Journal: Neural plasticity

Article Title: AAV-KLF7 Promotes Descending Propriospinal Neuron Axonal Plasticity after Spinal Cord Injury.

doi: 10.1155/2017/1621629

Figure Lengend Snippet: Figure 2: Efficacy of AAV2-KLF7 gene transfer to spinal cord neurons in vitro. (a) AAV-GFP-infected cultured spinal cord neurons. Scale bars: 50 μm. (b) Quantitative analysis of AAV2-GFP infection rate. Representative immunohistochemical staining of KLF7 (green) and β- III-tubulin (labeling neurite and dendrite outgrowth, red) with nuclear staining (DAPI, blue) in cultured spinal cord neurons in the AAV- GFP (c) and AAV-KLF7 groups (d). Block shows the KLF7 colabeling of nuclei and neurite outgrowth. Quantitative analyses of the KLF7 relative density (e) and neurite length/neuron (f) in the two groups. n = 6 wells/group. Student’s t-tests, ∗∗∗P < 0 001. Scale bars: 200 μm.

Article Snippet: Tissues were stained with antibodies against KLF7 (1 : 200; Novus Biologicals, Colorado, USA), peripheral myelin or P0 (1 : 200; Sigma-Aldrich, Missouri, USA), GFAP to identify astrocytes (1 : 200; Sigma-Aldrich), BDA to identify astrocytes (1 : 200; Sigma-Aldrich), SYP to identify presynaptic components Week SC I A A V -K LF 7 ‒1 0 1 2 3 5 Experimental timeline Weekly BMS testing Immunohistochemistry, Western blot, TEM, grid walking, tcMMEP 4 Ba se lin e FG tr ac in g BD A & CT B tr ac in g A ni m al Sa cr ifi ce Figure 1: Experimental timeline.

Techniques: In Vitro, Infection, Cell Culture, Immunohistochemical staining, Staining, Labeling, Blocking Assay

Journal: Neural plasticity

Article Title: AAV-KLF7 Promotes Descending Propriospinal Neuron Axonal Plasticity after Spinal Cord Injury.

doi: 10.1155/2017/1621629

Figure Lengend Snippet: Figure 3: AAV-KLF7 significantly increased the expression of KLF7 and target proteins in vitro. (a) Representative Western blot analysis of KLF7, NGF, and TrkA expression in the cultured spinal cord neurons of the three groups is shown at 4 days in vitro (6 wells/each group). The graphs represent the relative density of KLF7 (b), NGF (c), and TrkA (d) protein. (e) Representative RT-PCR analysis of KLF7, NGF, and TrkA expression in the cultured spinal cord neurons of the three groups is shown at 4 days in vitro (n = 6). The graphs represent the relative density of KLF7 (f), NGF (g), and TrkA (h) mRNA. Error bars denote SD. ∗P < 0 05, ∗∗P < 0 01, and ∗∗∗P < 0 001. NS: not significant. One-way ANOVA, Tukey’s post hoc test.

Article Snippet: Tissues were stained with antibodies against KLF7 (1 : 200; Novus Biologicals, Colorado, USA), peripheral myelin or P0 (1 : 200; Sigma-Aldrich, Missouri, USA), GFAP to identify astrocytes (1 : 200; Sigma-Aldrich), BDA to identify astrocytes (1 : 200; Sigma-Aldrich), SYP to identify presynaptic components Week SC I A A V -K LF 7 ‒1 0 1 2 3 5 Experimental timeline Weekly BMS testing Immunohistochemistry, Western blot, TEM, grid walking, tcMMEP 4 Ba se lin e FG tr ac in g BD A & CT B tr ac in g A ni m al Sa cr ifi ce Figure 1: Experimental timeline.

Techniques: Expressing, In Vitro, Western Blot, Cell Culture, Reverse Transcription Polymerase Chain Reaction

Journal: Neural plasticity

Article Title: AAV-KLF7 Promotes Descending Propriospinal Neuron Axonal Plasticity after Spinal Cord Injury.

doi: 10.1155/2017/1621629

Figure Lengend Snippet: Figure 5: KLF7 expression is dynamically altered following SCI. (a) Western blot analysis revealed the endogenous change in KLF7 expression in the thoracic spinal cord injury tissue after SCI. (b) Graphs represent the KLF7 relative density of the bands normalized to GAPDH across different time points. n = 6, error bars denote SD. ∗P < 0 05, ∗∗∗P < 0 001 versus the baseline. One-way ANOVA, Tukey’s post hoc test.

