Journal: Kidney International

Article Title: Early interleukin 6 production by leukocytes during ischemic acute kidney injury is regulated by TLR4

doi: 10.1038/ki.2011.140

Figure Lengend Snippet: Interleukin 6 (IL6) was measured by quantitative reverse transcriptase-PCR. Each sample normalized to glyceraldehyde 3-phosphate dehydrogenase (GAPDH). The calibrator gene is the media-treated BMDM. n = 4. Means and standard deviation are shown. *P<0.05 between TLR4 (+/+) media, HSP70, and HMGB1 groups. TLR4 (−/−) mice were C57BL/10ScNJ, and TLR4 (+/+) mice were C57BL/10SnJ.

Article Snippet: Macrophage cells were treated with either ROS-stressed S3 supernatant, recombinant human HMGB1 (rhHMGB1, R&D Systems) with or without 100 μmol/l glycyrrhizic acid (Sigma), or heat shock protein 70 (Stressgen, San Diego, CA) for 4 h. Treatment of S3 Cells with ROS The SV-40 transformed S3 tubular cells 83 were maintained as previously described.

Techniques: Reverse Transcription, Standard Deviation

Journal: Kidney International

Article Title: Early interleukin 6 production by leukocytes during ischemic acute kidney injury is regulated by TLR4

doi: 10.1038/ki.2011.140

Figure Lengend Snippet: (a) Experimental design. In stage I, the SV40-transformed S3 tubules were a gift from Dr Glenn Nagami.83 If they were injured by ROS, they released endogenous TLR4 ligands into their supernatant for 12 h. In stage II, the supernatant was cultured with murine bone marrow macrophages. (b) Only TLR4 (+/+) macrophages are activated by supernatant from ROS-stimulated S3 cells. TLR4 (+/+) macrophages are from C57BL/10SnJ mice. TLR4 (−/−) macrophages are from C57BL/10ScNJ mice. Results of typical experiment from four are shown. n = 3, mean and standard error are shown. *P<0.05 between S3 + ROS/TLR4 (+/+) macrophage and other groups by t-test. (c) High-mobility group protein B1 (HMGB1) in supernatants. n = 4. Mean and standard errors shown. P<0.05 between no treatment and ROS groups. IL6, interleukin 6.

Article Snippet: Macrophage cells were treated with either ROS-stressed S3 supernatant, recombinant human HMGB1 (rhHMGB1, R&D Systems) with or without 100 μmol/l glycyrrhizic acid (Sigma), or heat shock protein 70 (Stressgen, San Diego, CA) for 4 h. Treatment of S3 Cells with ROS The SV-40 transformed S3 tubular cells 83 were maintained as previously described.

Techniques: Transformation Assay, Cell Culture

Journal: Oncology reports

Article Title: Invasion potential of H22 hepatocarcinoma cells is increased by HMGB1-induced tumor NF-κB signaling via initiation of HSP70.

doi: 10.3892/or.2013.2595

Figure Lengend Snippet: Figure 1. Extracellular high-mobility group protein B1 (HMGB1) is increased in the tumor microenvironment during tumor progression by DTC-Ms and heat shock protein (HSP) 70. (A and B) Effect of DTC-Ms and HSP70 on tumor growth. (A) BALB/c mice were inoculated with H22 cells on day (d) 0 and received intramuscular injection of DTC-Ms and HSP70 (200 µg/mouse), once every 2 days from d1 to 13. Tumors (n=8 in each group) were dissected and weighed on d15 (d1-13 injection) after tumor inoculation. *P<0.05. (B) Detection of HMGB1 in tumors. Extracellular HMGB1 was detected by western blot analysis at the indicated time point after inoculation as described in Materials and methods. Tissues adjacent to the tumor were used as controls. (C) Expression of sTLR2 and sTLR4 after intramuscular transfection. Naked plasmid DNA was injected into muscle of mice. Tissues at sites of injection were surgically excised 72 h later and homogenized. The expressions of sTLR2 and sTLR4 were identified by reverse transcriptase-PCR and western blot analysis. (D) Flow cytometric analysis of TLR2 and TLR4 expression on H22 cells. (E and F) sTLR2 and sTLR4 suppressed HSP70 production but not HMGB1 production. (E) Mice inoculated with H22 cells received intramuscular injection of saline, pcDNA3.1, or psTLR2/psTLR4, with simultaneous injection of HSP70 as described in Materials and methods. Tumors (n=8 in each group) were dissected and weighed on d15 after tumor inoculation. *P<0.05. (F) Extracellular HMGB1 was detected by western blot analysis at the indicated time point after inoculation as described in Materials and methods.

