Journal: Nucleus

Article Title: Nucleocytoplasmic shuttling of HIV-1 integrase is controlled by the viral Rev protein

doi: 10.4161/nucl.1.2.11300

Figure Lengend Snippet: (See opposite page). Transient nuclear localization of HIV-1 IN in virus-infected cells. (A) HeLa-P4 cells were infected with WT HIV-1, and IN (red), Rev (green), LEDGF/p75 (gray) were immunostained and DNA (blue) was stained with DAPI. (B) Sup-T1 cells were infected with WT HIV-1 and the cytoplasmic and nuclear fractions were analyzed by western blot to detect Rev, IN, LEDGF/p75 and actin. (C) Same as (A) but LEDGF/p75-knockdown cells (HeLa-P4/shp75Cl15) were used for infection. (D) Same as in (B) but with the use of LEDGF/p75-knockdown cells (Sup-T1/TL3). In (A and C), scale bar represents 10 µm. All other experimental details are as described in Materials and Methods. Arrow indicate the presence of intranuclear IN (magenta), Rev (cyan) molecules, Rev-IN complex (white) or cytoplasmic Rev-IN complex (orange-yellow). Areas which have been magnified are marked by yellow boxes.

Article Snippet: Briefly, after fixation, cells were blocked with 5% (w/v) IgG-free BSA (Jackson) in PBS for 60 min. For detection of HIV-1 IN and Rev and the host LEDGF/p75, the cells were incubated with 1:50 rabbit α-IN(NIH AIDS Research & Reference Reagent Program catalog number 758), 1:50 rat α-Rev 50 and 1:100 goat α-LEDGF/p75 (R&D Systems) at room temperature for 60 min each.

Techniques: Virus, Infection, Staining, Western Blot

Journal: Nucleus

Article Title: Nucleocytoplasmic shuttling of HIV-1 integrase is controlled by the viral Rev protein

doi: 10.4161/nucl.1.2.11300

Figure Lengend Snippet: (See opposite page). Rev-induced inhibition of IN nuclear import in HIV-1 infected cells. (A) HeLa-P4 cells were infected with ΔRev HIV-1, and IN (red), Rev (green) were immunostained and DNA (blue) was stained with DAPI (B) H9 cells were infected with ΔRev HIV-1 and the cytoplasmic and nuclear fractions were analyzed by western blot to detect Rev, IN, LEDGF/p75 and actin. (C) Same as in (A), but Rev-overexpressing HeLa-P4 cells were infected with WT HIV-1. (D) Same as in (B), but Rev-overexpressing H9 cells were infected with WT HIV-1. (E) HeLa-P4 cells were transfected with vectors bearing either the IN, Rev or both as described in Material and Methods. IN (red), Rev (green) were immunostained and DNA (blue) was stained with DAPI. In (A, C and E), scale bar represents 10 µm. All other experimental details are as described in Materials and Methods. Arrow indicate the presence of intranuclear IN (magenta), Rev (cyan) molecules or cytoplasmic Rev-IN complex (orange-yellow). Areas which have been magnified are marked by yellow boxes.

Article Snippet: Briefly, after fixation, cells were blocked with 5% (w/v) IgG-free BSA (Jackson) in PBS for 60 min. For detection of HIV-1 IN and Rev and the host LEDGF/p75, the cells were incubated with 1:50 rabbit α-IN(NIH AIDS Research & Reference Reagent Program catalog number 758), 1:50 rat α-Rev 50 and 1:100 goat α-LEDGF/p75 (R&D Systems) at room temperature for 60 min each.

Techniques: Inhibition, Infection, Staining, Western Blot, Transfection

Journal: Nucleus

Article Title: Nucleocytoplasmic shuttling of HIV-1 integrase is controlled by the viral Rev protein

doi: 10.4161/nucl.1.2.11300

Figure Lengend Snippet: Effect of the Rev- and IN-derived peptides on nuclear localization of viral IN. (A) HeLa-P4 cells were infected with WT HIV-1 in the presence of Rev 13–23 peptide, and IN (red), Rev (green) were immunostained and DNA (blue) was stained with DAPI (B) H9 cells were infected with WT HIV-1 in the presence of Rev 13–23 peptide and the cytoplasmic and nuclear fractions were analyzed by western blot to detect Rev, IN, LEDGF/p75 and actin. (C and D), same as in (A and B), respectively, but in the presence of INr-1. In (A and C), scale bar represents 10 µm. All other experimental details are as described in Materials and Methods. Arrow indicate the presence of intranuclear IN (magenta), Rev (cyan) molecules or cytoplasmic Rev-IN complex (orange-yellow). Areas which have been magnified are marked by yellow boxes.