Article Snippet: Tissues were stained with antibodies against KLF7 (1 : 200; Novus Biologicals, Colorado, USA), peripheral myelin or P0 (1 : 200; Sigma-Aldrich, Missouri, USA), GFAP to identify astrocytes (1 : 200; Sigma-Aldrich), BDA to identify astrocytes (1 : 200; Sigma-Aldrich), SYP to identify presynaptic components Week SC I A A V -K LF 7 ‒1 0 1 2 3 5 Experimental timeline Weekly BMS testing Immunohistochemistry, Western blot, TEM, grid walking, tcMMEP 4 Ba se lin e FG tr ac in g BD A & CT B tr ac in g A ni m al Sa cr ifi ce Figure 1: Experimental timeline.

Techniques: Expressing, Western Blot

Journal: Neural plasticity

Article Title: AAV-KLF7 Promotes Descending Propriospinal Neuron Axonal Plasticity after Spinal Cord Injury.

doi: 10.1155/2017/1621629

Figure Lengend Snippet: Figure 6: AAV-KLF7 significantly increased the expression of KLF7 and target proteins NGF, TrkA, GAP43, and P0 following SCI. (a) Representative Western blot analyses of KLF7, NGF, TrkA, GAP43, and P0 expression in thoracic spinal cord section tissues following SCI across three groups. The graphs for the analyses represent the relative density of KLF7 (b), NGF (c), TrkA (d), GAP43 (e), and P0 (f). n = 6 mice/group. Error bars denote SD. ∗P < 0 05, ∗∗P < 0 01, and ∗∗∗P < 0 001. NS: not significant. One-way ANOVA, Tukey’s post hoc test.

Article Snippet: Tissues were stained with antibodies against KLF7 (1 : 200; Novus Biologicals, Colorado, USA), peripheral myelin or P0 (1 : 200; Sigma-Aldrich, Missouri, USA), GFAP to identify astrocytes (1 : 200; Sigma-Aldrich), BDA to identify astrocytes (1 : 200; Sigma-Aldrich), SYP to identify presynaptic components Week SC I A A V -K LF 7 ‒1 0 1 2 3 5 Experimental timeline Weekly BMS testing Immunohistochemistry, Western blot, TEM, grid walking, tcMMEP 4 Ba se lin e FG tr ac in g BD A & CT B tr ac in g A ni m al Sa cr ifi ce Figure 1: Experimental timeline.

Techniques: Expressing, Western Blot

Journal: Neural plasticity

Article Title: AAV-KLF7 Promotes Descending Propriospinal Neuron Axonal Plasticity after Spinal Cord Injury.

doi: 10.1155/2017/1621629

Figure Lengend Snippet: Figure 10: AAV-KLF7 promotes DPSN axonal plasticity and induced synapse formation with motor neurons at the caudal spinal cord. (a) Contact between motoneurons (CTB retrogradely labeled, red) and DPSN axons (BDA-labeled, green) and synapse formation (synaptophysin-labeled, purple) are shown. Block indicates the new synapses formation (white) between DPSN axons and motoneurons. Quantitative analyses of the motor neuron targeted CTB-labeled axons (b), BDA relative density (c), and SYP density per motoneuron per section (d) at the caudal spinal cord. n = 6 mice/group. ∗P < 0 05, ∗∗P < 0 01, and ∗∗∗P < 0 001. NS: not significant. One-way ANOVA, Tukey’s post hoc test. Scale bars: 100 μm.

Article Snippet: Tissues were stained with antibodies against KLF7 (1 : 200; Novus Biologicals, Colorado, USA), peripheral myelin or P0 (1 : 200; Sigma-Aldrich, Missouri, USA), GFAP to identify astrocytes (1 : 200; Sigma-Aldrich), BDA to identify astrocytes (1 : 200; Sigma-Aldrich), SYP to identify presynaptic components Week SC I A A V -K LF 7 ‒1 0 1 2 3 5 Experimental timeline Weekly BMS testing Immunohistochemistry, Western blot, TEM, grid walking, tcMMEP 4 Ba se lin e FG tr ac in g BD A & CT B tr ac in g A ni m al Sa cr ifi ce Figure 1: Experimental timeline.