Article Snippet: HMGB1 antibody was purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Injection, Western Blot, Expressing, Transfection, Plasmid Preparation, Reverse Transcription, Saline

Journal: Oncology reports

Article Title: Invasion potential of H22 hepatocarcinoma cells is increased by HMGB1-induced tumor NF-κB signaling via initiation of HSP70.

doi: 10.3892/or.2013.2595

Figure Lengend Snippet: Figure 2. Tumor cells exhibit an increased invasive growth by heat shock protein (HSP) 70‑induced expression of high-mobility group protein B1 (HMGB1). (A and B) H22 cells or HMGB1shRNA expressing H22 cells were treated in vitro and then inoculated to the liver of mice. Mice received no treatment or were administered HSP70 or HMGB1 as described in Materials and methods. (A) The invasive growth of tumor cells after HSP70 or HMGB1 treatment. The tumors in the liver of the mice are shown (upper). The main tumor nodules were measured (lower, left) and satellite tumor nodes counted (lower, right). *P<0.05, compared with control or saline groups. No satellite nodes were observed in control or the HMGB1shRNA mice administered H22 tumors. (B) On day (d) 16 after inoculation, tumor marginal tissues were surgically removed to extract RNA and proteins. The relative mRNA levels of MMP‑9 were detected by real‑time reverse transcriptase (RT)‑PCR (left). *P<0.05, compared with control groups. MMP‑9 in supernatants was detected by zymography assay (right). (C) Assay of MMP‑9 production. Cells were cultured in the presence or absence of HSP70 or HMGB1 for 5 or 24 h. The relative mRNA levels of MMP‑9 were detected by real‑time RT‑PCR (left). *P<0.05, compared with control groups. MMP‑9 in supernatants was detected by zymography assay (right).

Article Snippet: HMGB1 antibody was purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Expressing, In Vitro, Control, Saline, Reverse Transcription, Reverse Transcription Polymerase Chain Reaction, Zymography, Cell Culture, Quantitative RT-PCR

Journal: Oncology reports

Article Title: Invasion potential of H22 hepatocarcinoma cells is increased by HMGB1-induced tumor NF-κB signaling via initiation of HSP70.

doi: 10.3892/or.2013.2595

Figure Lengend Snippet: Figure 5. Receptor for advanced glycation end products (RAGE) but not Toll-like receptor (TLR)2/TLR4 is required for high-mobility group protein B1 (HMGB1)‑mediated MMP‑9 enhancement. (A) H22 cells were transfected with the indicated RAGE‑shRNA for 48 h and were then cultured in the presence of HMGB1 and/or resveratrol (Res, 30 µM). MMP‑9 in supernatants was detected by zymography assay. (B) H22 cells were cultured in the presence of HMGB1 and QNZ (20 nM) or Res (30 µM), and NF‑κB phosphorylation was analyzed by western blotting. (C) HepG2 cells were transfected with the indicated RAGE‑shRNA for 48 h and were then cultured in the presence of HMGB1 and/or Res (30 µM). MMP‑9 in supernatants was detected by zymography assay.

Article Snippet: HMGB1 antibody was purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Transfection, Cell Culture, Zymography, Phospho-proteomics, Western Blot

Journal: Oncology reports

Article Title: Invasion potential of H22 hepatocarcinoma cells is increased by HMGB1-induced tumor NF-κB signaling via initiation of HSP70.