Article Snippet: Briefly, after fixation, cells were blocked with 5% (w/v) IgG-free BSA (Jackson) in PBS for 60 min. For detection of HIV-1 IN and Rev and the host LEDGF/p75, the cells were incubated with 1:50 rabbit α-IN(NIH AIDS Research & Reference Reagent Program catalog number 758), 1:50 rat α-Rev 50 and 1:100 goat α-LEDGF/p75 (R&D Systems) at room temperature for 60 min each.

Techniques: Derivative Assay, Infection, Staining, Western Blot

Journal: Nucleus

Article Title: Nucleocytoplasmic shuttling of HIV-1 integrase is controlled by the viral Rev protein

doi: 10.4161/nucl.1.2.11300

Figure Lengend Snippet: Nuclear localization of viral IN and Rev in Rev M10 HIV-1-infected cells. (A) HeLa-P4 cells were infected with Rev M10 HIV-1, and IN (red), Rev (green) were immunostained and DNA (blue) was stained with DAPI. Scale bar represents 10 µm. (B) H9 cells were infected with Rev M10 HIV-1 and the cytoplasmic and nuclear fractions were analyzed by western blot to detect Rev, IN, LEDGF/p75 and actin. All other experimental details are as described in Materials and Methods. Arrow indicate the presence of intranuclear IN (magenta), Rev (cyan) molecules, Rev-IN complex (white) or cytoplasmic Rev-IN complex (orange-yellow). Areas which have been magnified are marked by yellow boxes.

Article Snippet: Briefly, after fixation, cells were blocked with 5% (w/v) IgG-free BSA (Jackson) in PBS for 60 min. For detection of HIV-1 IN and Rev and the host LEDGF/p75, the cells were incubated with 1:50 rabbit α-IN(NIH AIDS Research & Reference Reagent Program catalog number 758), 1:50 rat α-Rev 50 and 1:100 goat α-LEDGF/p75 (R&D Systems) at room temperature for 60 min each.

Techniques: Infection, Staining, Western Blot

Journal: Journal of Neuroscience

Article Title: Tumor Necrosis Factor Receptor 1 Is a Negative Regulator of Progenitor Proliferation in Adult Hippocampal Neurogenesis

doi: 10.1523/jneurosci.2723-06.2006

Figure Lengend Snippet: Figure1. DeletionofTNF-R1andTNF-R2differentiallyaffectsbasalhippocampalneurogen- esis. A, Number of new mature neurons (BrdU /NeuN ) in the dentate subgranular zone/ granulecelllayerat2weeksafter2weeksofdailyBrdUinjections.MeansSEM;n8,6,and 15 for wild-type, TNF-R1/, and TNF-R2/ mice, respectively. *p 0.05 compared with wild-type, one-way ANOVA with Bonferroni–Dunn post hoc test. B, Confocal images of a BrdU /NeuN cellshowingBrdUandNeuNimmunoreactivityseparatelyorasmergedimage. OrthogonalreconstructionsfromconfocalZ-seriesarepresentedasviewedinx–z(bottom)and y–z (right) planes.

Article Snippet: For immunocytochemistry, rat-derived progenitor cells were cultured on poly-L-ornithine/laminin-coated chamber slides, fixed with 4% PFA for 15 min, and stained with mouse anti-TNF-R1 and goat anti-TNF-R2 antibodies (Santa Cruz Biotechnology) overnight at 4°C.

