth (P1), co-immunolocalization between <t>neurofilament</t> NF68 (red) and eEF1A-2/S1 (green) reveals absence of eEF1A-2/S1 in neurons. (B) Later in development at P14, co-immunolocalization reveals that both eEF1A-2/S1 (green) and NF68 (red) co-localize in neurons. However, eEF1A-2/S1 expression is weak. (C) The same co-localization (yellow) 20 days after birth (P20) reveals an increase in eEF1A-2/S1 abundance in the neuronal soma. (D) At P26, eEF1A-2/S1 (green) is clearly visible in the soma of neurons. (E) Twelve-month-old mouse brain shows intense co-localization (yellow) between EF1A-2/S1 and NF68 (red). (F) Co-immunolocalization of eEF1A-2/S1 (red) and <t>GFAP</t> (green) reveals that eEF1A-2/S1 is not expressed in astrocytes. Solid arrows indicate neuronal soma, open arrows indicate axons, and asterisks indicate nuclei.
    Rabbit Anti Gfap Pab, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 88/100, based on 36 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Cell Signaling Technology Inc rabbit anti gfap
    Accumulation of newborn neurons. (A) The merged confocal images confirmed the presence of BrdU (red), <t>GFAP</t> (red), <t>DCX</t> (green), and Nestin (green) expressing cells in ipsilateral brain sections. Scale bar=20 μmol/L. (B) Quantitative data of each cell type. Means±SD. n =10. b P
    Rabbit Anti Gfap, supplied by Cell Signaling Technology Inc, used in various techniques. Bioz Stars score: 93/100, based on 140 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    88
    Agilent technologies rabbit anti cow gfap
    a Claudin-5 (×60 magnification) and IgG (×20 magnification) staining in ipsilateral and contralateral striatum for LPS and positive control AMPA-challenged animals. Claudin-5 staining revealed no differences between contralateral and ipsilateral sides in the LPS-challenged animals, while for AMPA-challenged animals, there was visible damage to vessels in the ipsilateral side and some disruption in the contralateral hemisphere. IgG appears normal in ipsilateral and contralateral sides of LPS animals but shows signs of leakage in the ipsilateral side of AMPA animals. b Staining for microglia (CD11b, red ) and astrocytes <t>(GFAP,</t> green ), merged at ×10 and ×20 ( white rectangles ) magnification; c NeuN staining at ×20 magnification in core and contralateral striatum
    Rabbit Anti Cow Gfap, supplied by Agilent technologies, used in various techniques. Bioz Stars score: 88/100, based on 144 PubMed citations. ZERO BIAS - scores, article reviews, protocol conditions and more
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    Image Search Results


    Up-regulation of astrocyte differentiation of hippocampal neural stem cells derived from both newborn and adult rats after exposure to Pb 2+ . Through the fluorescence microscopy using double labeling of 4’,6-diamidino-2-phenylindole (blue) and glial fibrillary acidic protein (red), the astrocyte differentiation of newborn rat hippocampal neural stem cells after exposure to Pb 2+ is shown in (A, 0 μM Pb 2+ ) and (C, 100 μM Pb 2+ ) respectively. Furthermore, the astrocyte commitment of adult rat hippocampal neural stem cells after exposure to Pb 2+ is shown in (B, 0 μM Pb 2+ ) and (D, 100 μM Pb 2+ ) respectively. Scale bars: 50 μm for A-D. The newborn rat hippocampal neural stem cells were more sensitive to Pb 2+ exposure in astrocyte differentiation indicated by a significant increase in the percentage of mature astrocytes even at a relatively low concentration (10 μM) of lead acetate ( b P

    Journal: Neural Regeneration Research

    Article Title: Are newborn rat-derived neural stem cells more sensitive to lead neurotoxicity? ★

    doi: 10.3969/j.issn.1673-5374.2013.07.001

    Figure Lengend Snippet: Up-regulation of astrocyte differentiation of hippocampal neural stem cells derived from both newborn and adult rats after exposure to Pb 2+ . Through the fluorescence microscopy using double labeling of 4’,6-diamidino-2-phenylindole (blue) and glial fibrillary acidic protein (red), the astrocyte differentiation of newborn rat hippocampal neural stem cells after exposure to Pb 2+ is shown in (A, 0 μM Pb 2+ ) and (C, 100 μM Pb 2+ ) respectively. Furthermore, the astrocyte commitment of adult rat hippocampal neural stem cells after exposure to Pb 2+ is shown in (B, 0 μM Pb 2+ ) and (D, 100 μM Pb 2+ ) respectively. Scale bars: 50 μm for A-D. The newborn rat hippocampal neural stem cells were more sensitive to Pb 2+ exposure in astrocyte differentiation indicated by a significant increase in the percentage of mature astrocytes even at a relatively low concentration (10 μM) of lead acetate ( b P

    Article Snippet: At the end of the incubation, the cells were rinsed three times with PBS and incubated with secondary antibodies [goat anti-rabbit Alexa Fluor® 488 for rabbit anti-microtubule-associated protein 2 (1:500 dilution in 1% bovine serum albumin/PBS), goat anti-rabbit Alexa Fluor® 647 for rabbit anti-glial fibrillary acidic protein (1:500 dilution in 1% bovine serum albumin/PBS) and goat anti-mouse Alexa Fluor® 488 for mouse anti-RIP (1:500 dilution in 1% bovine serum albumin/PBS); Life Technologies™ Inc.) at room temperature for 2 hours.

    Techniques: Derivative Assay, Fluorescence, Microscopy, Labeling, Concentration Assay

    Minocycline has anti-inflammatory effects in ts 1-infected mice brainstem and cultured C1 astrocytes. (A). Cell lysates or nuclear extracts from uninfected, ts 1-only and ts 1-mino C1 astrocytes are probed with anti-lκBα, NF-κB p65 and COX-2. (B). Mice from two groups were sacrificed at 30 dpi. Paraffin sections of the brainstem used for immunohistochemistry (IHC) assay. These sections were incubated with anti-GFAP (brown) and anti-COX-2 antibodies as specified in Experimental Procedures. Strong COX-2 immunoreactivity (red) is evident in astrocyte (arrow) of ts 1-only mouse (left panel), but weak in ts 1-mino mouse (right panel). Magnifications: 40X. (C). Expression of CD68, which marks activated microglia and macrophages, was examined by immunofluorescence staining using brainstem sections from uninfected, ts 1-only and ts 1-mino mice. Magnifications: 20X.

    Journal: Brain Research

    Article Title: Attenuation of oxidative stress, inflammation and apoptosis by minocycline prevents retrovirus-induced neurodegeneration in mice

    doi: 10.1016/j.brainres.2009.06.007

    Figure Lengend Snippet: Minocycline has anti-inflammatory effects in ts 1-infected mice brainstem and cultured C1 astrocytes. (A). Cell lysates or nuclear extracts from uninfected, ts 1-only and ts 1-mino C1 astrocytes are probed with anti-lκBα, NF-κB p65 and COX-2. (B). Mice from two groups were sacrificed at 30 dpi. Paraffin sections of the brainstem used for immunohistochemistry (IHC) assay. These sections were incubated with anti-GFAP (brown) and anti-COX-2 antibodies as specified in Experimental Procedures. Strong COX-2 immunoreactivity (red) is evident in astrocyte (arrow) of ts 1-only mouse (left panel), but weak in ts 1-mino mouse (right panel). Magnifications: 40X. (C). Expression of CD68, which marks activated microglia and macrophages, was examined by immunofluorescence staining using brainstem sections from uninfected, ts 1-only and ts 1-mino mice. Magnifications: 20X.

    Article Snippet: The sections were then subjected to an antigen retrieval protocol, in which they were heated in 10 mM citrate buffer (pH 6.0) for 10 min, blocked with 5% normal goat serum in TBS, and then incubated overnight at 4°C with rat anti-COX-2 antibody (Chemicon International) at a dilution of 1:100, or with rabbit anti- GFAP (Sigma; at 1:200).

    Techniques: Infection, Mouse Assay, Cell Culture, Immunohistochemistry, Incubation, Expressing, Immunofluorescence, Staining

    Expression of Cx43 at the cell surface of GFAP + primary astrocytes. Primary astrocytes were double immune labeled with mouse anti-GFAP (astrocyte marker) and rabbit anti-Cx43 antisera. Cells subsequently were labeled with FITC goat anti-mouse IgG and Texas Red goat anti-rabbit IgG, respectively. Immunostained cells were counterstained with DAPI (blue). Merged images show that GFAP-positive astrocytes express high levels of punctate Cx43 staining at the cell surface. Insets show magnifications of the expression of Cx43 at the cell surface of GFAP + astrocytes.

    Journal: Journal of Virology

    Article Title: Mouse Hepatitis Virus Infection Remodels Connexin43-Mediated Gap Junction Intercellular Communication In Vitro and In Vivo

    doi: 10.1128/JVI.02420-15

    Figure Lengend Snippet: Expression of Cx43 at the cell surface of GFAP + primary astrocytes. Primary astrocytes were double immune labeled with mouse anti-GFAP (astrocyte marker) and rabbit anti-Cx43 antisera. Cells subsequently were labeled with FITC goat anti-mouse IgG and Texas Red goat anti-rabbit IgG, respectively. Immunostained cells were counterstained with DAPI (blue). Merged images show that GFAP-positive astrocytes express high levels of punctate Cx43 staining at the cell surface. Insets show magnifications of the expression of Cx43 at the cell surface of GFAP + astrocytes.

    Article Snippet: At 24 h p.i., total protein was extracted and the concentration was measured, and 5 μg of total protein was loaded for immunoblotting, probed with either rabbit anti-Cx43 antibody (Sigma), rabbit anti-GFAP antibody (Sigma), or internal control rabbit anti-γ-actin antibody (Biobharati Lifescience Pvt.

