Journal: Frontiers in Aging Neuroscience

Article Title: Multimodal Approaches for Regenerative Stroke Therapies: Combination of Granulocyte Colony-Stimulating Factor with Bone Marrow Mesenchymal Stem Cells is Not Superior to G-CSF Alone

doi: 10.3389/fnagi.2014.00130

Figure Lengend Snippet: Post-stroke neurogenesis and angiogenesis . At 8 weeks post-stroke, none of the DCX + cells in the SVZ of control animals co-localized with BrdU-labeled nuclei. Instead, the BrdU-positive nuclei were distributed mainly in the “pinwheel” architecture of the ventricular epithelium (A) . The DCX + cells occupied an adjacent, distinct position [ (A) , arrows]. Some of the DCX + migrated away from the ventricular wall (B) . We noted vigorous neurogenesis with many DCX + (arrows) co-localizing with BrdU nuclei in the G-CSF-treated animals [ (C) ; arrowheads] and animals treated with G-CSF + BM MSC [ (D) , arrows]. (E–G) Post-stroke angiogenesis. In regions adjacent to the infarct scar, we found numerous BrdU + nuclei in the endothelium of newly formed blood vessels in the formerly infarct core [ (E) , green]. The border to the healthy brain region was abruptly demarcated to the left by NeuN-positive nuclei [ (E) , red]. Beyond the formerly infarct core, we noted vigorous sprouting angiogenesis as evidenced by RECA/BrdU double positive blood vessels [ (F) , violet] as well as numerous BrdU + nuclei in the newly formed endothelium [ (F) , blue] and reconstruction of the basal lamina [ (F) , green] during the resolution phase of angiogenesis. By number of laminin/BrdU co-localizations, the density of the newly formed blood vessels was significantly higher (threefold, p = 0.01) in the brains of animals treated with the combination G-CSF + BM MSC as compared to controls and G-CSF alone (G) . Cc, corpus callosum; IC, infarct core; IR, islet of regeneration; LV, lateral ventricle; PI, periinfract.

Article Snippet: For phenotyping in the periinfarcted area and beyond the fibrotic scar, sections were triple-immunolabeled with rabbit anti-laminin-specific antibodies (1:5000, Sigma, Munich), mouse anti-NeuN antibodies (1:500, Millipore, Germany), mouse anti-RECA (1:1000, Millipore, Germany), and rat anti-BrdU-specific antibodies (1:3000; Serotec, UK).

Techniques: Control, Labeling

Journal: Scientific Reports

Article Title: Jak/Stat signaling regulates the proliferation and neurogenic potential of Müller glia-derived progenitor cells in the avian retina

doi: 10.1038/srep35703

Figure Lengend Snippet: Antibodies, working dilutions, host and source.

Article Snippet: BrdU , 1:100 , mouse , G3G4 , Developmental Studies Hybridoma Bank (DSHB).

Techniques: Diagnostic Assay

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Nestin expression in repopulating mesangial cells promotes their proliferation.

doi: 10.1038/labinvest.2008.5

Figure Lengend Snippet: Figure 2 Colocalization studies confirm de novo expression of nestin by activated and proliferating MCs. Double immunostaining in kidney biopsies from anti-Thy1 nephritic rats on day 5 confirms that nestin (a, d, g, j, m, green fluorescence) colocalizes with the MC marker Ox-7 on days 3 (b, red fluorescence; c, yellow fluorescence) and 5 (e, red fluorescence; f, yellow fluorescence), the podocyte marker ezrin (h, red fluorescence; i, yellow fluorescence) and with the activation marker for MCs, a-sm-actin (k, red fluorescence; l, yellow fluorescence). In contrast, glomerular endothelial cells as identified by anti-RECA-1 antibody (n, red fluorescence) show no colocalization with nestin-positive cells (m, o, green fluorescence).

Article Snippet: Tissues were embedded in paraffin and cut into 3-mm sections for indirect immunoperoxidase staining or immunofluorescence as described elsewhere.21,26 The following antibodies were used in this study: nestin, clone rat 401 a murine IgG1 mAb specific for rat and murine nestin (Chemicon International, Temecula, CA, USA); nestin, clone 4D4 a murine IgG1 mAB specific for murine nestin (Biotrend, Köln, Germany); nestin, a murine IgG1 mAb specific for human nestin (Chemicon); OX-7, a murine IgG1 mAb specific for rat CD90 expressed on MCs (Serotec Ltd, Oxford, UK);21 a-smooth muscle actin (a-sm-actin), a murine IgG2 mAb specific for activated MCs (Dako, Hamburg, Germany);27 RECA-1, a murine IgG mAb specific for rat endothelial cells (Serotec Ltd);28,29 ezrin, a murine IgG1 mAb specific for ezrin (Labvision Corporation, Fremont, CA, USA); and26 proliferating-cell nuclear antigen (PCNA), a murine IgG1 mAb (clone PC10) specific for the PCNA (Dako, Glostrup, Denmark).