Techniques:

Journal: Journal of Neuroscience

Article Title: Tumor Necrosis Factor Receptor 1 Is a Negative Regulator of Progenitor Proliferation in Adult Hippocampal Neurogenesis

doi: 10.1523/jneurosci.2723-06.2006

Figure Lengend Snippet: Figure 2. Relative distribution of different convulsive behaviors and EEG characteristics for wild-type and TNF-R1/, TNF-R2/, and TNF-R1/R2/ mice during the 2 h of self- sustained status epilepticus. Ratings are based on Racine’s (1972) scale. Grades 0–2 and 3–5 represent partial and generalized convulsions, respectively. Each segment depicts the mean percentageoftimespentexhibitingthebehavior.ExamplesoftypicalhippocampalEEGrecord- ings during SE are presented for each group.

Article Snippet: For immunocytochemistry, rat-derived progenitor cells were cultured on poly-L-ornithine/laminin-coated chamber slides, fixed with 4% PFA for 15 min, and stained with mouse anti-TNF-R1 and goat anti-TNF-R2 antibodies (Santa Cruz Biotechnology) overnight at 4°C.

Techniques:

Journal: Journal of Neuroscience

Article Title: Tumor Necrosis Factor Receptor 1 Is a Negative Regulator of Progenitor Proliferation in Adult Hippocampal Neurogenesis

doi: 10.1523/jneurosci.2723-06.2006

Figure Lengend Snippet: Figure 3. Deletion of TNF-R1 and TNF-R2 differentially affects hippocampal neurogenesis after status epilepticus. A, Number ofnewmatureneurons(BrdU /NeuN )inthedentateSGZ/GCLat4weeksafter1weekofdailyBrdUinjections,whichstarted 1 week after 2 h of electrically induced status epilepticus. Means SEM; n 6, 6, and 9 for wild-type, TNF-R1/, and TNF-R2/ mice, respectively. *p 0.05 compared with wild-type, one-way ANOVA with Bonferroni–Dunn post hoc test. B, Photomicrographs showing distribution of BrdU (red) and NeuN (green) cells in dentate gyrus. Some BrdU /NeuN cells (yellow) are depicted by arrows. Scale bar, 70 m.

Article Snippet: For immunocytochemistry, rat-derived progenitor cells were cultured on poly-L-ornithine/laminin-coated chamber slides, fixed with 4% PFA for 15 min, and stained with mouse anti-TNF-R1 and goat anti-TNF-R2 antibodies (Santa Cruz Biotechnology) overnight at 4°C.

Techniques:

Journal: Journal of Neuroscience

Article Title: Tumor Necrosis Factor Receptor 1 Is a Negative Regulator of Progenitor Proliferation in Adult Hippocampal Neurogenesis

doi: 10.1523/jneurosci.2723-06.2006

Figure Lengend Snippet: Figure 4. Deletion of both TNF-R1 and TNF-R2 increases hippocampal neurogenesis during basal conditions and after status epilepticus. A, C, Number of new mature neurons (BrdU /NeuN ) in the dentate SGZ/GCL of TNF-R1/R2/ mice at 2 weeks after 2 weeks of daily BrdU injections (A) and at 4 weeks after 1 week of daily BrdU injections, which started 1 week after 2 h of electrically induced status epilepticus (C). Means SEM; n 5 and 6, and 6 and 10 for wild-type and TNF-R1/R2/ mice, respectively. *p 0.05 compared with wild-type, one-way ANOVA with Bonferroni–Dunn post hoc test. B, Photomicrographs showingdistributionofBrdU (red)andNeuN (green)cellsindentategyrus.SomeBrdU /NeuN cells(yellow)aredepicted by arrows. Scale bar, 70 m.

Article Snippet: For immunocytochemistry, rat-derived progenitor cells were cultured on poly-L-ornithine/laminin-coated chamber slides, fixed with 4% PFA for 15 min, and stained with mouse anti-TNF-R1 and goat anti-TNF-R2 antibodies (Santa Cruz Biotechnology) overnight at 4°C.