    Techniques: Expressing, Labeling, Marker, Staining

    Reduction of Cx43 protein expression due to MHV-A59 infection. Primary astrocytes were either mock infected or infected with MHV-A59, and total protein was extracted at 24 h p.i. Protein (5 μg) was resolved in SDS-PAGE, transferred to a PVDF membrane, and immunoprobed. (A) Upon infection with MHV-A59, whole-cell Cx43 protein levels were decreased compared to those of mock-infected cells. The internal loading control γ-actin showed similar signal intensity for all experiments. (B) Densitometric analysis showed that there was a 36.3% ± 3.3% (at an MOI of 2) and 40.8% ± 6.8% (at an MOI of 5) reduction in total Cx43 for MHV-infected cells compared to the level for mock-infected control cells. The total proteins isolated from primary astrocytes also were probed for GFAP, but expression was found to be similar in all experimental groups. The mean ± SEM incidences of six experimental replicates from three different experiments are shown (****, P

    Journal: Journal of Virology

    Article Title: Mouse Hepatitis Virus Infection Remodels Connexin43-Mediated Gap Junction Intercellular Communication In Vitro and In Vivo

    doi: 10.1128/JVI.02420-15

    Figure Lengend Snippet: Reduction of Cx43 protein expression due to MHV-A59 infection. Primary astrocytes were either mock infected or infected with MHV-A59, and total protein was extracted at 24 h p.i. Protein (5 μg) was resolved in SDS-PAGE, transferred to a PVDF membrane, and immunoprobed. (A) Upon infection with MHV-A59, whole-cell Cx43 protein levels were decreased compared to those of mock-infected cells. The internal loading control γ-actin showed similar signal intensity for all experiments. (B) Densitometric analysis showed that there was a 36.3% ± 3.3% (at an MOI of 2) and 40.8% ± 6.8% (at an MOI of 5) reduction in total Cx43 for MHV-infected cells compared to the level for mock-infected control cells. The total proteins isolated from primary astrocytes also were probed for GFAP, but expression was found to be similar in all experimental groups. The mean ± SEM incidences of six experimental replicates from three different experiments are shown (****, P

    Article Snippet: At 24 h p.i., total protein was extracted and the concentration was measured, and 5 μg of total protein was loaded for immunoblotting, probed with either rabbit anti-Cx43 antibody (Sigma), rabbit anti-GFAP antibody (Sigma), or internal control rabbit anti-γ-actin antibody (Biobharati Lifescience Pvt.

    Techniques: Expressing, Infection, SDS Page, Isolation

    In situ immunofluorescence data on infected brain tissue demonstrated alteration of Cx43 in GFAP-positive astrocytes. Cryosections of brain tissue from MHV-A59-infected and mock-infected mice were double immunolabeled for either GFAP (red) and viral N (green) protein (A to F) or Cx43 (red) and viral N (green) protein (G to L). Cells were counterstained with DAPI. Mock-infected (A to C) and virus-infected (D to F) astrocytes appeared to be morphologically normal (thin arrow in panels A and C and thick arrow in panels D and F). Abundant punctate Cx43 staining was observed (thin arrow in panels G and I) in mock-infected brain tissue. In contrast, significant loss of Cx43 staining was observed in MHV-A59-infected brain tissue (thick arrow in panels J and L).

    Journal: Journal of Virology

    Article Title: Mouse Hepatitis Virus Infection Remodels Connexin43-Mediated Gap Junction Intercellular Communication In Vitro and In Vivo

    doi: 10.1128/JVI.02420-15

    Figure Lengend Snippet: In situ immunofluorescence data on infected brain tissue demonstrated alteration of Cx43 in GFAP-positive astrocytes. Cryosections of brain tissue from MHV-A59-infected and mock-infected mice were double immunolabeled for either GFAP (red) and viral N (green) protein (A to F) or Cx43 (red) and viral N (green) protein (G to L). Cells were counterstained with DAPI. Mock-infected (A to C) and virus-infected (D to F) astrocytes appeared to be morphologically normal (thin arrow in panels A and C and thick arrow in panels D and F). Abundant punctate Cx43 staining was observed (thin arrow in panels G and I) in mock-infected brain tissue. In contrast, significant loss of Cx43 staining was observed in MHV-A59-infected brain tissue (thick arrow in panels J and L).

    Article Snippet: At 24 h p.i., total protein was extracted and the concentration was measured, and 5 μg of total protein was loaded for immunoblotting, probed with either rabbit anti-Cx43 antibody (Sigma), rabbit anti-GFAP antibody (Sigma), or internal control rabbit anti-γ-actin antibody (Biobharati Lifescience Pvt.

    Techniques: In Situ, Immunofluorescence, Infection, Mouse Assay, Immunolabeling, Staining

    In vitro phenotypes of hNPCs differentiated in the neuroblast assay (NBA), followed by enrichment of the immature neuron subpopulation using MACS. ( A ) Overall experimental design. hNPC neurospheres were dissociated into single cells and plated in the NBA, generating a mixed population of neurons (N) and astrocytes ( A ). The neurons were then isolated from the astrocytes via MACS. Lastly, phenotyping was performed on the cells during the three major phases of the experiment. ( B–D ) Fluorescent images of immunocytochemical staining show hNPCs significantly lose expression of the proliferation marker Ki67 during the NBA. Enrichment of the neuron subgroup after the NBA, using MACS, reduces Ki67 expression still further (scale bar = 20 μm) (49,6-diamidino-2-phenylindole (DAPI), blue). ( E–G ) hNPCs significantly gain expression of the immature neuron marker DCX during the NBA. Enrichment of the neuron subgroup after the NBA yields an even higher percentage of DCX+ cells (scale bar = 20 μm) (49,6-diamidino-2-phenylindole (DAPI), blue). ( H–J ) Expression of the astrocyte marker GFAP increases slightly during differentiation, but MACS purification removes virtually all the percentage of GFAP+ cells (scale bar = 20 μm) (49,6-diamidino-2-phenylindole (DAPI), blue). ( K ) Quantification of immunolabeled cells indicates the differentiation of hNPCs into immature neurons during the NBA, and the subsequent purification of neurons made possible by MACS. Means + s.e.m. * P

    Journal: Scientific Reports

    Article Title: Transplantation of Defined Populations of Differentiated Human Neural Stem Cell Progeny

    doi: 10.1038/srep23579

    Figure Lengend Snippet: In vitro phenotypes of hNPCs differentiated in the neuroblast assay (NBA), followed by enrichment of the immature neuron subpopulation using MACS. ( A ) Overall experimental design. hNPC neurospheres were dissociated into single cells and plated in the NBA, generating a mixed population of neurons (N) and astrocytes ( A ). The neurons were then isolated from the astrocytes via MACS. Lastly, phenotyping was performed on the cells during the three major phases of the experiment. ( B–D ) Fluorescent images of immunocytochemical staining show hNPCs significantly lose expression of the proliferation marker Ki67 during the NBA. Enrichment of the neuron subgroup after the NBA, using MACS, reduces Ki67 expression still further (scale bar = 20 μm) (49,6-diamidino-2-phenylindole (DAPI), blue). ( E–G ) hNPCs significantly gain expression of the immature neuron marker DCX during the NBA. Enrichment of the neuron subgroup after the NBA yields an even higher percentage of DCX+ cells (scale bar = 20 μm) (49,6-diamidino-2-phenylindole (DAPI), blue). ( H–J ) Expression of the astrocyte marker GFAP increases slightly during differentiation, but MACS purification removes virtually all the percentage of GFAP+ cells (scale bar = 20 μm) (49,6-diamidino-2-phenylindole (DAPI), blue). ( K ) Quantification of immunolabeled cells indicates the differentiation of hNPCs into immature neurons during the NBA, and the subsequent purification of neurons made possible by MACS. Means + s.e.m. * P

    Article Snippet: For immunostaining, free-floating sections were blocked in a solution of 1X PBS with 0.1–0.3% Triton X-100 and 5% normal goat serum, and then incubated at 4 °C overnight in primary antibody cocktails containing the following, as needed: rabbit anti-GFAP (1:500; catalog #Z0334; Dako), rabbit anti-Ki67 (1:500; NCL-Ki67p; Novocastra), rabbit anti-Doublecortin (1:1,000; ab77450; Abcam).

    Techniques: In Vitro, Magnetic Cell Separation, Isolation, Staining, Expressing, Marker, Purification, Immunolabeling

    In vivo survival and phenotypes of hNPCs and their neuronal progeny transplanted in P-zero NSG mice. ( A) Overall experimental design. One group of mice was grafted with hNPCs, while the other group was grafted with NBA-generated hNPC progeny. Mice in both groups were sacrificed after 10 and 56 days. ( B,C ) Representative images of GFP+ surviving cells in mice grafted with either hNPCs ( B ) or immature neurons ( C ) 8 weeks after transplantation. ( D ) Quantification of surviving GFP+ cells in mice grafted with either hNPCs sacrificed 8 weeks after transplantation. The number of surviving GFP+ cells per section were not significantly different between hNPC and neuron grafts. ( E–J ) Fluorescent images of immunohistochemical staining, 8 weeks after transplantation, show GFP+/GFAP+ hNPCs to be upregulated in the hNPC ( E–G ) versus neuron ( H–J ) grafts (scale bar = 20 μm). ( K ) Quantification of donor cell phenotypes in the ipsilateral ventricle, cortex, hippocampus, RMS, olfactory bulb and cerebellum. While no difference in DCX frequency was seen between grafted cell groups 8 weeks after transplantation, the hNPC graft did yield significantly more GFP+/GFAP+ cells. ( L–R ) Likewise GFP+/Ki67+ cells are more observable in hNPC ( L–N ) versus neuron ( O – Q ) grafts, 10 days after transplantation (scale bar = 20 μm). Quantitatively, ( R ) GFP+/Ki67+ cells were more frequent in the hNPC grafts than in the neuron grafts 10 days after transplantation (**), but this difference gradually disappeared by 8 weeks post implant. (OB = olfactory bulb, Ctx = cortex) Means + s.e.m. ### P