Techniques: Labeling, Blocking Assay

Journal: Neural plasticity

Article Title: AAV-KLF7 Promotes Descending Propriospinal Neuron Axonal Plasticity after Spinal Cord Injury.

doi: 10.1155/2017/1621629

Figure Lengend Snippet: Figure 11: AAV-KLF7 promotes axon myelination around the lesion. Transmission electron microscope (TEM) analysis was used to assess remyelination of spinal axons across the sham (a), SCI + AAV-GFP (b), and SCI + AAV-KLF7 groups (c) (n = 10). Quantitative analyses of the myelinated ratio (d). ∗P < 0 05, ∗∗∗P < 0 001, one-way ANOVA, Tukey’s post hoc test. Bar indicates 1 μm.

Article Snippet: Tissues were stained with antibodies against KLF7 (1 : 200; Novus Biologicals, Colorado, USA), peripheral myelin or P0 (1 : 200; Sigma-Aldrich, Missouri, USA), GFAP to identify astrocytes (1 : 200; Sigma-Aldrich), BDA to identify astrocytes (1 : 200; Sigma-Aldrich), SYP to identify presynaptic components Week SC I A A V -K LF 7 ‒1 0 1 2 3 5 Experimental timeline Weekly BMS testing Immunohistochemistry, Western blot, TEM, grid walking, tcMMEP 4 Ba se lin e FG tr ac in g BD A & CT B tr ac in g A ni m al Sa cr ifi ce Figure 1: Experimental timeline.

Techniques: Transmission Assay, Microscopy

Journal: Neural plasticity

Article Title: AAV-KLF7 Promotes Descending Propriospinal Neuron Axonal Plasticity after Spinal Cord Injury.

doi: 10.1155/2017/1621629

Figure Lengend Snippet: Figure 12: Cellular localization of KLF7 expression in the mouse spinal cord after spinal cord injury (SCI). (a) Little KLF7 immunoreactivity (IR) was localized in oligodendrocytes indicated by CC1 IR (arrows). (b) KLF7 immunoreactivity (IR) was localized in oligodendrocyte precursors indicated by NG2 IR (arrows). (c) No KLF7 immunoreactivity (IR) was localized in astrocytes indicated by GFAP IR (arrows). Scale bars: 100 μm.

Article Snippet: Tissues were stained with antibodies against KLF7 (1 : 200; Novus Biologicals, Colorado, USA), peripheral myelin or P0 (1 : 200; Sigma-Aldrich, Missouri, USA), GFAP to identify astrocytes (1 : 200; Sigma-Aldrich), BDA to identify astrocytes (1 : 200; Sigma-Aldrich), SYP to identify presynaptic components Week SC I A A V -K LF 7 ‒1 0 1 2 3 5 Experimental timeline Weekly BMS testing Immunohistochemistry, Western blot, TEM, grid walking, tcMMEP 4 Ba se lin e FG tr ac in g BD A & CT B tr ac in g A ni m al Sa cr ifi ce Figure 1: Experimental timeline.

Techniques: Expressing

Journal: Neural plasticity

Article Title: AAV-KLF7 Promotes Descending Propriospinal Neuron Axonal Plasticity after Spinal Cord Injury.

doi: 10.1155/2017/1621629

Figure Lengend Snippet: Figure 14: AAV-KLF7 significantly enhanced motor functional recovery following SCI. (a) Quantitative analyses of the Basso Mouse Scale (BMS) locomotor scores post-SCI (b), number of foot drops during grid walking five weeks postinjury (c), and tcMMEP responses five weeks postinjury (d), including the amplitude (mV) (e) and latency period (ms) from each group (n = 8), are shown. Error bars denote SD. ∗P < 0 05, ∗∗P < 0 01, and ∗∗∗P < 0 001, one-way ANOVA, Tukey’s post hoc test.

Article Snippet: Tissues were stained with antibodies against KLF7 (1 : 200; Novus Biologicals, Colorado, USA), peripheral myelin or P0 (1 : 200; Sigma-Aldrich, Missouri, USA), GFAP to identify astrocytes (1 : 200; Sigma-Aldrich), BDA to identify astrocytes (1 : 200; Sigma-Aldrich), SYP to identify presynaptic components Week SC I A A V -K LF 7 ‒1 0 1 2 3 5 Experimental timeline Weekly BMS testing Immunohistochemistry, Western blot, TEM, grid walking, tcMMEP 4 Ba se lin e FG tr ac in g BD A & CT B tr ac in g A ni m al Sa cr ifi ce Figure 1: Experimental timeline.