Techniques:

Journal: Journal of Neuroscience

Article Title: Tumor Necrosis Factor Receptor 1 Is a Negative Regulator of Progenitor Proliferation in Adult Hippocampal Neurogenesis

doi: 10.1523/jneurosci.2723-06.2006

Figure Lengend Snippet: Figure 5. Deletion of TNF-R1 and TNF-R2 differentially affects hippocampal progenitor proliferation under basal conditions andafterstatusepilepticus.A–F,NumberofcellsinthedentateSGZexpressingtheproliferationmarkersBrdU(A,D),p-H3(B,E), andPCNA(C,F)inintactmice(A–C)andat1weekafterelectricallyinducedstatusepilepticus(D–F).BrdUhadbeeninjectedfour times with 2 h interval, and animals were perfused 2 h thereafter. Means SEM; n 5 and 7, 4 and 7, 5 and 7, and 5 and 7 for wild-type, TNF-R1/, TNF-R2/, and TNF-R1/R2/ mice, respectively. *p 0.05 compared with wild-type, one-way ANOVAwithBonferroni–Dunnposthoctest.G–I,PhotomicrographsshowingdistributionofBrdU (G),p-H3 (H),andPCNA

Article Snippet: For immunocytochemistry, rat-derived progenitor cells were cultured on poly-L-ornithine/laminin-coated chamber slides, fixed with 4% PFA for 15 min, and stained with mouse anti-TNF-R1 and goat anti-TNF-R2 antibodies (Santa Cruz Biotechnology) overnight at 4°C.

Techniques:

Journal: Journal of Neuroscience

Article Title: Tumor Necrosis Factor Receptor 1 Is a Negative Regulator of Progenitor Proliferation in Adult Hippocampal Neurogenesis

doi: 10.1523/jneurosci.2723-06.2006

Figure Lengend Snippet: Figure 6. Deletion of TNF-R1 and TNF-R2 affects microglia in neurogenic area after status epilepticus. A, B, Numbers of Iba1 immunoreactive cells (showing both active and quiescent microglia) in the dentate SGZ/GCL under basal conditions (A) and at 6 weeks after SE (B). Means SEM; n 11, 6, 10, and 10 for wild-type, TNF-R1/, TNF-R2/, and TNF-R1/R2/ mice, respectively. *p 0.05 compared with wild-type, one-way ANOVA with Bonferroni–Dunn post hoc test. C, Photomicrographs showing distribution of Iba1 cells in dentate hilus, SGZ, and GCL at 6 weeks after SE. Some Iba1 cells (black) are depicted by arrows. Scale bar, 70 m.

Article Snippet: For immunocytochemistry, rat-derived progenitor cells were cultured on poly-L-ornithine/laminin-coated chamber slides, fixed with 4% PFA for 15 min, and stained with mouse anti-TNF-R1 and goat anti-TNF-R2 antibodies (Santa Cruz Biotechnology) overnight at 4°C.

Techniques:

Journal: Journal of Neuroscience

Article Title: Tumor Necrosis Factor Receptor 1 Is a Negative Regulator of Progenitor Proliferation in Adult Hippocampal Neurogenesis

doi: 10.1523/jneurosci.2723-06.2006

Figure Lengend Snippet: Figure7. HippocampalprogenitorsexpressTNF-R1,TNF-R2,andTNF-.A,DetectionofTNF-,TNF-R1,andTNF-R2mRNAin sorted hippocampal progenitors from adult nestin–GFP mice and in adult rat hippocampal progenitor cultures (AHP) using RT-PCR. First lane is a DNA ladder to indicate size of amplified products. B–G, TNF-R1 (B) and TNF-R2 (E) immunoreactivity, Hoechststaining(C,F),andmergedimages(D,G)asobservedinculturesofadultrathippocampalprogenitors.Arrowsdepictone TNF-R1 (B–D) and one TNF-R2 (E–G) cell. Scale bar, 50 m.

Article Snippet: For immunocytochemistry, rat-derived progenitor cells were cultured on poly-L-ornithine/laminin-coated chamber slides, fixed with 4% PFA for 15 min, and stained with mouse anti-TNF-R1 and goat anti-TNF-R2 antibodies (Santa Cruz Biotechnology) overnight at 4°C.