    Journal: Scientific Reports

    Article Title: Transplantation of Defined Populations of Differentiated Human Neural Stem Cell Progeny

    doi: 10.1038/srep23579

    Figure Lengend Snippet: In vivo survival and phenotypes of hNPCs and their neuronal progeny transplanted in P-zero NSG mice. ( A) Overall experimental design. One group of mice was grafted with hNPCs, while the other group was grafted with NBA-generated hNPC progeny. Mice in both groups were sacrificed after 10 and 56 days. ( B,C ) Representative images of GFP+ surviving cells in mice grafted with either hNPCs ( B ) or immature neurons ( C ) 8 weeks after transplantation. ( D ) Quantification of surviving GFP+ cells in mice grafted with either hNPCs sacrificed 8 weeks after transplantation. The number of surviving GFP+ cells per section were not significantly different between hNPC and neuron grafts. ( E–J ) Fluorescent images of immunohistochemical staining, 8 weeks after transplantation, show GFP+/GFAP+ hNPCs to be upregulated in the hNPC ( E–G ) versus neuron ( H–J ) grafts (scale bar = 20 μm). ( K ) Quantification of donor cell phenotypes in the ipsilateral ventricle, cortex, hippocampus, RMS, olfactory bulb and cerebellum. While no difference in DCX frequency was seen between grafted cell groups 8 weeks after transplantation, the hNPC graft did yield significantly more GFP+/GFAP+ cells. ( L–R ) Likewise GFP+/Ki67+ cells are more observable in hNPC ( L–N ) versus neuron ( O – Q ) grafts, 10 days after transplantation (scale bar = 20 μm). Quantitatively, ( R ) GFP+/Ki67+ cells were more frequent in the hNPC grafts than in the neuron grafts 10 days after transplantation (**), but this difference gradually disappeared by 8 weeks post implant. (OB = olfactory bulb, Ctx = cortex) Means + s.e.m. ### P

    Article Snippet: For immunostaining, free-floating sections were blocked in a solution of 1X PBS with 0.1–0.3% Triton X-100 and 5% normal goat serum, and then incubated at 4 °C overnight in primary antibody cocktails containing the following, as needed: rabbit anti-GFAP (1:500; catalog #Z0334; Dako), rabbit anti-Ki67 (1:500; NCL-Ki67p; Novocastra), rabbit anti-Doublecortin (1:1,000; ab77450; Abcam).

    Techniques: In Vivo, Mouse Assay, Generated, Transplantation Assay, Immunohistochemistry, Staining

    Longitudinal sections of the ON–PN interface and distal part of the PN graft of one wt mouse. A , The ON–PN interface immunostained with an antibody against laminin. The laminin-positive PN graft (∗) runs horizontally along the eye wall (†) and connects with the proximal ON stump ( arrow ). In the ON stump, only the meningeal sheath and the blood vessels are laminin-positive. B , The adjacent section immunostained for GFAP. The inner tissue of the ON stump is GFAP-positive ( arrow ). C , The distal part of the PN graft after anterograde labeling with rhodamine, with the focal point on a long fiber of regenerating RGC axon ( arrow ). Scale bar: A , B , 160 μm; C , 100 μm.

    Journal: The Journal of Neuroscience

    Article Title: Bcl-2 Overexpression Does Not EnhanceIn Vivo Axonal Regeneration of Retinal Ganglion Cells after Peripheral Nerve Transplantation in Adult Mice

    doi: 10.1523/JNEUROSCI.22-11-04468.2002

    Figure Lengend Snippet: Longitudinal sections of the ON–PN interface and distal part of the PN graft of one wt mouse. A , The ON–PN interface immunostained with an antibody against laminin. The laminin-positive PN graft (∗) runs horizontally along the eye wall (†) and connects with the proximal ON stump ( arrow ). In the ON stump, only the meningeal sheath and the blood vessels are laminin-positive. B , The adjacent section immunostained for GFAP. The inner tissue of the ON stump is GFAP-positive ( arrow ). C , The distal part of the PN graft after anterograde labeling with rhodamine, with the focal point on a long fiber of regenerating RGC axon ( arrow ). Scale bar: A , B , 160 μm; C , 100 μm.

    Article Snippet: The following primary antibodies (Abs) were used: rabbit anti-GFAP Ab (Dako Japan, Kyoto, Japan) and rabbit anti-laminin Ab (Sigma, St. Louis, MO).

    Techniques: Labeling

    Attenuated reactive astrocytosis after stroke in Smad1 cKO mice. (A) Images of IHC for reactive astrocyte marker GFAP on ipsilateral hemisphere at 7 days (top two panels) or 3 months (bottom panels) post-stroke. The border between the stroke core and peri-infarct area is outlined by dotted lines. Arrows point to striatal infarct core. (B) Enlarged IHC images of boxed areas in (A) at the cortical peri-infarct area (blue box in D) with the indicated reactive astrocyte markers GFAP, Nestin, and Vimentin. Enlarged images of GFAP IHC highlight the hypertrophic morphology of GFAP + astrocytes in mutants. (C) Quantification of the number of GFAP + astrocytes and the intensity of GFAP immunoreactivity (IR) at the peri-infarct area shown in (B). n = 4, one-way ANOVA for the number of astrocytes, unpaired Student’s t-test for GFAP intensity, ***p

    Journal: PLoS ONE

    Article Title: Attenuation of Cerebral Ischemic Injury in Smad1 Deficient Mice

    doi: 10.1371/journal.pone.0136967

    Figure Lengend Snippet: Attenuated reactive astrocytosis after stroke in Smad1 cKO mice. (A) Images of IHC for reactive astrocyte marker GFAP on ipsilateral hemisphere at 7 days (top two panels) or 3 months (bottom panels) post-stroke. The border between the stroke core and peri-infarct area is outlined by dotted lines. Arrows point to striatal infarct core. (B) Enlarged IHC images of boxed areas in (A) at the cortical peri-infarct area (blue box in D) with the indicated reactive astrocyte markers GFAP, Nestin, and Vimentin. Enlarged images of GFAP IHC highlight the hypertrophic morphology of GFAP + astrocytes in mutants. (C) Quantification of the number of GFAP + astrocytes and the intensity of GFAP immunoreactivity (IR) at the peri-infarct area shown in (B). n = 4, one-way ANOVA for the number of astrocytes, unpaired Student’s t-test for GFAP intensity, ***p

    Article Snippet: Primary antibodies used for immunohistochemistry include: rabbit GFAP (Dako Z0334, 1:800), mouse Nestin (Abcam ab6142, 1:100), chicken Vimentin (Novus Biologicals NB300-223, 1:2000), rabbit S100 (Dako Z0311, 1:400), mouse NeuN (Millipore MAB377, 1:200), rabbit Iba1 (Wako 019–19741, 1:500), rabbit activated Caspase 3 (R & D Systems AF835, 1:400), mouse 8-hydroxyl-2’-deoxyguanosine (8-oxo-dG) (Trevigen 4354-MC-050, 1:350), rabbit p53 (Santa Cruz sc6243, 1:50), rat CD31 (BD 553370, 1:250), rabbit Olig2 (Millipore AB9610, 1:500), and chicken GFP (Aves Labs GFP-1020, 1:500).

    Techniques: Mouse Assay, Immunohistochemistry, Marker

    Fibrinogen-induced SVZ astrocytes add to cortical scar. a Representation of the different lesion regions quantified by immunoreactivity. b Fibrinogen deposition (red) and GFAP + astrocytes in the brain of fibrinogen-depleted mice (Ancrod) compared to controls (NaCl) at 6 days after PT (left). Yellow dotted line delineates the lesion area. The white boxes indicate the enlargement of the lesion region quantified for GFAP (right, top) and CSPGs (right, bottom) immunoreactivities. Quantifications of GFAP and CSPG immunoreactivities in the different lesion regions ( n = 4 mice, mean ± s.e.m, one-way ANOVA and Bonferroni’s multiple comparisons test, * P

    Journal: Nature Communications

    Article Title: Fibrinogen induces neural stem cell differentiation into astrocytes in the subventricular zone via BMP signaling

    doi: 10.1038/s41467-020-14466-y

    Figure Lengend Snippet: Fibrinogen-induced SVZ astrocytes add to cortical scar. a Representation of the different lesion regions quantified by immunoreactivity. b Fibrinogen deposition (red) and GFAP + astrocytes in the brain of fibrinogen-depleted mice (Ancrod) compared to controls (NaCl) at 6 days after PT (left). Yellow dotted line delineates the lesion area. The white boxes indicate the enlargement of the lesion region quantified for GFAP (right, top) and CSPGs (right, bottom) immunoreactivities. Quantifications of GFAP and CSPG immunoreactivities in the different lesion regions ( n = 4 mice, mean ± s.e.m, one-way ANOVA and Bonferroni’s multiple comparisons test, * P

    Article Snippet: Primary antibodies used were rabbit anti-Aqp4 (1:300, Santa Cruz), rat anti-BrdU (1:300, Abcam), goat anti-CD13 (1:500, R & D Systems), rabbit anti-CD31 (1:50, Abcam), mouse anti-CSPG (1:200, Sigma), guinea pig anti-Doublecortin (1:1000, Millipore), rabbit anti-δ-GFAP (1:500, Millipore), rabbit anti-GFAP (1:2000, Abcam), rat anti-GFAP (1:2000, Invitrogen), goat anti-GFP (1:2000, Abcam), rabbit anti-GFP (1:2000, Abcam), rabbit anti-Fibrinogen (1:10000, USBiological), sheep anti-Fibrinogen (1:500, USBiological), rabbit anti-Id3 (1:1000, Calbioreagent), goat anti-Lcn2 (1:1000, R & D Systems), goat anti-Nestin (1:200, Santa Cruz), goat anti-Nestin (1:500, Antibodies-Online), rabbit anti NeuN (1:1000, Abcam), rabbit anti-S100β (1:2000, Abcam), rabbit anti-Thbs4 (1:300, Genetex), sheep anti-Thbs4 (1:1000, R & D Systems) and secondary antibodies used included donkey antibodies to rabbit, rat, guinea pig, mouse, sheep, and goat conjugated with Alexa Fluor 488, 594, or 405 (1:200, Jackson ImmunoResearch Laboratories).