doi: 10.3892/or.2013.2595

Figure Lengend Snippet: Figure 4. Time course analysis of heat shock protein (HSP) 70‑induced high-mobility group protein B1 (HMGB1) and Beclin‑1 production and effect of Jun N-terminal kinase (JNK) inhibitors on HSP70‑induced NF‑κB activation. (A) Effect of HSP70 on production of HMGB1 and Beclin‑1. H22 cells were incubated with HSP70 (20 µg/ml) for various times and Beclin‑1 and HMGB1 protein production was analyzed over a 6‑h period. Supernatants were collected at various times after HSP70 stimulation and the concentration of Beclin‑1 and HMGB1 was determined by western blotting and compared with basal levels (time 0). (B) A JNK specific inhibitor inhibited JNK activation. Cells were treated with and/or without JNK specific inhibitor (SP600125, 10 µmol/l) followed by stimulation with HSP70 (20 µg/ml) for 4 h. JNK and p‑JNKs were measured by western blotting. (C) Activation of JNK is required for HSP70‑induced Beclin‑1 and HMGB1 production. Cells were treated with and/or without a JNK specific inhibitor (SP600125, 10 µmol/l) and/or Beclin‑1 shRNA followed by stimulation with HSP70 (20 µg/ml) for 4 h. Empty vector served as a negative control. HMGB1 and Beclin‑1 were measured by western blotting. (D) Effects on NF‑κB activation in H22 cells. H22 cells were incubated with HMGB1 (20 µg/ml) or with a JNK specific inhibitor (SP600125, 10 µmol/l) followed by stimulation with HSP70 (20 µg/ml) for various times (0‑8 h), and NF‑κB phosphorylation was analyzed by western blotting.

Article Snippet: HMGB1 antibody was purchased from Cell Signaling Technology (Danvers, MA, USA).

Techniques: Activation Assay, Incubation, Concentration Assay, Western Blot, shRNA, Plasmid Preparation, Negative Control, Phospho-proteomics

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a Prediction of candidate HMGB1 crosstalk RNAs using the ceRDB database ( https://www.oncomir.umn.edu/cefinder ). The top 12 candidate RNAs and scores were shown. b The RNA expression levels of HMGB1 and the 12 candidate RNAs were analyzed in early stage HCC (BCLC stages 0 and A HCC) tissues by qRT-PCR. HBV + HCC (above) n = 26; HBV − HCC (below), n = 9. The results are means ± SD. Names marked red and black represent significantly ( p < 0.05) and nonsignificant genes, respectively. c Pearson correlation coefficient analysis between expression levels of HMGB1 and the 12 candidate RNAs, respectively, in early stage HCC tumor tissues

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: RNA Expression, Quantitative RT-PCR, Expressing

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a The mRNA expression of HMGB1 and RICTOR in the 11 indicated cell lines as determined by qRT-PCR. * p < 0.05. b Pearson correlation coefficient analysis between the expression level of HMGB1 and RICTOR in the cell lines from ( a ). c Immunohistochemical staining of HMGB1 and RICTOR in liver tissues in the DEN + CCl4-induced liver cancer mouse model. d , e The expression levels of RICTOR and HMGB1 mRNA (left) and protein (right) after the interference of HMGB1 or RICTOR in the HCCLM3 ( d ) and PLC/PRF/5 ( e ) cell lines (abbreviated as “KD-HMGB1 or KD-RICTOR”, respectively). f The expression levels of RICTOR and HMGB1 mRNA (left) and protein (right) after overexpression of HMGB1 or RICTOR 3′UTR in the QSG-7701 cell line (abbreviated as “OE-HMGB1 or OE-RICTOR”, respectively). The mRNA levels (left) were determined by qRT-PCR and protein level (right) was determined by Western blot. * p < 0.05, *** p < 0.001

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: Expressing, Quantitative RT-PCR, Immunohistochemical staining, Staining, Over Expression, Western Blot

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a Luciferase assays with reporter constructs containing the wild-type or mutant HMGB1/RICTOR 3′UTR downstream of the luciferase gene were performed after co-transfection with miR-200a/200b/429 mimics in HEK293T cells. * p < 0.05, ** p < 0.01, *** p < 0.001. b The mRNA levels of HMGB1 and RICTOR were determined by qRT-PCR 48 h after transfection of 100 nM miR-200a/200b/429 mimics in indicated cells. ** p < 0.01, *** p < 0.001. c RNA FISH assays detecting the cellular localization of HMGB1/RICTOR and miR-429 in indicated cells

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: Luciferase, Construct, Mutagenesis, Cotransfection, Quantitative RT-PCR, Transfection