Techniques: Expressing, Double Immunostaining, Fluorescence, Marker, Activation Assay

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Nestin expression in repopulating mesangial cells promotes their proliferation.

doi: 10.1038/labinvest.2008.5

Figure Lengend Snippet: Figure 5 Nestin is differentially expressed during cell cycle and promotes cell proliferation. Double immunostaining in kidney biopsies from anti-Thy1 nephritic rats on days 3 and 5 for nestin (brown color) and the proliferation marker PCNA (green nuclei) demonstrates that most nestin-positive cells in a typical mesangial position are also proliferating (a, day 3, arrows; b, day 5). After staining of DNA in cultured MCs by propidium iodide, cell-cycle analysis was performed by flow cytometry analysis (c). In parallel, MCs were stained for nestin for cell-cycle phase-dependent analysis (d). The percentage of cells expressing high levels of nestin were determined during G1-, S- and G2-phases, and demonstrated increased nestin expression during S- and G2-phases (e). # Indicates statistic significant changes between G1- and S- phases (Po0.008) and G2-phase (n ¼ 4, Po0.0001) tested by two-way ANOVA analysis. Transfection of MCs with siRNA-targeting nestin successfully downregulated nestin expression as demonstrated by flow cytometry analysis (f, g, n ¼ 4; Po0.0001). MC proliferation, as measured by BrdU incorporation (h) or cell-cycle analysis (i), was significantly reduced in MCs after transfection with nestin siRNA compared with MCs transfected with nonspecific control siRNA. * Indicates statistical significant changes between cells treated with transfection reagent only and siRNA-treated cells tested by two-way ANOVA analysis (n ¼ 4; Po0.05).

Article Snippet: Tissues were embedded in paraffin and cut into 3-mm sections for indirect immunoperoxidase staining or immunofluorescence as described elsewhere.21,26 The following antibodies were used in this study: nestin, clone rat 401 a murine IgG1 mAb specific for rat and murine nestin (Chemicon International, Temecula, CA, USA); nestin, clone 4D4 a murine IgG1 mAB specific for murine nestin (Biotrend, Köln, Germany); nestin, a murine IgG1 mAb specific for human nestin (Chemicon); OX-7, a murine IgG1 mAb specific for rat CD90 expressed on MCs (Serotec Ltd, Oxford, UK);21 a-smooth muscle actin (a-sm-actin), a murine IgG2 mAb specific for activated MCs (Dako, Hamburg, Germany);27 RECA-1, a murine IgG mAb specific for rat endothelial cells (Serotec Ltd);28,29 ezrin, a murine IgG1 mAb specific for ezrin (Labvision Corporation, Fremont, CA, USA); and26 proliferating-cell nuclear antigen (PCNA), a murine IgG1 mAb (clone PC10) specific for the PCNA (Dako, Glostrup, Denmark).

Techniques: Double Immunostaining, Marker, Staining, Cell Culture, Cell Cycle Assay, Flow Cytometry, Expressing, Transfection, BrdU Incorporation Assay, Control

Journal: The Journal of Neuroscience

Article Title: Neonatal and Adult Neurogenesis Provide Two Distinct Populations of Newborn Neurons to the Mouse Olfactory Bulb

doi: 10.1523/JNEUROSCI.1114-05.2005

Figure Lengend Snippet: Targeting of newborn granule cells changes during bulbar development. a, e, Quantifying BrdU+ cell distribution throughout the internal-external (a) and ventrodorsal (e) axes of the granule cell layer (see Materials and Methods for details). EPL, External plexiform layer; GL, glomerular layer. b, c, Quantification of BrdU+ cells throughout the internal-external axis expressed as percentage (b) and density (c). Note the different age-dependent distribution profiles of BrdU+ cells. d, Internal-external distribution of BrdU+ cells measured after short-term (2 h; black circles) or long-term (from 20 to 28 d, depending on postnatal age of animals; white circles) survival. f, g, Distribution of BrdU+ cells throughout the ventrodorsal axis expressed as percentage (f) and density (g). Note the equal distribution of BrdU+ profiles across ages. Scale bars, 200 μm.