    Techniques: Mouse Assay

    Fibrinogen deposition in the SVZ after cortical injury. a GFAP immunostaining (green) around the lesion core (indicated by yellow dotted line) and in the SVZ. White box indicating enlargement of SVZ showing GFAP expression in the adult SVZ 7 days after PT (right) ( n = 4 mice). Scale bars, 369 μm, left and 57 µm, enlargement. b Fibrinogen immunostaining (red) in the lesion core and in the SVZ 1 day after PT. White box indicating enlargement of SVZ showing fibrinogen deposition in the SVZ 1 day after PT (right) ( n = 4 mice). Scale bars, 428 μm, left and 120 µm, enlargement. c GFAP (green) and fibrinogen (red) immunostainings in the lesion core (left) and in the SVZ (right) 3, 5 and 7 days after stroke (MCAO). White box indicating enlargement of SVZ showing increased GFAP expression colocalizing with fibrinogen (red) deposition in the SVZ 3 days after stroke. Scale bars, 543 μm, left and 50 µm, right ( n = 3 mice). d δ-GFAP (green) and fibrinogen (red) immunostainings in the SVZ of patients after stroke compared to healthy controls. White box indicating quantification area. Scale bar, 25 μm. Quantification of fibrinogen immunoreactivity in the SVZ per area ( n = 5 (healthy control), n = 6 (stroke patients, mean ± s.e.m., unpaired Student’s t -test, * P

    Journal: Nature Communications

    Article Title: Fibrinogen induces neural stem cell differentiation into astrocytes in the subventricular zone via BMP signaling

    doi: 10.1038/s41467-020-14466-y

    Figure Lengend Snippet: Fibrinogen deposition in the SVZ after cortical injury. a GFAP immunostaining (green) around the lesion core (indicated by yellow dotted line) and in the SVZ. White box indicating enlargement of SVZ showing GFAP expression in the adult SVZ 7 days after PT (right) ( n = 4 mice). Scale bars, 369 μm, left and 57 µm, enlargement. b Fibrinogen immunostaining (red) in the lesion core and in the SVZ 1 day after PT. White box indicating enlargement of SVZ showing fibrinogen deposition in the SVZ 1 day after PT (right) ( n = 4 mice). Scale bars, 428 μm, left and 120 µm, enlargement. c GFAP (green) and fibrinogen (red) immunostainings in the lesion core (left) and in the SVZ (right) 3, 5 and 7 days after stroke (MCAO). White box indicating enlargement of SVZ showing increased GFAP expression colocalizing with fibrinogen (red) deposition in the SVZ 3 days after stroke. Scale bars, 543 μm, left and 50 µm, right ( n = 3 mice). d δ-GFAP (green) and fibrinogen (red) immunostainings in the SVZ of patients after stroke compared to healthy controls. White box indicating quantification area. Scale bar, 25 μm. Quantification of fibrinogen immunoreactivity in the SVZ per area ( n = 5 (healthy control), n = 6 (stroke patients, mean ± s.e.m., unpaired Student’s t -test, * P

    Article Snippet: Primary antibodies used were rabbit anti-Aqp4 (1:300, Santa Cruz), rat anti-BrdU (1:300, Abcam), goat anti-CD13 (1:500, R & D Systems), rabbit anti-CD31 (1:50, Abcam), mouse anti-CSPG (1:200, Sigma), guinea pig anti-Doublecortin (1:1000, Millipore), rabbit anti-δ-GFAP (1:500, Millipore), rabbit anti-GFAP (1:2000, Abcam), rat anti-GFAP (1:2000, Invitrogen), goat anti-GFP (1:2000, Abcam), rabbit anti-GFP (1:2000, Abcam), rabbit anti-Fibrinogen (1:10000, USBiological), sheep anti-Fibrinogen (1:500, USBiological), rabbit anti-Id3 (1:1000, Calbioreagent), goat anti-Lcn2 (1:1000, R & D Systems), goat anti-Nestin (1:200, Santa Cruz), goat anti-Nestin (1:500, Antibodies-Online), rabbit anti NeuN (1:1000, Abcam), rabbit anti-S100β (1:2000, Abcam), rabbit anti-Thbs4 (1:300, Genetex), sheep anti-Thbs4 (1:1000, R & D Systems) and secondary antibodies used included donkey antibodies to rabbit, rat, guinea pig, mouse, sheep, and goat conjugated with Alexa Fluor 488, 594, or 405 (1:200, Jackson ImmunoResearch Laboratories).

    Techniques: Immunostaining, Expressing, Mouse Assay

    Single neurons were grown on microislands consisting of purified astrocytes. Neurons were identified by labeling with the fluorescein-conjugated C fragment of tetanus toxin (CFITC; Left ), and anti-glial fibrillary acidic protein (GFAP) antibody was used to identify astrocytes ( Right ).

    Journal: Proceedings of the National Academy of Sciences of the United States of America

    Article Title: Physiological astrocytic calcium levels stimulate glutamate release to modulate adjacent neurons

    doi:

    Figure Lengend Snippet: Single neurons were grown on microislands consisting of purified astrocytes. Neurons were identified by labeling with the fluorescein-conjugated C fragment of tetanus toxin (CFITC; Left ), and anti-glial fibrillary acidic protein (GFAP) antibody was used to identify astrocytes ( Right ).

    Article Snippet: A polyclonal antibody (Sigma; catalogue no. G9269; 1:100 dilution) was used to probe for glial fibrillary acidic protein, confirming a greater than 95% purity of astrocyte type-1 culture.

    Techniques: Purification, Labeling

    SIRT2 is an oligodendroglial protein. A–H , Double-immunofluorescence labeling of SIRT2 (red in all panels) plus CNP ( A–D ), GFAP ( E ), OX42 ( F ), neurofilament ( G , H ), or MBP ( I , J ) in the rat cerebellar cortex ( A , E ), caudate–putamen ( B , F ), cerebral cortex ( I ; layers III and IV), dentate gyrus ( J ), cross ( C , G ) or longitudinal sections ( D , H ) of cervical spinal white ( C , G ) or gray ( D , H ) matter. The inset in A or E is the observation of the marked area at a higher magnification. In A–D , note that, although it overlapped CNP (green) in most oligodendrocytic processes, SIRT2 (red) was undetected or only faintly positive in most oligodendroglial cell bodies that showed strong CNP staining (open arrowheads). K , L , On longitudinal sections of cervical spinal white matter, double-immunofluorescence labeling ( K ) of SIRT2 (red) together with pan-sodium channel (NavP, green), or triple labeling ( L ) of SIRT2 (red) together with NavP (green) and potassium channel Kv1.2 (green) showing SIRT2 distribution in the juxtanodal and paranodal domains of myelin sheath. The arrows point to examples of the NavP clusters at the nodes of Ranvier. The inset in L shows an example from the marked area at a higher magnification. Note the SIRT2-positive bands flanking the node of Ranvier and in between the strongly Kv1.2-positive juxtaparanodes. Gr, Granule cell layer of cerebellar cortex ( A , E ) or the dentate gyrus ( J ). Mol, Molecular layer of cerebellar cortex; Pur, Purkinje cell layer of cerebellar cortex. Scale bars: B , D , F , H–J , 20 μm; A , E , 50 μm; C , G , K , L , 10 μm.

    Journal: The Journal of Neuroscience

    Article Title: Sirtuin 2, a Mammalian Homolog of Yeast Silent Information Regulator-2 Longevity Regulator, Is an Oligodendroglial Protein That Decelerates Cell Differentiation through Deacetylating α-Tubulin

    doi: 10.1523/JNEUROSCI.4181-06.2007

    Figure Lengend Snippet: SIRT2 is an oligodendroglial protein. A–H , Double-immunofluorescence labeling of SIRT2 (red in all panels) plus CNP ( A–D ), GFAP ( E ), OX42 ( F ), neurofilament ( G , H ), or MBP ( I , J ) in the rat cerebellar cortex ( A , E ), caudate–putamen ( B , F ), cerebral cortex ( I ; layers III and IV), dentate gyrus ( J ), cross ( C , G ) or longitudinal sections ( D , H ) of cervical spinal white ( C , G ) or gray ( D , H ) matter. The inset in A or E is the observation of the marked area at a higher magnification. In A–D , note that, although it overlapped CNP (green) in most oligodendrocytic processes, SIRT2 (red) was undetected or only faintly positive in most oligodendroglial cell bodies that showed strong CNP staining (open arrowheads). K , L , On longitudinal sections of cervical spinal white matter, double-immunofluorescence labeling ( K ) of SIRT2 (red) together with pan-sodium channel (NavP, green), or triple labeling ( L ) of SIRT2 (red) together with NavP (green) and potassium channel Kv1.2 (green) showing SIRT2 distribution in the juxtanodal and paranodal domains of myelin sheath. The arrows point to examples of the NavP clusters at the nodes of Ranvier. The inset in L shows an example from the marked area at a higher magnification. Note the SIRT2-positive bands flanking the node of Ranvier and in between the strongly Kv1.2-positive juxtaparanodes. Gr, Granule cell layer of cerebellar cortex ( A , E ) or the dentate gyrus ( J ). Mol, Molecular layer of cerebellar cortex; Pur, Purkinje cell layer of cerebellar cortex. Scale bars: B , D , F , H–J , 20 μm; A , E , 50 μm; C , G , K , L , 10 μm.