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a Determination of spheroid formation after 3000 cells were seeded in low-adhesion plates for 10 days (left). The number of tumor spheroids was quantitated (right). *** p < 0.001. b Cell proliferation was evaluated using CCK8 assays. The results are means ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001. c Male nude mice ( n = 5) were subcutaneously injected with 1 × 10 4 HMGB1/RICTOR 3′UTR over-expressed liver cells (abbreviated as “OE-HMGB1 or OE-RICTOR”, respectively) or HMGB1/RICTOR mRNA interference HCC cells (abbreviated as “KD-HMGB1 or KD-RICTOR”, respectively). Tumorigenesis was assessed after 48 days as shown

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: Injection

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a PMBC killing assay scheme. HMGB1/RICTOR mRNA interference HCC cells (abbreviated as “KD-HMGB1 or KD-RICTOR”, respectively) were cocultured with activated PBMCs and treated with or without Atezolizumab for 48 h before apoptosis detection. b Cell apoptosis in treated HCC cells was evaluated by TUNEL assay (above). The apoptotic cell ratios were shown (below). c Exosomes isolated from cell culture supernatants in treated HCC cells were determined by flow cytometry using PE-fluorescein-conjugated anti-PD-L1 antibody (red). Isotype matched antibody (PE-fluorescein-conjugated anti-IgG antibody) was used as gating controls (black). Percentages of PD-L1 + Exo resulted from the relative fluorescence values gated by isotype controls. Exo: exosomes. ** p < 0.01, *** p < 0.001

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: TUNEL Assay, Isolation, Cell Culture, Flow Cytometry, Fluorescence

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a A model of the RNA–RNA crosstalk network involving HMGB1 and RICTOR acting in the early stage HCC

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques:

Journal: The Journals of Gerontology Series A: Biological Sciences and Medical Sciences

Article Title: Role of mTOR in the Development of Asthma in Mice With Cigarette Smoke-Induced Cellular Senescence

doi: 10.1093/gerona/glab303

Figure Lengend Snippet: Effects of rapamycin on the CSE-induced changes of inflammatory and senescence-associated markers in lung and glutathione in serum. ( A ) Experimental protocol (5–6 mice in each group). ( B ) Inflammatory cells in bronchoalveolar lavage fluid. ( C ) SA-β-gal activity in lung homogenate. Y axis represents relative fluorescence unit (RFU). ( D ) A plot of p21+ population in lung cells. ( E ) The frequency of p21+ population in lung cells. ( F ) Total glutathione, oxidized glutathione and reduced glutathione levels, and reduced/oxidized glutathione ratios in serum were measured, and the detection time was 2 minutes. TNF-α, Pro-MMP9, IL-6, IL-1β, S100A8/9, and HMGB1 levels in bronchoalveolar lavage fluid were measured. # p < .05, ## p < .01 compared to control (PBS) group; * p < .05, ** p < .01 between 2 groups. PBS = phosphate-buffered saline; SA-β-gal = senescence-associated beta-galactosidase; CSE = cigarette smoke extract.

Article Snippet: Tumor necrosis factor (TNF)-α (BioLegend, CA), Pro-MMP9 (R&D Systems, Abingdon, UK), interleukin(IL)-6 (BioLegend), IL-1β (BioLegend), S100A8/9 (R&D Systems), and HMGB1 (CUSABIO, Houston, TX) levels in bronchoalveolar lavage (BAL) fluid and Der p-specific immunoglobulin E (IgE) level in serum were measured using enzyme-linked immunosorbent assay according to the manufacturer’s instructions.

Techniques: Activity Assay, Fluorescence, Control, Saline

Journal: The Journals of Gerontology Series A: Biological Sciences and Medical Sciences

Article Title: Role of mTOR in the Development of Asthma in Mice With Cigarette Smoke-Induced Cellular Senescence