Techniques: Functional Assay

Journal: International Journal of Molecular Sciences

Article Title: Differential Expression of Peroxisomal Proteins in Distinct Types of Parotid Gland Tumors

doi: 10.3390/ijms22157872

Figure Lengend Snippet: List of antibodies used for the detection of peroxisomal proteins in the parotid gland.

Article Snippet: Peroxin 6 (PEX6p), , Rabbit, polyclonal , 104 kDa , , 1:1000 , Novus Biologicals, cat. no.: NBP1-80955.

Techniques: Molecular Weight, Phospho-proteomics, Marker

Journal: bioRxiv

Article Title: KDM2B is a histone H3K79 demethylase and induces transcriptional repression via SIRT1-mediated chromatin silencing

doi: 10.1101/228379

Figure Lengend Snippet: KDM2B demethylates histone H3K79me2/3. (A) Effects of overexpression and knockdown of KDM2B on H3K79 demethylation. FLAG-tagged KDM2B wild-type and catalytically deficient H242A mutant were transfected into 293T cells. H3K79me1/2/3 levels were detected by western blotting. KDM2B knockdown stable 293T cells were analyzed in the same way. (B) Immunocytochemistry of 293T cells expressing FLAG-KDM2B wild-type and KDM2B-H242A. Changes in signal intensity were detected after double immunostaining with anti-FLAG and anti-H3K79me1/3 antibodies. Arrows indicate transfected cells. (C) KDM2B in vitro demethylase assay. Upper: Core histones and nucleosomes were used as substrates. Demethylation activity of KDM2B against di- and tri-methyl H3K79 is shown. Lower: Purified GST and GST-KDM2B 1–734 recombinant proteins used were visualized by Coomassie staining. (D) Formaldehyde production was detected by FDH assay. NADH production was measured at OD 340 nm after 3-hour reaction using GST-KDM2B together with recombinant MLA histone H3K79me3 as substrates. Absence of demethylase and GST were used as negative controls.

Article Snippet: The following antibodies were used: anti-FLAG, anti-H3K79me3 (Epigentek), anti-H3K79me1, goat anti-mouse IgG-h+l fluorescein isothiocyanate (FITC) conjugated (Bethyl Laboratories), Cy™3-conjugated AffiniPure goat anti-rabbit IgG (H+L) (Jackson Immunoresearch).

Techniques: Over Expression, Mutagenesis, Transfection, Western Blot, Immunocytochemistry, Expressing, Double Immunostaining, In Vitro, Activity Assay, Purification, Recombinant, Staining

Journal: bioRxiv

Article Title: KDM2B is a histone H3K79 demethylase and induces transcriptional repression via SIRT1-mediated chromatin silencing

doi: 10.1101/228379

Figure Lengend Snippet: KDM2B lowers H3K79me3 occupancy at target gene promoters. (A) ChIP-seq heat map covering 10 kb regions across H3K79me3 peaks. (B) H3K79me3 ChIP-seq signal normalized by input sample in the control (blue line) and shKDM2B cells (red line). (C) Change in H3K79me3 occupancy following KDM2B knockdown. (D) Input and H3K79me3 ChIP-seq profiles over two regions of the genome in control and shKDM2B cells. Below the sequencing traces, GATA4 and PDE3B genes are indicated. (E) H3K79me3 mark at PDE3B promoter in KDM2B knockdown cells analyzed by ChIP- qPCR. (F) qRT-PCR analysis for detection of target gene expression. KDM2B was depleted in 293T cells. (G) ChIP-qPCR analysis for measurement of KDM2B enrichment and H3K79me3 levels on target gene promoters. KDM2B was stably knocked down in 293T cells. Promoters of HOXA7 , MEIS1 , ALX1 , and GATA4 were immunoprecipitated and amplified using specific primers.

Article Snippet: The following antibodies were used: anti-FLAG, anti-H3K79me3 (Epigentek), anti-H3K79me1, goat anti-mouse IgG-h+l fluorescein isothiocyanate (FITC) conjugated (Bethyl Laboratories), Cy™3-conjugated AffiniPure goat anti-rabbit IgG (H+L) (Jackson Immunoresearch).

Techniques: ChIP-sequencing, Sequencing, Quantitative RT-PCR, Expressing, Stable Transfection, Immunoprecipitation, Amplification