    Article Snippet: Anti-pan-actin C4 (IB, 1:2000), anti-myelin basic protein (MBP) (ICC, 1:200), and anti-glial fibrillary acidic protein (GFAP) (ICC, 1:200) were obtained from Chemicon (Temecula, CA).

    Techniques: Immunofluorescence, Labeling, Staining

    Double immunofluorescence straining for expression and cell-specific distribution of SIRT3 protein. Typical photomicrographs illustrated immunofluorescence staining of SIRT3 in cerebral cortex of rats in the sham group and the 24 h post-SAH group. NeuN, GFAP, and DAPI (blue) were used as markers for neurons, astrocytes, and nuclei, respectively. SIRT3 was seldom colocalized with GFAP in the astrocytes in all rats (white circles). By contrast, SIRT3 was abundant and colocalized with NeuN in neurons of sham groups rats (orange color in the merged photomicrographs, indicated by white arrows), which markedly decreased after experimental SAH.

    Journal: BioMed Research International

    Article Title: SIRT3 Expression Decreases with Reactive Oxygen Species Generation in Rat Cortical Neurons during Early Brain Injury Induced by Experimental Subarachnoid Hemorrhage

    doi: 10.1155/2016/8263926

    Figure Lengend Snippet: Double immunofluorescence straining for expression and cell-specific distribution of SIRT3 protein. Typical photomicrographs illustrated immunofluorescence staining of SIRT3 in cerebral cortex of rats in the sham group and the 24 h post-SAH group. NeuN, GFAP, and DAPI (blue) were used as markers for neurons, astrocytes, and nuclei, respectively. SIRT3 was seldom colocalized with GFAP in the astrocytes in all rats (white circles). By contrast, SIRT3 was abundant and colocalized with NeuN in neurons of sham groups rats (orange color in the merged photomicrographs, indicated by white arrows), which markedly decreased after experimental SAH.

    Article Snippet: The sections were first cocultured with 5% normal fetal bovine serum in PBS including 0.1% Triton X-100 for 2 h and then with anti-neuron-specific nuclear protein (NeuN) antibody (1 : 200, Millipore, USA) and anti-SIRT3 antibody (1 : 100) or anti-glial fibrillary acidic protein (GFAP) antibody (1 : 200, Millipore, USA) and anti-SIRT3 antibody (1 : 100, Santa Cruz Biotechnology Inc., USA) overnight at 4°C.

    Techniques: Immunofluorescence, Expressing, Staining

    Intraperitoneal injection of curcumin (200 μg/g BW) in P7 mice immediately post-HI has no effect on the levels of proliferation at 48 h. (A) Assessment of the number of BrdU+ cells in P7 naïve untreated animals or naïve and HI animals treated with curcumin or DMSO did not show differences between the groups in any of the regions. (naïve n = 4, naïve + curcumin n = 5, HI + DMSO n = 7, HI + curcumin n = 8). (B) Curcumin treatment in naïve or HI animals immediately after insult did not affect the percentage of IBA1 and BrdU double positive over BrdU+ cells per region and group. The percentage of proliferating IBA1+ cells was low (10–20% of the BrdU+ cells). (C) The percentage of GFAP and BrdU double positive over BrdU+ cells per region and group was not affected by curcumin treatment in naïve or HI animals. The percentage of proliferating GFAP+ cells was low (10–20% of the BrdU+ cells). (D) The percentage of NG2 and BrdU double positive over BrdU+ cells per region and group was not affected by curcumin treatment in naïve or HI animals. The percentage of proliferating NG2+ cells was low (10–20% of the BrdU+ cells). (E) The percentage of NeuN and BrdU double positive over BrdU+ cells per region and group was not affected by curcumin treatment in naïve or HI animals. The percentage of proliferating NeuN+ cells comprised 80–90% of the BrdU+ cells and was the highest of all four studied cell types. (F,J,N) Immunofluorescence for rat polyclonal anti-BrdU (F,N) in green superimposed on the rabbit polyclonal anti-IBA1 (J,N) in red, and nuclear DAPI fluorescence in blue (N) . Note the co-localization of BrdU and IBA1 (white arrows) and the lack of such co- localization (empty arrows). (G,K,O) Immunofluorescence for rat polyclonal anti-BrdU (G,O) in green superimposed on the rabbit polyclonal anti-GFAP (K,O) in red, and nuclear DAPI fluorescence in blue (O) . Note the co-localization of BrdU and GFAP (full arrows) and the lack of such co-localization (empty arrows). (H,L,P) Immunofluorescence for rat polyclonal anti-BrdU (H,P) in green superimposed on the guinea pig polyclonal anti-NG2 (L,P) in red, and nuclear DAPI fluorescence in blue (P) . Note the co-localization of BrdU and NG2 (full arrows) and the lack of such co-localization (empty arrows). (I,M,Q) Immunofluorescence for rat polyclonal anti-BrdU (I,Q) in green superimposed on the mouse monoclonal anti-NeuN (M,Q) in red, and nuclear DAPI fluorescence in blue (Q) . Note the high level of co-localization of BrdU and NeuN (full arrows) and the lack of such co-localization (empty arrows). Abbreviations: CTX, cerebral cortex; EC – external capsule; HIP – hippocampus; PYR, pyriform cortex; STR, striatum; THAL, thalamus. Scale bar: 126 μm.

    Journal: Frontiers in Physiology

    Article Title: Curcumin: Novel Treatment in Neonatal Hypoxic-Ischemic Brain Injury

    doi: 10.3389/fphys.2019.01351

    Figure Lengend Snippet: Intraperitoneal injection of curcumin (200 μg/g BW) in P7 mice immediately post-HI has no effect on the levels of proliferation at 48 h. (A) Assessment of the number of BrdU+ cells in P7 naïve untreated animals or naïve and HI animals treated with curcumin or DMSO did not show differences between the groups in any of the regions. (naïve n = 4, naïve + curcumin n = 5, HI + DMSO n = 7, HI + curcumin n = 8). (B) Curcumin treatment in naïve or HI animals immediately after insult did not affect the percentage of IBA1 and BrdU double positive over BrdU+ cells per region and group. The percentage of proliferating IBA1+ cells was low (10–20% of the BrdU+ cells). (C) The percentage of GFAP and BrdU double positive over BrdU+ cells per region and group was not affected by curcumin treatment in naïve or HI animals. The percentage of proliferating GFAP+ cells was low (10–20% of the BrdU+ cells). (D) The percentage of NG2 and BrdU double positive over BrdU+ cells per region and group was not affected by curcumin treatment in naïve or HI animals. The percentage of proliferating NG2+ cells was low (10–20% of the BrdU+ cells). (E) The percentage of NeuN and BrdU double positive over BrdU+ cells per region and group was not affected by curcumin treatment in naïve or HI animals. The percentage of proliferating NeuN+ cells comprised 80–90% of the BrdU+ cells and was the highest of all four studied cell types. (F,J,N) Immunofluorescence for rat polyclonal anti-BrdU (F,N) in green superimposed on the rabbit polyclonal anti-IBA1 (J,N) in red, and nuclear DAPI fluorescence in blue (N) . Note the co-localization of BrdU and IBA1 (white arrows) and the lack of such co- localization (empty arrows). (G,K,O) Immunofluorescence for rat polyclonal anti-BrdU (G,O) in green superimposed on the rabbit polyclonal anti-GFAP (K,O) in red, and nuclear DAPI fluorescence in blue (O) . Note the co-localization of BrdU and GFAP (full arrows) and the lack of such co-localization (empty arrows). (H,L,P) Immunofluorescence for rat polyclonal anti-BrdU (H,P) in green superimposed on the guinea pig polyclonal anti-NG2 (L,P) in red, and nuclear DAPI fluorescence in blue (P) . Note the co-localization of BrdU and NG2 (full arrows) and the lack of such co-localization (empty arrows). (I,M,Q) Immunofluorescence for rat polyclonal anti-BrdU (I,Q) in green superimposed on the mouse monoclonal anti-NeuN (M,Q) in red, and nuclear DAPI fluorescence in blue (Q) . Note the high level of co-localization of BrdU and NeuN (full arrows) and the lack of such co-localization (empty arrows). Abbreviations: CTX, cerebral cortex; EC – external capsule; HIP – hippocampus; PYR, pyriform cortex; STR, striatum; THAL, thalamus. Scale bar: 126 μm.

    Article Snippet: Briefly, the sections were incubated overnight with rat anti-CD11b αM integrin subunit (1:5,000, Serotec, UK), rabbit polyclonal anti-GFAP (1:6,000, DAKO, UK), rabbit anti-iNOS (1:500, Santa Cruz, USA), polyclonal rat anti-BrdU (1:400, Abcam, UK), or rabbit anti-myelin basic protein (MBP) (1:200, Abcam, Cambridge, UK) primary antibodies, for 1 h with biotinylated goat anti-rabbit or -rat (1:100, Vector, UK) secondary antibodies, followed by incubation with Avidin-Biotinylated horseradish peroxidase Complex (Vector, UK) and visualization with diaminobenzidine/H2 O2 (Fisher Scientific, UK).