Techniques: Reverse Transcription Polymerase Chain Reaction, Amplification

Journal: Oncotarget

Article Title: Cell activity during peripheral nerve defect repair process using a nerve scaffold

doi: 10.18632/oncotarget.22978

Figure Lengend Snippet: ( A 1) Autologous sciatic nerve at 1 week, immunofluorescence staining with DAPI (blue), integrin-α(red) and NGFR P75 (green). ( A 2) Autologous sciatic nerve at 2 weeks, immunofluorescence staining with DAPI (blue), integrin-α(red) and NGFR P75 (green). ( B 1) Nerve scaffold at 1 week, immunofluorescence staining with DAPI (blue), integrin-α(red) and fibronectin (green). ( B 2) Nerve scaffold at 2 weeks, immunofluorescence staining with DAPI (blue), integrin-α(red) and fibronectin (green). ( C 1) Longitudinal section of the proximal end of PLGA tube at 2 weeks (HE). ( C 2) Longitudinal section of the distal end of PLGA tube at 2 weeks (HE). ( C 3) A film preparation smeared with the content of PLGA tube at 2 weeks (HE). White arrows: The field of vision show in C3. Ba: Basophile. Eo: Eosinophile. Le: Leukocyte. Ly: Lymphocyte. Mo: Monocyte.

Article Snippet: Primary antibody for low-affinity nerve growth factor receptor (NGFR) P75 (goat anti-rat; Santa Cruz Biotechnology, USA) was used to identify Schwann cells.

Techniques: Immunofluorescence, Staining

Journal: Oncotarget

Article Title: Cell activity during peripheral nerve defect repair process using a nerve scaffold

doi: 10.18632/oncotarget.22978

Figure Lengend Snippet: ( A ) Autologous, PLGA tube and nerve scaffold at 3 weeks after implantation were stained with DAPI (blue), NF200 (red), and NGFR P75 (green). ( B ) PLGA tube and nerve scaffold at 3 weeks after implantation were stained with DAPI (blue), NF200 (red), and fibronectin (green). ( C ) Autologous , PLGA tube and nerve scaffold at 20 weeks after implantation were stained with DAPI (blue), NF200 (red), and NGFR P75 (green). ( D ) Autologous , PLGA tube and nerve scaffold at 20 weeks after implantation were stained with DAPI (blue), NF200 (red), and fibronectin (green). SF: Silk fibers.

Article Snippet: Primary antibody for low-affinity nerve growth factor receptor (NGFR) P75 (goat anti-rat; Santa Cruz Biotechnology, USA) was used to identify Schwann cells.

Techniques: Staining

Journal: bioRxiv

Article Title: Cholinergic basal forebrain neurons regulate vascular dynamics and cerebrospinal fluid flux

doi: 10.1101/2024.08.25.609536

Figure Lengend Snippet: a) An immunotoxin, p75-saporin, which targets the p75 neurotrophin receptor-expressing cholinergic neurons was injected into the medial septum (MS) of the mouse basal forebrain. This resulted in significant reduction of cholinergic neurons (green fluorescence) and their density in the MS compared to that of sham mice injected with the non-targeted IgG-saporin. b) 3-4 weeks after the surgery, resting-state fMRI was conducted using a 9.4T MRI to measure the cortical BOLD (dash line) and ventricular CSF inflow (solid line) signals. c) The cross-correlation between the cortical BOLD and CSF signals shows anticorrelation at 0 lag time in both sham and lesioned mice. However, the anticorrelation between the regional BOLD and CSF signals were abolished in the lesioned mice in the hippocampus, which receives the MS cholinergic projection, but not in the cortex. Error bars represent the SEM. d) The cortical BOLD signal coupling with the CSF signal (top) and fluctuation amplitude (bottom) were comparable between the lesioned and sham mice. e) In the hippocampus, however, the hippocampal BOLD-CSF coupling strength was reduced toward 0, with the BOLD signal amplitude showed a trend of decrease. f) The hippocampal BOLD-CSF coupling, BOLD signal amplitude, and the lag time of maximal anticorrelation between the BOLD and CSF signals, were all significantly correlated with the p75-positive cholinergic neuron density of the MS of the imaged mice.

Article Snippet: Immunofluorescence labeling of the cholinergic cells were stained with primary goat anti-p75 antibody (1:400, AF1157, R&D Systems) overnight at room temperature.