doi: 10.1093/gerona/glab303

Figure Lengend Snippet: Murine asthma model with CSE-induced cellular senescence and effects of rapamycin on this model. ( A ) Experimental protocol (4–5 mice in each group). ( B ) SA-β-gal activity in lung homogenate. Y axis represents relative fluorescence unit (RFU). ( C ) Airway hyperresponsiveness. Y axis represents cmH 2 O.s/mL. ( D ) Inflammatory cells in bronchoalveolar lavage fluid. ( E ) Serum Der p -specific IgE. Y axis represents OD (optimal density). ( F ) Gating plot of MHCII+ CD86+ population in CD11c+ cells from lung-draining lymph node. ( G ) The frequency of MHCII+ CD86+ population with CD11c+ in total cells from lung-draining lymph node. ( H ) Lung tissue (H&E stain, ×200). ( I ) The frequencies of IL-5+, IL-13+, IL-17+, and IFN-γ+ populations in CD4+ cells. ( J ) Gating plot of S100A8/9+ HMGB1+ population in CD45− Epcam+ cells. ( K ) The frequency of S100A8/9+ HMGB1+ population in CD45− Epcam+ cells. # p < .05, ## p < .01 compared to control (PBS) group; * p < .05, ** p < .01 between 2 groups. Der p = Dermatophagoides pteronyssinus ; Meth con = methacholine concentration; Macro = macrophage; Neu = neutrophil; Eos = eosinophil; Lym = lymphocyte; Rapa = rapamycin; CSE = cigarette smoke extract.

Article Snippet: Tumor necrosis factor (TNF)-α (BioLegend, CA), Pro-MMP9 (R&D Systems, Abingdon, UK), interleukin(IL)-6 (BioLegend), IL-1β (BioLegend), S100A8/9 (R&D Systems), and HMGB1 (CUSABIO, Houston, TX) levels in bronchoalveolar lavage (BAL) fluid and Der p-specific immunoglobulin E (IgE) level in serum were measured using enzyme-linked immunosorbent assay according to the manufacturer’s instructions.

Techniques: Activity Assay, Fluorescence, Staining, Control, Concentration Assay

Journal: The Journals of Gerontology Series A: Biological Sciences and Medical Sciences

Article Title: Role of mTOR in the Development of Asthma in Mice With Cigarette Smoke-Induced Cellular Senescence

doi: 10.1093/gerona/glab303

Figure Lengend Snippet: Effects of rapamycin on the CSE-induced changes of S100A8/9+ or HMGB1+ populations in various lung cells. ( A ) Gating plot and the frequency of S100A8/9+ population in CD45− EpCAM+ cells. ( B ) Gating plot and the frequency of S100A8/9+ F4/80+ population in CD45+ cells. ( C ) Gating plot and the frequency of S100A8/9+ CD11c+ population in CD45+ cells. ( D ) Gating plot and the frequency of S100A8/9+ population in CD4+ cells. ( E ) Gating plot and the frequency of HMGB1+ population in CD45− EpCAM+ cells. ( F ) Gating plot and the frequency of HMGB1+ F4/80+ population in CD45+ cells. ( G ) Gating plot and the frequency of HMGB1+ CD11c+ population in CD45+ cells. ( H ) Gating plot and frequency of HMGB1+ population in CD4+ cells. # p < .05 compared to control (PBS) group; * p < .05, ** p < .01 between 2 groups. PBS = phosphate-buffered saline; CSE = cigarette smoke extract.

Article Snippet: Tumor necrosis factor (TNF)-α (BioLegend, CA), Pro-MMP9 (R&D Systems, Abingdon, UK), interleukin(IL)-6 (BioLegend), IL-1β (BioLegend), S100A8/9 (R&D Systems), and HMGB1 (CUSABIO, Houston, TX) levels in bronchoalveolar lavage (BAL) fluid and Der p-specific immunoglobulin E (IgE) level in serum were measured using enzyme-linked immunosorbent assay according to the manufacturer’s instructions.

Techniques: Control, Saline

Journal: The Journals of Gerontology Series A: Biological Sciences and Medical Sciences

Article Title: Role of mTOR in the Development of Asthma in Mice With Cigarette Smoke-Induced Cellular Senescence

