Journal: Molecular neurobiology

Article Title: Piezo2 Channel Upregulation is Involved in Mechanical Allodynia in CYP-Induced Cystitis Rats.

doi: 10.1007/s12035-023-03386-9

Figure Lengend Snippet: Fig. 4 Upregulation of Piezo2 protein expression in L6-S1 DRG neurons in CYP rats. (A) Repre sentative western blots of protein expression of Piezo2 channels. Total membrane proteins were extracted from bilateral L6-S1 DRGs from four group of rats. β-actin (42 kDa) was used as the control. (B) Summary western blot data showing upregulated protein expression of Piezo2 in L6-S1 DRGs in CYP rats; and this increase was significantly reduced in antisense + CYP rat (two-way ANOVA, p < 0.01). Data are presented as mean ± SEM. n is the number of rats in each group. ** p < 0.01

Article Snippet: In the current study, we have further shown that preadsorption of the Piezo2 antibody blocking peptide (Novus (containing 1% Triton-X100, 2% bovine serum albumin [BSA], and 4% normal goat serum in phosphate-buffered saline) for 4 h. The tissues were then incubated with rabbit anti-Piezo2 (diluted at 1:100 in phosphate-buffered saline, APC090; Alomone Labs, Jerusalem, Israel) and neurofilament 200 (NF200) (1:200; #2836; Cell Signaling Technology, United States) or mouse anti-calcitonin gene-related peptide (anti-CGRP) (1:100, ab81887; Abcam, Cambridge, UK), TRPV1 (1:100, 66983-1-Ig; Proteintech, China), or isolectin B4 (IB4) (diluted at 10 μg/mL in phosphatebuffered saline, DL-1207; Vector Laboratories, Burlingame, CA, USA) for almost 40 h at 4 °C.

Techniques: Expressing, Western Blot, Membrane, Control

Journal: Molecular neurobiology

Article Title: Piezo2 Channel Upregulation is Involved in Mechanical Allodynia in CYP-Induced Cystitis Rats.

doi: 10.1007/s12035-023-03386-9

Figure Lengend Snippet: Fig. 6 Knockdown Piezo2 expression in sensory neurons attenuated mechanical allodynia in CYP rats. Mechanical allodynia in lower abdominal region was assessed using a series of von Frey filaments (0.04-2 g) on the day after the third CYP injection. CYP rats and CYP + mismatch rats showed an increased response to mechanical stimulation (> 0.4 g) compared with control rats, and these increased responses were significantly reduced in antisence + CYP rats (Two- way repeated measure ANOVA, p < 0.05). *p < 0.05 compared with control, #p < 0.05 compared with CYP + Mismatch. n indicates the number of rats

Article Snippet: In the current study, we have further shown that preadsorption of the Piezo2 antibody blocking peptide (Novus (containing 1% Triton-X100, 2% bovine serum albumin [BSA], and 4% normal goat serum in phosphate-buffered saline) for 4 h. The tissues were then incubated with rabbit anti-Piezo2 (diluted at 1:100 in phosphate-buffered saline, APC090; Alomone Labs, Jerusalem, Israel) and neurofilament 200 (NF200) (1:200; #2836; Cell Signaling Technology, United States) or mouse anti-calcitonin gene-related peptide (anti-CGRP) (1:100, ab81887; Abcam, Cambridge, UK), TRPV1 (1:100, 66983-1-Ig; Proteintech, China), or isolectin B4 (IB4) (diluted at 10 μg/mL in phosphatebuffered saline, DL-1207; Vector Laboratories, Burlingame, CA, USA) for almost 40 h at 4 °C.

Techniques: Knockdown, Expressing, Injection, Control

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 1: Expression of p33ING1b and p29ING4 in renal cell carcinoma. (a) mRNA expression of p33ING1b and p29ING4 was increased at early and late stages of disease with higher expression of both genes at late stages of disease compared to early stages (Robson stages III/IV vs. I/II) (p33ING1b and p29ING4: stages I/II and III/IV vs. normal tissue p<0.001 and p<0.0001, respectively; p33ING1b and p29ING4: stages III/IV vs. I/II p<0.01 and p<0.001, respectively). (b) Results of the real time RT-PCR were normalized to kidney normal tissue and expressed as x-fold difference (2- ∆∆Ct). Expression of p33ING1b and p29ING4 by renal carcinoma cells was confirmed (Cy3 red; DAPI blue (nuclear counterstaining)). (c) Sections are representative of n=30 patients/group. Representative western blots of p33ING1b and p29ING4 (loading control ß-actin).

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

Techniques: Expressing, Quantitative RT-PCR, Western Blot, Control

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 4: p33ING1b- and p29ING4-specific T cell reactivity. Four peptides resulted in significantly increased expression of IFN-γ compared to controls, independent of tumor stage (p33ING1b (aa109-118) and aa259-268, and p29ING4 (aa149-158) and aa239-248; p<0.001) Stages I/ II (a) and Stages III/IV (b). Elispot analysis was measured as spots/5 x 105 cells/patient.