Techniques: Expressing, Injection, Fluorescence

Journal: Journal of Neuroscience

Article Title: Olfactory Ensheathing Cells Do Not Exhibit Unique Migratory or Axonal Growth-Promoting Properties after Spinal Cord Injury

doi: 10.1523/jneurosci.3264-06.2006

Figure Lengend Snippet: Figure 1. p75 immunolabeling of GFP-expressing OECs in vitro and in vivo. A–C, After passage 6, the majority of GFP- transduced OECs (A; green) express p75 (B, red; C, yellow). D, Double florescent immunolabeling shows that the majority of GFP-expressingOECsexpressp75,7daftertransplantation.E,xy,xz,andyxplaneimagesofatypicalGFPandp75double-labeled OEC taken from the central field of D. Scale bars: A–C, 35 m; D, 63 m; E, 10 m.

Article Snippet: After washes, sections were incubated either with Alexa 488, Alexa 594- or Cy5-conjugated donkey anti-mouse, donkey anti-rabbit, or donkey anti-goat secondary antibodies (1:150; Invitrogen) or biotinylated donkey anti-rabbit secondary antibody (for p75 labeling, 1:150; Vector Laboratories, Burlingame, CA) for 2.5 h at room temperature.

Techniques: Immunolabeling, Expressing, In Vitro, In Vivo, Labeling

Journal:

Article Title: p75 NTR Mediates Neurotrophin-Induced Apoptosis of Vascular Smooth Muscle Cells

doi:

Figure Lengend Snippet: Immunofluorescent analysis for expression of activated caspases in p75-TsTmSMC. Cells were counterstained with DAPI for detection of condensed chromatin as an index of apoptosis.

Article Snippet: Adjacent sections were then incubated with either the anti-p75 NTR antibody (goat polyclonal, Santa Cruz Biotechnology, Santa Cruz, CA), anti-full length trk B antibody (goat polyclonal; directed against the carboxy terminus of trk B); anti-truncated trk B antibody (rabbit polyclonal; Santa Cruz Biotechnology); anti-full length trk C antibody (rabbit polyclonal directed against carboxy terminus against trk C; Santa Cruz Biotechnology) anti-truncated trk C antisera (rabbit polyclonal 2 ), anti-smooth muscle cell α-actin antibody (monoclonal, Clone 1A4, Dako Corp., Carpinteria, CA), anti-CD31 antibody (Hec-7 26 kindly provided by Dr. William Muller, Weill Medical College of Cornell University) or anti-macrophage antibody, (HAM 56 mouse monoclonal; DAKO Corp.).

Techniques: Expressing

Journal:

Article Title: p75 NTR Mediates Neurotrophin-Induced Apoptosis of Vascular Smooth Muscle Cells

doi:

Figure Lengend Snippet: Western blot analysis of p75NTR, trk, and FAS in native TsTmSMC and p75-TsTmSMC. For FAS Western blot analysis African green monkey kidney cells and the murine lymphocyte cell line A20 are used as a negative and positive control, respectively. The pheochromocytoma cell line PC12, genetically manipulated to overexpress trk receptors, 28 was used as a positive control for trk and p75NTR Western blot analysis. This cell line expresses approximately 40,000 trk receptors and 40,000 p75NTR cell. Lane 1, TsTmSMC. Lane 2, A20 cells. Lane 3, COS cells. Lane 4, PC12. Lane 5, p75-TsTmSMC. Lane 6, p75-TsTmSMC-2.

Article Snippet: Adjacent sections were then incubated with either the anti-p75 NTR antibody (goat polyclonal, Santa Cruz Biotechnology, Santa Cruz, CA), anti-full length trk B antibody (goat polyclonal; directed against the carboxy terminus of trk B); anti-truncated trk B antibody (rabbit polyclonal; Santa Cruz Biotechnology); anti-full length trk C antibody (rabbit polyclonal directed against carboxy terminus against trk C; Santa Cruz Biotechnology) anti-truncated trk C antisera (rabbit polyclonal 2 ), anti-smooth muscle cell α-actin antibody (monoclonal, Clone 1A4, Dako Corp., Carpinteria, CA), anti-CD31 antibody (Hec-7 26 kindly provided by Dr. William Muller, Weill Medical College of Cornell University) or anti-macrophage antibody, (HAM 56 mouse monoclonal; DAKO Corp.).

Techniques: Western Blot, Positive Control