    Techniques: Injection, Mouse Assay, Immunofluorescence, Fluorescence

    DPI reduced ethanol-induced caspase-3 activation and Fluoro-Jade B staining . Male C57BL/6 mice were treated intragastrically with ethanol (5 g/kg, i.g.) daily for 10 days. DPI (3 mg/kg) was injected intraperitoneally 0.5 hr and 24 hr after the last dose of ethanol. Mice were sacrificed 3 hrs after the last dose of DPI. Brain sections were stained with polyclonal cleave caspase-3 antibody and Fluoro-Jade B, respectively. (A) Ethanol increased the number of caspase-3+IR cells. DPI significantly reduced ethanol-induced increases in the number of caspase-3+IR cells. (B) Fluorescent intensity of Fluoro-Jade B positive cells in cortex was quantified by BioQuant image analysis system. Ethanol-treated mouse brains showed more Fluoro-Jade B positive cells than water-treated mouse brains. DPI significantly decreased ethanol-induced increases in the fluorescent intensity of Fluoro-Jade B positive cells. ** P

    Journal: Journal of Neuroinflammation

    Article Title: NADPH oxidase and reactive oxygen species contribute to alcohol-induced microglial activation and neurodegeneration

    doi: 10.1186/1742-2094-9-5

    Figure Lengend Snippet: DPI reduced ethanol-induced caspase-3 activation and Fluoro-Jade B staining . Male C57BL/6 mice were treated intragastrically with ethanol (5 g/kg, i.g.) daily for 10 days. DPI (3 mg/kg) was injected intraperitoneally 0.5 hr and 24 hr after the last dose of ethanol. Mice were sacrificed 3 hrs after the last dose of DPI. Brain sections were stained with polyclonal cleave caspase-3 antibody and Fluoro-Jade B, respectively. (A) Ethanol increased the number of caspase-3+IR cells. DPI significantly reduced ethanol-induced increases in the number of caspase-3+IR cells. (B) Fluorescent intensity of Fluoro-Jade B positive cells in cortex was quantified by BioQuant image analysis system. Ethanol-treated mouse brains showed more Fluoro-Jade B positive cells than water-treated mouse brains. DPI significantly decreased ethanol-induced increases in the fluorescent intensity of Fluoro-Jade B positive cells. ** P

    Article Snippet: Polyclonal Rabbit anti-Glial Fibrillary Acidic Protein was from DakoCytomation (Glostrup, Denmark).

    Techniques: Activation Assay, Staining, Mouse Assay, Injection

    Antibody validation and morphometric analysis of brainstem astrocytes. (a–c) PreBötC GFAP‐positive astrocytes immunostained with rabbit anti‐GFAP polyclonal antibody (green; a) and mouse anti‐GFAP monoclonal antibody (red; b). Merged low magnification and high magnification (inset scale bar: 50 μm) images (c) illustrates colocalization of antibody labeling. (d) 2D maximum projection image of individual astrocyte (blue), morphologically reconstructed in 3D using Neurolucida 360, within the field of astrocytes identified by GFAP immunoreactivity (green; RRID: http://scicrunch.org/resolver/AB-10013382 ) in the preBötC region. Maximum projection image of astrocyte field and reconstructed astrocyte was rendered from Z stack of confocal images. (e) Sholl analysis to characterize astrocyte arbour complexity, including number of processes, process lengths, and number of branch points, was performed by applying nested concentric spheres increasing in size by a constant change in radius (5 μm increments) from the center of the astroglial soma (maximum projection image shown, and see Section 2.6 for detail). (f) Convex hull analysis was performed by connecting the tips of distal processes (terminal points) to generate a convex polygon (projection image shown) to determine the volume and surface area of the physical space occupied by the polygon (see Section 2.6)

    Journal: The Journal of Comparative Neurology

    Article Title: Morphometric analysis of astrocytes in brainstem respiratory regions, et al. Morphometric analysis of astrocytes in brainstem respiratory regions

    doi: 10.1002/cne.24472

    Figure Lengend Snippet: Antibody validation and morphometric analysis of brainstem astrocytes. (a–c) PreBötC GFAP‐positive astrocytes immunostained with rabbit anti‐GFAP polyclonal antibody (green; a) and mouse anti‐GFAP monoclonal antibody (red; b). Merged low magnification and high magnification (inset scale bar: 50 μm) images (c) illustrates colocalization of antibody labeling. (d) 2D maximum projection image of individual astrocyte (blue), morphologically reconstructed in 3D using Neurolucida 360, within the field of astrocytes identified by GFAP immunoreactivity (green; RRID: http://scicrunch.org/resolver/AB-10013382 ) in the preBötC region. Maximum projection image of astrocyte field and reconstructed astrocyte was rendered from Z stack of confocal images. (e) Sholl analysis to characterize astrocyte arbour complexity, including number of processes, process lengths, and number of branch points, was performed by applying nested concentric spheres increasing in size by a constant change in radius (5 μm increments) from the center of the astroglial soma (maximum projection image shown, and see Section 2.6 for detail). (f) Convex hull analysis was performed by connecting the tips of distal processes (terminal points) to generate a convex polygon (projection image shown) to determine the volume and surface area of the physical space occupied by the polygon (see Section 2.6)

    Article Snippet: The rabbit polyclonal anti‐GFAP antibody (1:1000; DAKO, catalog #z‐0334, RRID: http://scicrunch.org/resolver/AB_10013382 ) was isolated from cow spinal cord and cross‐reacts with the intra‐cytoplasmic filamentous protein of an epitope of the astrocytic cytoskeleton in mouse, rat, and human [manufacturer's technical information; also see (L F Eng, Ghirnikar, & Lee, )].

    Techniques: Antibody Labeling

    Induction of αS aggregates in primary neuronal-glial cultures by exogenous treatment with αS fibrils Neuronal-glial cultures were maintained for 6 days, incubated with 21–140 human αS fibrils (20 ug/ml) for 10 days ( A, B, D ) or maintained untreated for an additional 10 days ( C ), and analyzed by immunofluorescence. Cultures were labeled with antibodies ( A ) pSer129/81A (green) and polyclonal NFL (red), ( B ) pSer129/81A (green) and rabbit monoclonal C28E10-NFL, or ( C, D ) Syn 506 and βIII-tubulin. Arrows depict pSer129/81A staining that does not co-localize with NFL. Merged images are shown. Bar = 100 um and 250 um for the higher magnification images on the right.

    Journal: Journal of neurochemistry

    Article Title: Proteolysis of α-Synuclein Fibrils in the Lysosomal Pathway Limits Induction of Inclusion Pathology

    doi: 10.1111/jnc.13743

    Figure Lengend Snippet: Induction of αS aggregates in primary neuronal-glial cultures by exogenous treatment with αS fibrils Neuronal-glial cultures were maintained for 6 days, incubated with 21–140 human αS fibrils (20 ug/ml) for 10 days ( A, B, D ) or maintained untreated for an additional 10 days ( C ), and analyzed by immunofluorescence. Cultures were labeled with antibodies ( A ) pSer129/81A (green) and polyclonal NFL (red), ( B ) pSer129/81A (green) and rabbit monoclonal C28E10-NFL, or ( C, D ) Syn 506 and βIII-tubulin. Arrows depict pSer129/81A staining that does not co-localize with NFL. Merged images are shown. Bar = 100 um and 250 um for the higher magnification images on the right.

    Article Snippet: A polyclonal rabbit anti-glial fibrillary acidic protein antibody was purchased from Dako (Carpinteria, CA).

    Techniques: Incubation, Immunofluorescence, Labeling, Staining

    DGC activation during the course of status epilepticus. ( A ) TRAPing of DGCs at T0 + 60 to T0 + 300 min. ( B ) Co-localization of TRAPed neurons in the dentate gyrus with the neuronal marker protein NeuN, microglial marker protein Iba1 and astrocyte marker protein GFAP. The insets show boxed areas in the respective image. ( C ) The number of activated DGCs was plotted over time; n = 3 each for T0, T0 + 60, +120, +180, n = 4 for T0 + 240 and n = 2 for T0 + 300. The line depicts a linear regression fit of the data.

    Journal: Brain

    Article Title: Parallel pathways of seizure generalization

    doi: 10.1093/brain/awz170

    Figure Lengend Snippet: DGC activation during the course of status epilepticus. ( A ) TRAPing of DGCs at T0 + 60 to T0 + 300 min. ( B ) Co-localization of TRAPed neurons in the dentate gyrus with the neuronal marker protein NeuN, microglial marker protein Iba1 and astrocyte marker protein GFAP. The insets show boxed areas in the respective image. ( C ) The number of activated DGCs was plotted over time; n = 3 each for T0, T0 + 60, +120, +180, n = 4 for T0 + 240 and n = 2 for T0 + 300. The line depicts a linear regression fit of the data.

    Article Snippet: The antibodies used were anti-cFos antibody (1:300, sc52, Santa Cruz Biotechnology), anti-NeuN antibody (1:200, MAB377, Millipore), anti-GFAP antibody (1:300, Ab7260, Abcam), anti-Iba1 antibody (1: 300, CP 290 A, Biocare Medical), and anti-Parvalbumin antibody (1:500, Ab11427, Abcam).