Article Snippet: The primary antibodies used were rat monoclonal anti-BrdU antibody (1:200; Harlan Sera-Lab, Loughborough, UK) and mouse monoclonal anti- neuronal-specific nuclear protein (NeuN) antibody (1:200; Chemicon, Temecula, CA).

Techniques:

Journal: The Journal of Neuroscience

Article Title: Neonatal and Adult Neurogenesis Provide Two Distinct Populations of Newborn Neurons to the Mouse Olfactory Bulb

doi: 10.1523/JNEUROSCI.1114-05.2005

Figure Lengend Snippet: Local cellular proliferation in the postnatal olfactory bulb decreases with age. a-e, Representative images of BrdU+ cells located in the granule cell layer of 3-d-old (a), 7-d-old (b), 14-d-old (c), 21-d-old (d), and 60-d-old (e) mice. f, Mean density of BrdU+ cells (per mm2) in the granule cell layer determined 2 h after a single injection of BrdU (n = 2-4 animals per group). Scale bar, 50 μm.

Article Snippet: The primary antibodies used were rat monoclonal anti-BrdU antibody (1:200; Harlan Sera-Lab, Loughborough, UK) and mouse monoclonal anti- neuronal-specific nuclear protein (NeuN) antibody (1:200; Chemicon, Temecula, CA).

Techniques: Injection

Journal: The Journal of Neuroscience

Article Title: Neonatal and Adult Neurogenesis Provide Two Distinct Populations of Newborn Neurons to the Mouse Olfactory Bulb

doi: 10.1523/JNEUROSCI.1114-05.2005

Figure Lengend Snippet: Age-dependent changes in the migration duration required for neuroblasts to reach the OB. a, Representative images of a sagittal section of a 19-d-old mouse brain demonstrating the pattern of distribution of BrdU+ cells generated at P3 along the SVZ/OB pathway. LV, Lateral ventricle. b, Coronal sections of the OB of animals injected with BrdU at P3, P7, P14, P21, and P60 and killed following different survival periods as indicated in the parentheses. The coronal sections were taken at the level of the dotted line indicated in the sagittal section in a. Note the disappearance of BrdU+ cells in the RMSOB following the chosen survival times. c, Ratio of BrdU+ cells in the RMSOB to those in the OB. Scale bars: a, 500 μm; b, 50 μm.

Article Snippet: The primary antibodies used were rat monoclonal anti-BrdU antibody (1:200; Harlan Sera-Lab, Loughborough, UK) and mouse monoclonal anti- neuronal-specific nuclear protein (NeuN) antibody (1:200; Chemicon, Temecula, CA).

Techniques: Migration, Generated, Injection

Journal: The Journal of Neuroscience

Article Title: Neonatal and Adult Neurogenesis Provide Two Distinct Populations of Newborn Neurons to the Mouse Olfactory Bulb

doi: 10.1523/JNEUROSCI.1114-05.2005

Figure Lengend Snippet: Age-dependent decreases in the density of surviving newly generated cells in the granule cell layer. a, Mean number of BrdU+ cells (per mm2) in the granule cell layer at different ages. Animals (n = 3-5 per group) were killed following a time window required to empty the RMS (as indicated in Fig. 2). b, Mean number of BrdU+ cells (per mm2) throughout the rostrocaudal axis of the OB. c, Ratio of SVZ/RMS-generated cells in the OB after migration versus that of locally proliferating cells at different ages. Note that, because of the decreased local OB proliferation with age, this ratio increases during postnatal development.

Article Snippet: The primary antibodies used were rat monoclonal anti-BrdU antibody (1:200; Harlan Sera-Lab, Loughborough, UK) and mouse monoclonal anti- neuronal-specific nuclear protein (NeuN) antibody (1:200; Chemicon, Temecula, CA).