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

Techniques: Expressing, Enzyme-linked Immunospot

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 5: p33ING1b- and p29ING4-specific T cell reactivity. Two peptides resulted in increased expression of IL-10 compared to controls; peptide p33ING1b (aa109-118) at all stages and p29ING4 (aa239-248) at late stages (p<0.001). Stages I/ II (a) and Stages III/IV (b). Luminex analysis was measured as pg/ml.

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

Techniques: Expressing, Luminex

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 6: p33ING1b- and p29ING4-specific T cell reactivity. Evaluation of IL-2 expression revealed that only a single epitope (p29ING4 (aa149-158)) resulted in high responsiveness of T cells from RCC patients after stimulation with the peptide (stages I/II vs. III/IV p<0.001). PBMCs from healthy volunteers (n=30) had no significant IL-2 expression after stimulation with all peptide epitopes. Luminex analysis for IL-2 expression was measured as pg/ml and presented as percent stimulation index.

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

Techniques: Expressing, Luminex

Journal: Journal of Clinical & Cellular Immunology

Article Title: Tumor Suppressor Gene P29ing4 is Overexpressed and Induces a CD8 T Effector Cell Response in Human Renal Cell Carcinoma

doi: 10.4172/2155-9899.1000511

Figure Lengend Snippet: Figure 7: p29ING4 presentation by MHC class I (a) and II (b) molecules. p29ING4 presentation by major histocompatibility complex (MHC) class I and II antigens was shown to be increased in RCC but not in healthy kidneys.

Article Snippet: Analysis of single staining was performed for p33ING1 (Santa Cruz Biotechnology, CA, USA), p29ING4 (Rockland/Biomol, Hamburg, Germany) CD4, CD25, Foxp3, CD8, (Abcam, Cambridge, UK), IFN-γ (BD Pharmingen, Heidelberg, Germany), IL-2, and IL-10 (R&D Systems, Minneapolis, USA).

Techniques: Immunopeptidomics

Journal: Biomolecules

Article Title: Herpesvirus Entry Mediator as an Immune Checkpoint Target and a Potential Prognostic Biomarker in Myeloid and Lymphoid Leukemia

doi: 10.3390/biom14050523

Figure Lengend Snippet: Light microscopy images and the expression of herpesvirus entry mediator (HVEM) surface protein on breast cancer (MCF-7), hepatocellular carcinomas (HepG2), chronic myelogenous leukemia (K562), and human embryonic kidney (Hek293) cells. The left column represents the microscope and a magnification of 100 pt using NIS-Elements F 4.00.00 software (Nikon Instruments, USA) under a light microscope (Nikon Eclipse, Fujisawa, Japan). Data collected from four separate experiments represents cell proliferation, morphology, and attachment nature. The right column shows histogram graphs representing the measurements of HVEM expression after staining with anti-HVEM-PE. The dotted-line histograms show the unstained control cells, whereas the line histograms represent the HVEM-stained cells. The numbers at the top of each histogram indicate the MFI of the unstained ( left ) and HVEM-PE ( right ). The number in the middle represents the percentage of HVEM-positive cells. The results were collected from two wells in three separate experiments. Magnification: 100 pt.

Article Snippet: The efficacy of blocking HVEM expressed on K562 leukemic cells by NBP1-76690PEP (Novus Biologicals, USA) on the proliferation of naïve CFSE CD4 + T cells showed an increase in CD4 + T cell proliferation in vitro.

Techniques: Light Microscopy, Expressing, Microscopy, Software, Staining, Control

Journal: Biomolecules

Article Title: Herpesvirus Entry Mediator as an Immune Checkpoint Target and a Potential Prognostic Biomarker in Myeloid and Lymphoid Leukemia

doi: 10.3390/biom14050523

Figure Lengend Snippet: The relative gene expression level of HVEM , normalized to the housekeeping gene GAPDH , is represented as a fold change.

Article Snippet: The efficacy of blocking HVEM expressed on K562 leukemic cells by NBP1-76690PEP (Novus Biologicals, USA) on the proliferation of naïve CFSE CD4 + T cells showed an increase in CD4 + T cell proliferation in vitro.

Techniques: Gene Expression

Journal: Biomolecules

Article Title: Herpesvirus Entry Mediator as an Immune Checkpoint Target and a Potential Prognostic Biomarker in Myeloid and Lymphoid Leukemia

doi: 10.3390/biom14050523

Figure Lengend Snippet: Light microscopic images of K562 ( left ) and Hek293 ( right ): ( A ) alone, ( B ) HVEM untreated cells with CFSE CD4 + T cells at one tumor or normal to ten T cells, or ( C ) HVEM blocker treated cells with CFSE CD4 + T cells after K562 and Hek293 for 72 h of incubation. Data are representative of three separate experiments. Magnification: 100 μm.