    Techniques: Activation Assay, Marker

    Representative and quantitative immunohistochemistry of the peri-infarct cortex before the bregma (coordinate + 0.58) in normal WT, normal sEH KO, MCAO WT, MCAO sEH KO, MCAO with vehicle treatment and MCAO with AUDA treatment mice (n = 2 for normal groups, n = 5–6 for all groups subjected to MCAO). ( A – F , S ) The number of CD31-positive endothelial cells significantly increased in the sEH KO before and after MCAO and post-MCAO AUDA-treated mice, while ( G – L ) Note the increased phosphorylated TrkB (p-TrkB) immunoreactivity that partially co-localized with NeuN-positive neurons, particularly in the sEH KO and AUDA-treated mice (arrows) ( T ). The percentage of NeuN- and p-TrkB-positive double labeling in the neuronal population was significantly increased compared to that in the corresponding controls. ( M – R , U – V ) GFAP-positive astrogliosis and Iba1-positive microglial infiltration decreased in post-MCAO sEH KO and AUDA treatment groups relative to that in the corresponding controls. The scale bar represents 50 µm. * p

    Journal: Scientific Reports

    Article Title: Blockade of soluble epoxide hydrolase attenuates post-ischemic neuronal hyperexcitation and confers resilience against stroke with TrkB activation

    doi: 10.1038/s41598-017-18558-6

    Figure Lengend Snippet: Representative and quantitative immunohistochemistry of the peri-infarct cortex before the bregma (coordinate + 0.58) in normal WT, normal sEH KO, MCAO WT, MCAO sEH KO, MCAO with vehicle treatment and MCAO with AUDA treatment mice (n = 2 for normal groups, n = 5–6 for all groups subjected to MCAO). ( A – F , S ) The number of CD31-positive endothelial cells significantly increased in the sEH KO before and after MCAO and post-MCAO AUDA-treated mice, while ( G – L ) Note the increased phosphorylated TrkB (p-TrkB) immunoreactivity that partially co-localized with NeuN-positive neurons, particularly in the sEH KO and AUDA-treated mice (arrows) ( T ). The percentage of NeuN- and p-TrkB-positive double labeling in the neuronal population was significantly increased compared to that in the corresponding controls. ( M – R , U – V ) GFAP-positive astrogliosis and Iba1-positive microglial infiltration decreased in post-MCAO sEH KO and AUDA treatment groups relative to that in the corresponding controls. The scale bar represents 50 µm. * p

    Article Snippet: The slides were incubated in a blocking solution containing 3% donkey serum albumin (Abcam) and 0.3% Triton X-100 (Sigma-Aldrich) in TBS for 1 hour at room temperature and immunostained with primary antibodies including rabbit anti-CD31 (1:200, BioRad, MCA23886A), anti-p-TrkB (1:200, Abcam, ab109684), anti-NeuN (1:200, Abcam, ab104224), anti-GFAP (1:500, Abcam, ab7260), and anti-Iba1 (1:200, Novus, NB100-1028) overnight at 4 °C.

    Techniques: Immunohistochemistry, Mouse Assay, Labeling

    Detection of GFAP and cit-GFAP in livers from sham-operated control and BDL mice. The expression levels of GFAP and cit-GFAP in the liver tissues of control and BDL mice were analyzed by western blot using rabbit polyclonal anti-GFAP (Abcam, Cambridge UK) and citrullinated GFAP (CTGF-122R and CTGF-1221) antibodies. Note that GFAP and cit-GFAP were increased in the liver tissues of BDL mice compared with controls.

    Journal: PLoS ONE

    Article Title: Accumulation of citrullinated glial fibrillary acidic protein in a mouse model of bile duct ligation-induced hepatic fibrosis

    doi: 10.1371/journal.pone.0201744

    Figure Lengend Snippet: Detection of GFAP and cit-GFAP in livers from sham-operated control and BDL mice. The expression levels of GFAP and cit-GFAP in the liver tissues of control and BDL mice were analyzed by western blot using rabbit polyclonal anti-GFAP (Abcam, Cambridge UK) and citrullinated GFAP (CTGF-122R and CTGF-1221) antibodies. Note that GFAP and cit-GFAP were increased in the liver tissues of BDL mice compared with controls.

    Article Snippet: The membranes were then incubated with mouse monoclonal anti-PAD2 [ ], mouse monoclonal anti-F95 (anti-peptidyl-citrulline antibody, Millipore), rabbit polyclonal anti-GFAP (Abcam, Cambridge UK), mouse monoclonal citrullinated GFAP (CTGF-122R and CTGF-1221)[ ] and rabbit polyclonal glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (Santa Cruz Biotechnology, Santa Cruz, CA) antibodies in PBST overnight at 4°C.

    Techniques: Mouse Assay, Expressing, Western Blot

    PALS1 localizes with CRB1 and MUPP1 to the subapical region adjacent to adherens junctions at the outer limiting membrane. Immunohistochemistry on 3-month-old retinas showing confocal images of staining for proteins associated with the Crumbs complex, PALS1, CRB1, and MUPP1, and for GFAP, a Müller glial cell marker. Staining for the Crumbs complex shows fragmented staining (arrows) of PALS1, CRB1, and MUPP1 in the shPals1 -Rx Cre retinas, which indicate the dislocalization of the Crumbs complex proteins at the SAR adjacent to adherens junctions at the OLM. GFAP staining shows increased reactive gliosis in the mutant retinas. Scale bar, 50 μm.

    Journal: The Journal of Neuroscience

    Article Title: PALS1 Is Essential for Retinal Pigment Epithelium Structure and Neural Retina Stratification

    doi: 10.1523/JNEUROSCI.4430-11.2011

    Figure Lengend Snippet: PALS1 localizes with CRB1 and MUPP1 to the subapical region adjacent to adherens junctions at the outer limiting membrane. Immunohistochemistry on 3-month-old retinas showing confocal images of staining for proteins associated with the Crumbs complex, PALS1, CRB1, and MUPP1, and for GFAP, a Müller glial cell marker. Staining for the Crumbs complex shows fragmented staining (arrows) of PALS1, CRB1, and MUPP1 in the shPals1 -Rx Cre retinas, which indicate the dislocalization of the Crumbs complex proteins at the SAR adjacent to adherens junctions at the OLM. GFAP staining shows increased reactive gliosis in the mutant retinas. Scale bar, 50 μm.

    Article Snippet: Immunostainings were performed on cryosections using the following primary and secondary antibodies: rabbit anti-PALS1 (1/200, Millipore Bioscience Research Reagents, ProteinTech), mouse anti-β-catenin (1/300, BD), rabbit anti-CRB1 (1/100, AK2) , rabbit anti-glial fibrillary acidic protein (GFAP) (1/200, DAKO), mouse anti-MUPP1 (1/300, BD), mouse anti-glutamine synthetase (1/300, BD), rabbit anti-MPP3 (1/100, ProteinTech), rabbit anti-Recoverin (1/200, Millipore Bioscience Research Reagents), mouse anti-PKC α (1/300, BD), and rabbit anti-Caspase-3 (cleaved form, 1/200, Cell Signaling).

    Techniques: Immunohistochemistry, Staining, Marker, Mutagenesis

    iNSC transplantation suppressed the inflammatory response in mice brain. (a–l) Immunohistochemical staining for glial fibrillary acidic protein (GFAP) in the ipsilateral side of a poststroke brain. Scale bar: 1 mm. (b, c, e, f, h, i, k, l) High-magnification views of the field indicated by the boxed areas in (a), (d), (g), and (j), respectively. Scale bar: 50 μm. (m, n) Relative intensity of GFAP and Iba1 in the peri-infarcted lesion of the cortex was measured on days 7 and 28 of transplantation ( ∗ p

    Journal: Cell Transplantation

    Article Title: Novel Therapeutic Transplantation of Induced Neural Stem Cells for Stroke

    doi: 10.3727/096368916X692988

    Figure Lengend Snippet: iNSC transplantation suppressed the inflammatory response in mice brain. (a–l) Immunohistochemical staining for glial fibrillary acidic protein (GFAP) in the ipsilateral side of a poststroke brain. Scale bar: 1 mm. (b, c, e, f, h, i, k, l) High-magnification views of the field indicated by the boxed areas in (a), (d), (g), and (j), respectively. Scale bar: 50 μm. (m, n) Relative intensity of GFAP and Iba1 in the peri-infarcted lesion of the cortex was measured on days 7 and 28 of transplantation ( ∗ p

    Article Snippet: For immunohistochemistry, the following primary antibodies were used: rabbit anti-nestin antibody (1:100; Santa Cruz Biotechnology, Santa Cruz, CA, USA); rabbit anti-Sox2 antibody (1:100; Santa Cruz Biotechnology); rabbit anti-microtubule-associated protein 2 (MAP2) antibody (1:200; Millipore, Billerica, MA, USA); rabbit anti-β III tubulin (Tuj1) antibody (1:50; Covance Research, Princeton, NJ, USA); mouse anti-galactocerebroside (GalC) antibody (1:400; Millipore); rabbit anti-glial fibrillary acidic protein (GFAP) antibody (1:500; Dako, Glostrup, Denmark); rabbit anti-Iba1 antibody (1:1,000; Wako, Osaka, Japan); mouse anti-NeuN antibody (1:100; Millipore); and mouse anti-O4 antibody (1:100; R & D Systems, Minneapolis, MN, USA).

    Techniques: Transplantation Assay, Mouse Assay, Immunohistochemistry, Staining

    Induced neural stem cells (iNSCs) can differentiate into both neuronal and glial lineages in vitro. (a) Left: Morphology of undifferentiated iNSCs as assessed by bright-field microscopy. Scale bars: 50 μm. Right: Immunofluorescence microscopy images of undifferentiated iNSCs, using antibodies against nestin and (sex-determining region Y)-box 2 (Sox2). Scale bars: 50 μm. (b) Immunofluorescence microscopy images of neuronal differentiated iNSCs using antibodies against microtubule-associated protein 2 (MAP2) and rabbit anti-β III tubulin (Tuj1). Scale bars: 20 μm. (c) Immunofluorescence microscopy images of glial differentiated iNSCs using antibodies against the oligodendrocyte marker, anti-galactocerebroside (GalC), and an astrocyte marker, glial fibrillary acidic protein (GFAP). Scale bars: 20 μm.