Techniques: Generated, Migration

Journal: The Journal of Neuroscience

Article Title: Neonatal and Adult Neurogenesis Provide Two Distinct Populations of Newborn Neurons to the Mouse Olfactory Bulb

doi: 10.1523/JNEUROSCI.1114-05.2005

Figure Lengend Snippet: Cell death and fate of newborn cells remain constant in the olfactory bulb at different postnatal ages. a, TUNEL+ cells (arrows) in the middle of granule cell layer of 1-month-old (left) and 3-month-old (right) mice. b, Number of apoptotic nuclei (per mm2) in the granule cell layer of young and adult mice. c, Distribution of TUNEL+ cells (expressed as percentage) within the internal-external axis of the granule cell layer of 1- and 3-month-old animals (n = 4 per group). d, Example of a 3D-reconstructed BrdU+ cell (red) colabeled with the neuronal marker NeuN (green) in the granule cell layer. Reconstructed orthogonal projections are presented as viewed in the x-z (top) and y-z (right) planes. e, Percentage of double-labeled cells in the granule cell layer of 7-, 14-, 21-, and 60-d-old mice. More than 1000 cells, taken from three animals per group, were analyzed for each postnatal age. Scale bars: a, 20 μm; d, 10 μm. Error bars indicate SEM.

Article Snippet: The primary antibodies used were rat monoclonal anti-BrdU antibody (1:200; Harlan Sera-Lab, Loughborough, UK) and mouse monoclonal anti- neuronal-specific nuclear protein (NeuN) antibody (1:200; Chemicon, Temecula, CA).

Techniques: TUNEL Assay, Marker, Labeling

Journal: The Journal of Neuroscience

Article Title: Neonatal and Adult Neurogenesis Provide Two Distinct Populations of Newborn Neurons to the Mouse Olfactory Bulb

doi: 10.1523/JNEUROSCI.1114-05.2005

Figure Lengend Snippet: Different survival rates of newborn granule cells in the neonate and the adult olfactory bulb. a, Photomicrographs of BrdU-immunostained sections of the granule cell layer in early postnatal (top; P4) and adult mice (bottom; P45) 22 d (left) and 60 d (right) after BrdU injections. b, Corresponding densities of BrdU+ cells generated during different periods of life and allowed to survive for short (22 d) and long (60 d) BrdU postinjection times (n = 4-10 animals per group). Note the differences in age-dependent persistence of newborn cells in young and adult mice. c, Distribution of early- and late-generated GCs following different survival periods. Note the distribution differences between early postnatal (P4) and adult animals (P45) versus the similar distributions observed following different survival periods. **p < 0.001. Scale bar, 20 μm. Error bars indicate SEM.

Article Snippet: The primary antibodies used were rat monoclonal anti-BrdU antibody (1:200; Harlan Sera-Lab, Loughborough, UK) and mouse monoclonal anti- neuronal-specific nuclear protein (NeuN) antibody (1:200; Chemicon, Temecula, CA).

Techniques: Generated

Journal: The Journal of Neuroscience

Article Title: Neonatal and Adult Neurogenesis Provide Two Distinct Populations of Newborn Neurons to the Mouse Olfactory Bulb

doi: 10.1523/JNEUROSCI.1114-05.2005

Figure Lengend Snippet: Early olfactory experience alters early but not late postnatal neurogenesis. a, Boxed area, Pups were reared in a citral- or saline-scented environment from E18 to P9. Top graph, P8 and P60 mice were subjected to a preference test. Histograms indicate the ratio of time spent by saline- and citral-reared animals (black and white bars, respectively) investigating citral or carvon shavings versus clean shavings (n = 9-33 animals per group). At both ages, animals show a significant preference to citral compared with saline. The specificity of this preference is shown by the absence of preference to carvon shavings at any age. Bottom graph, Quantification of BrdU+ cells following early olfactory experience. BrdU was administered either from P4 to P7 (one injection per day) or at P45 (four injections), and animals were killed for immunohistochemistry 20 d later. The mean number of newborn GCs was significantly higher in citral-reared animals compared with control mice when measured at P27 but not at P65. b, Spatial distribution of BrdU+ cells counted in the granule cell layer along the rostrocaudal axis (left panel), the internal-external axis (middle panel), and the ventrodorsal axis (right panel) of the OB taken from saline- or citral-reared mice. Animals were injected with BrdU from P4 to P7 (n = 5-7 animals per group). ***p < 0.001 with a Student's t test.

Article Snippet: The primary antibodies used were rat monoclonal anti-BrdU antibody (1:200; Harlan Sera-Lab, Loughborough, UK) and mouse monoclonal anti- neuronal-specific nuclear protein (NeuN) antibody (1:200; Chemicon, Temecula, CA).

Techniques: Saline, Injection, Immunohistochemistry, Control