Article Snippet: The efficacy of blocking HVEM expressed on K562 leukemic cells by NBP1-76690PEP (Novus Biologicals, USA) on the proliferation of naïve CFSE CD4 + T cells showed an increase in CD4 + T cell proliferation in vitro.

Techniques: Incubation

Journal: Biomolecules

Article Title: Herpesvirus Entry Mediator as an Immune Checkpoint Target and a Potential Prognostic Biomarker in Myeloid and Lymphoid Leukemia

doi: 10.3390/biom14050523

Figure Lengend Snippet: Proliferation of CFSE-labeled CD4 + T cells in response to ( A ) K562 tumor cells or ( B ) Hek293 healthy control cells without HVEM blocker treatment ( left column), or after K562 and Hek293 treatment with HVEM blocker ( middle column); and non-proliferated CFSE CD4 + T cells ( right column) after 72 h of incubation at 37 °C. Data are representative of two separate experiments.

Article Snippet: The efficacy of blocking HVEM expressed on K562 leukemic cells by NBP1-76690PEP (Novus Biologicals, USA) on the proliferation of naïve CFSE CD4 + T cells showed an increase in CD4 + T cell proliferation in vitro.

Techniques: Labeling, Control, Incubation

Journal: Biomolecules

Article Title: Herpesvirus Entry Mediator as an Immune Checkpoint Target and a Potential Prognostic Biomarker in Myeloid and Lymphoid Leukemia

doi: 10.3390/biom14050523

Figure Lengend Snippet: Mean ± SD of fluorescence intensity of CFSE-labeled CD4 + T cells before proliferation on day zero and after 72 h of incubation with either K562 tumor cells or with Hek293 normal cells that were left untreated or treated with 20 ng HVEM blocker. Cells were cultured at a ratio of one tumor or normal cell to ten CFSE CD4 + T cells. ** indicates strong significant differences between groups with p = 0.0033, whereas “ns” indicates no significant differences with p = 0.5918. The results were collected from three separate experiments.

Article Snippet: The efficacy of blocking HVEM expressed on K562 leukemic cells by NBP1-76690PEP (Novus Biologicals, USA) on the proliferation of naïve CFSE CD4 + T cells showed an increase in CD4 + T cell proliferation in vitro.

Techniques: Fluorescence, Labeling, Incubation, Cell Culture

Journal: eLife

Article Title: Arf1/COPI machinery acts directly on lipid droplets and enables their connection to the ER for protein targeting

doi: 10.7554/elife.01607

Figure Lengend Snippet: Figure 3. The COPI machinery localizes to the LD surface. (A) The endogenous COPI machinery stained with αCOP or garz antibodies (red) localizes to LDs in S2 cells. Frequencies of colocalization of αCOP and garz spots with LDs from experiments are higher than expected from a random distribution. (B) The endogenous COPI machinery localizes to LDs in NRK cells. NRK cells stained for βCOP or GBF1 by immunofluorescence (red) show partial colocalization with LDs stained with BODIPY (green). Colocalization of βCOP with LDs in NRK cells is not random. Relative frequencies of βCOP, KDEL receptor and clathrin spots colocalizing with LDs determined in experiments are respectively compared to the frequencies of colocalization from a binomial random distribution. From the two frequencies (experiment vs simulation), a significant overrepresentation of βCOP on LDs is observed, whereas clathrin and KDEL receptor (KDELR) are not found on LDs. For (A) and (B) scale bars are 10 μm (overview) or 1 μm (first inlay) or 250 nm (second inlay). Statistical significance was tested by a student t test with p<0.01 (n = 30). (C) Localization of β’COP (green) to the LD surface (perilipin3, red) using confocal (upper panel) and super-resolution STED microscopy (lower panel). Scale bar = 500 nm (overview) or 100 nm (inlay). (D) Localization of β’COP to LDs is efficiently blocked by treatment of cells with the Arf1 GEF inhibitors brefeldin A or golgicide A. Scale bar = 10 μm (overview) or 1 μm (inlay). DOI: 10.7554/eLife.01607.008 The following figure supplements are available for figure 3:

Article Snippet: Rabbit polyclonal antibodies used: anti-GPAT4 (Wilfling et al., 2013), anti-CCT1 (Wilfling et al., 2013), anti-GBF1 (BD Biosciences, San Jose, CA), anti-KDEL-receptor (KDELR; gift from Dr JE Rothman; Yale University), anti-βCOP (gift from Dr JE Rothman; Yale University), anti-perilipin3 (TIP47; Novus Biologicals, Littleton, CO), anti-αCOP (Abcam, Cambridge, MA), anti-GRP78/BiP (ET-21) (Sigma–Aldrich, St. Louis, MO) and anti-garz (Wang et al., 2012) (gift from Dr A Paululat; University of Osnabrück).

Techniques: Staining, Immunofluorescence, Microscopy