    Journal: Cell Transplantation

    Article Title: Novel Therapeutic Transplantation of Induced Neural Stem Cells for Stroke

    doi: 10.3727/096368916X692988

    Figure Lengend Snippet: Induced neural stem cells (iNSCs) can differentiate into both neuronal and glial lineages in vitro. (a) Left: Morphology of undifferentiated iNSCs as assessed by bright-field microscopy. Scale bars: 50 μm. Right: Immunofluorescence microscopy images of undifferentiated iNSCs, using antibodies against nestin and (sex-determining region Y)-box 2 (Sox2). Scale bars: 50 μm. (b) Immunofluorescence microscopy images of neuronal differentiated iNSCs using antibodies against microtubule-associated protein 2 (MAP2) and rabbit anti-β III tubulin (Tuj1). Scale bars: 20 μm. (c) Immunofluorescence microscopy images of glial differentiated iNSCs using antibodies against the oligodendrocyte marker, anti-galactocerebroside (GalC), and an astrocyte marker, glial fibrillary acidic protein (GFAP). Scale bars: 20 μm.

    Article Snippet: For immunohistochemistry, the following primary antibodies were used: rabbit anti-nestin antibody (1:100; Santa Cruz Biotechnology, Santa Cruz, CA, USA); rabbit anti-Sox2 antibody (1:100; Santa Cruz Biotechnology); rabbit anti-microtubule-associated protein 2 (MAP2) antibody (1:200; Millipore, Billerica, MA, USA); rabbit anti-β III tubulin (Tuj1) antibody (1:50; Covance Research, Princeton, NJ, USA); mouse anti-galactocerebroside (GalC) antibody (1:400; Millipore); rabbit anti-glial fibrillary acidic protein (GFAP) antibody (1:500; Dako, Glostrup, Denmark); rabbit anti-Iba1 antibody (1:1,000; Wako, Osaka, Japan); mouse anti-NeuN antibody (1:100; Millipore); and mouse anti-O4 antibody (1:100; R & D Systems, Minneapolis, MN, USA).

    Techniques: In Vitro, Microscopy, Immunofluorescence, Marker

    TRPV4 activation was involved in glial activation and proinflammatory cytokines release. (A) Representative glial fibrillary acidic protein antibody (GFAP) and Iba-1 immunofluorescent images for the corpus callosum. Scale bar = 20 μm. (B,C) Quantitative analysis of GFAP- and Iba-1-positive cell numbers. The increased numbers of GFAP- and Iba-1-positive cells were reversed by the TRPV4 inhibitor RN-1734 ( n = 4). (D) Protein levels of tumor necrosis factor alpha (TNF-α) and interleukin 1beta (IL-1β) were detected by enzyme-linked immunosorbent assay (ELISA) ( n = 3). The data are expressed as the mean ± SEM. ** P

    Journal: Frontiers in Cellular Neuroscience

    Article Title: TRPV4 Inhibition Improved Myelination and Reduced Glia Reactivity and Inflammation in a Cuprizone-Induced Mouse Model of Demyelination

    doi: 10.3389/fncel.2018.00392

    Figure Lengend Snippet: TRPV4 activation was involved in glial activation and proinflammatory cytokines release. (A) Representative glial fibrillary acidic protein antibody (GFAP) and Iba-1 immunofluorescent images for the corpus callosum. Scale bar = 20 μm. (B,C) Quantitative analysis of GFAP- and Iba-1-positive cell numbers. The increased numbers of GFAP- and Iba-1-positive cells were reversed by the TRPV4 inhibitor RN-1734 ( n = 4). (D) Protein levels of tumor necrosis factor alpha (TNF-α) and interleukin 1beta (IL-1β) were detected by enzyme-linked immunosorbent assay (ELISA) ( n = 3). The data are expressed as the mean ± SEM. ** P

    Article Snippet: The primary antibodies used were mouse anti-2′,3′-cyclic nucleotide 3′ phosphodiesterase antibody (CNP; IgG, 1:1,000, Santa Cruz, Santa Cruz, CA, USA), rabbit anti-TRPV4 and rabbit anti-olig2 antibodies (both IgG, 1:200, Abcam Cambridge, UK), rabbit anti-glial fibrillary acidic protein antibody (GFAP; IgG, 1:500, Abcam, USA), rabbit anti-ionized calcium binding adaptor molecule-1 antibody (Iba-1; IgG, 1:1,000, Cell Signaling Technology/CST, Danvers, MA, USA) and rabbit anti-cleaved caspase3 antibody (IgG, 1:300, CST, Danvers, MA, USA).

    Techniques: Activation Assay, Enzyme-linked Immunosorbent Assay

    Confocal analysis of eEF1A-2/S1 during murine brain development. (A) One day after birth (P1), co-immunolocalization between neurofilament NF68 (red) and eEF1A-2/S1 (green) reveals absence of eEF1A-2/S1 in neurons. (B) Later in development at P14, co-immunolocalization reveals that both eEF1A-2/S1 (green) and NF68 (red) co-localize in neurons. However, eEF1A-2/S1 expression is weak. (C) The same co-localization (yellow) 20 days after birth (P20) reveals an increase in eEF1A-2/S1 abundance in the neuronal soma. (D) At P26, eEF1A-2/S1 (green) is clearly visible in the soma of neurons. (E) Twelve-month-old mouse brain shows intense co-localization (yellow) between EF1A-2/S1 and NF68 (red). (F) Co-immunolocalization of eEF1A-2/S1 (red) and GFAP (green) reveals that eEF1A-2/S1 is not expressed in astrocytes. Solid arrows indicate neuronal soma, open arrows indicate axons, and asterisks indicate nuclei.

    Journal: Brain research. Developmental brain research

    Article Title: Immuno-characterization of the switch of peptide elongation factors eEF1A-1/EF-1α and eEF1A-2/S1 in the central nervous system during mouse development

    doi: 10.1016/j.devbrainres.2003.10.011

    Figure Lengend Snippet: Confocal analysis of eEF1A-2/S1 during murine brain development. (A) One day after birth (P1), co-immunolocalization between neurofilament NF68 (red) and eEF1A-2/S1 (green) reveals absence of eEF1A-2/S1 in neurons. (B) Later in development at P14, co-immunolocalization reveals that both eEF1A-2/S1 (green) and NF68 (red) co-localize in neurons. However, eEF1A-2/S1 expression is weak. (C) The same co-localization (yellow) 20 days after birth (P20) reveals an increase in eEF1A-2/S1 abundance in the neuronal soma. (D) At P26, eEF1A-2/S1 (green) is clearly visible in the soma of neurons. (E) Twelve-month-old mouse brain shows intense co-localization (yellow) between EF1A-2/S1 and NF68 (red). (F) Co-immunolocalization of eEF1A-2/S1 (red) and GFAP (green) reveals that eEF1A-2/S1 is not expressed in astrocytes. Solid arrows indicate neuronal soma, open arrows indicate axons, and asterisks indicate nuclei.

    Article Snippet: Neuron-specific monoclonal antibody to neurofilament NF-68 and rabbit anti-glial fibrillary acidic protein (GFAP) were purchased from Sigma-Aldrich.

    Techniques: Expressing

    Accumulation of newborn neurons. (A) The merged confocal images confirmed the presence of BrdU (red), GFAP (red), DCX (green), and Nestin (green) expressing cells in ipsilateral brain sections. Scale bar=20 μmol/L. (B) Quantitative data of each cell type. Means±SD. n =10. b P

    Journal: Acta Pharmacologica Sinica

    Article Title: Ginsenoside Rd promotes neurogenesis in rat brain after transient focal cerebral ischemia via activation of PI3K/Akt pathway

    doi: 10.1038/aps.2014.156

    Figure Lengend Snippet: Accumulation of newborn neurons. (A) The merged confocal images confirmed the presence of BrdU (red), GFAP (red), DCX (green), and Nestin (green) expressing cells in ipsilateral brain sections. Scale bar=20 μmol/L. (B) Quantitative data of each cell type. Means±SD. n =10. b P

    Article Snippet: Materials and reagents The mouse anti-BrdU antibody was purchased from Santa Cruz Biotechnology (Santa Cruz, CA, USA), while the rabbit anti-DCX, rabbit anti-GFAP and mouse anti-Nestin antibodies were obtained from Cell Signaling Technology (Danvers, MA, USA).

    Techniques: Expressing

    a Claudin-5 (×60 magnification) and IgG (×20 magnification) staining in ipsilateral and contralateral striatum for LPS and positive control AMPA-challenged animals. Claudin-5 staining revealed no differences between contralateral and ipsilateral sides in the LPS-challenged animals, while for AMPA-challenged animals, there was visible damage to vessels in the ipsilateral side and some disruption in the contralateral hemisphere. IgG appears normal in ipsilateral and contralateral sides of LPS animals but shows signs of leakage in the ipsilateral side of AMPA animals. b Staining for microglia (CD11b, red ) and astrocytes (GFAP, green ), merged at ×10 and ×20 ( white rectangles ) magnification; c NeuN staining at ×20 magnification in core and contralateral striatum

    Journal: Molecular Imaging and Biology

    Article Title: Comparative Evaluation of Three TSPO PET Radiotracers in a LPS-Induced Model of Mild Neuroinflammation in Rats

    doi: 10.1007/s11307-016-0984-3

    Figure Lengend Snippet: a Claudin-5 (×60 magnification) and IgG (×20 magnification) staining in ipsilateral and contralateral striatum for LPS and positive control AMPA-challenged animals. Claudin-5 staining revealed no differences between contralateral and ipsilateral sides in the LPS-challenged animals, while for AMPA-challenged animals, there was visible damage to vessels in the ipsilateral side and some disruption in the contralateral hemisphere. IgG appears normal in ipsilateral and contralateral sides of LPS animals but shows signs of leakage in the ipsilateral side of AMPA animals. b Staining for microglia (CD11b, red ) and astrocytes (GFAP, green ), merged at ×10 and ×20 ( white rectangles ) magnification; c NeuN staining at ×20 magnification in core and contralateral striatum

    Article Snippet: Double immunohistochemical staining was performed against GFAP with rabbit anti-cow GFAP (Dako, 1:400) and CD11b (OX42) with mouse anti-rat CD11b (Serotec, 1:1000).

    Techniques: Staining, Positive Control