doi: 10.1093/gerona/glab303

Figure Lengend Snippet: Effects of rapamycin on CSE-stimulated MLE-12 cells and effects of S100A9 or HMGB1 overexpressing MLE-12 cells treated by low-dose Dermatophagoides pteronyssinus allergen on dendritic cell activation. ( A ) Plot (upper) and histogram (lower) of DCF+ population in CSE-stimulated MLE-12 cells (with or without rapamycin). ( B ) The frequency of DCF+ population. Y axis of graph represents % of DCF+ cells. ( C ) Gating plot of SA-β-gal+ population in total cells from CSE-stimulated MLE-12 cells (with or without rapamycin). ( D ) The frequency of SA-β-gal+ population. ( E ) Gating plot of p-mTOR+ population in S100A8/9+ HMGB1+ cells in singlets from CSE-stimulated MLE-12 cells (with or without rapamycin). ( F ) The frequency of p-mTOR+ population in S100A8/9+HMGB1+ cells. ( G ) Experimental procedure. ( H ) Gating plot of MHCII+ CD11c+ population in BMDC after coculture with S100A9 or HMGB1 overexpressing MLE-12 cells. ( I ) The frequency of MHCII+ CD11c+ cells in BMDCs after coculture with S100A9 or HMGB1 overexpressing MLE-12 cells. # p < .05, ## p < .01 compared to media; * p < .05, ** p < .01 between 2 groups. DCF = dichlorofluorescein; CSE = cigarette smoke extract.

Article Snippet: Tumor necrosis factor (TNF)-α (BioLegend, CA), Pro-MMP9 (R&D Systems, Abingdon, UK), interleukin(IL)-6 (BioLegend), IL-1β (BioLegend), S100A8/9 (R&D Systems), and HMGB1 (CUSABIO, Houston, TX) levels in bronchoalveolar lavage (BAL) fluid and Der p-specific immunoglobulin E (IgE) level in serum were measured using enzyme-linked immunosorbent assay according to the manufacturer’s instructions.

Techniques: Activation Assay

Journal: European Journal of Histochemistry : EJH

Article Title: Inhibition of HMGB1 suppresses inflammation and catabolism in temporomandibular joint osteoarthritis via NF-κB signaling pathway

doi: 10.4081/ejh.2022.3357

Figure Lengend Snippet: Inhibition of HMGB1 decreases the expression of pro-inflammatory cytokines and catabolic mediators in rat synovium of CFA induced TMJOA. A) The schematic chart for the timeline of treating rats. B) The expression of ADAMTS5, MMP13, IL-1β, IL-6 and negative controls was detected by immunohistochemistry in the synovium of CFA groups, CFA with anti-HMGB1 antibody treatment groups at day 21, or in the control synovium; scale bar: 100 μm. C) Quantitative analysis of ADAMTS5, MMP13, IL-1β and IL- 6 in the synovium of rats; data are shown as the means with 95% CI and analyzed by one-way analysis of variance (ANOVA), n=5, *p<0.05, **p<0.01, ***p<0.001.

Article Snippet: After 2 weeks, 5 rats previously injected with CFA were intraperitoneally injected with 100 μg/kg/day anti HMGB1 antibody (MAB1690-100; R&D System) for 7 days.

Techniques: Inhibition, Expressing, Immunohistochemistry

Journal: European Journal of Histochemistry : EJH

Article Title: Inhibition of HMGB1 suppresses inflammation and catabolism in temporomandibular joint osteoarthritis via NF-κB signaling pathway

doi: 10.4081/ejh.2022.3357

Figure Lengend Snippet: Inhibition of HMGB1 alleviates the damage of cartilage and subchondral bone in CFA induced TMJOA. A) H&E, Safranin O and Masson trichrome staining of rat models at day 21; scale bar: 100 μm. B-D) Analysis of the condylar cartilage thickness, the Safranin O positive areas and unmineralized bone areas; data are shown as the means with 95% CI and analyzed by one-way analysis of variance (ANOVA), n=5, *p<0.05, **p<0.01, ***p<0.001.

Article Snippet: After 2 weeks, 5 rats previously injected with CFA were intraperitoneally injected with 100 μg/kg/day anti HMGB1 antibody (MAB1690-100; R&D System) for 7 days.

Techniques: Inhibition, Staining

Journal: European Journal of Histochemistry : EJH

Article Title: Inhibition of HMGB1 suppresses inflammation and catabolism in temporomandibular joint osteoarthritis via NF-κB signaling pathway

doi: 10.4081/ejh.2022.3357

Figure Lengend Snippet: Inhibition of HMGB1 decreases the expression of pro-inflammatory cytokines and catabolic mediators in rat cartilage of CFA induced TMJOA. A) The expression of ADAMTS5, MMP13, IL-1β, IL-6 and negative controls was detected by immunohistochemistry in the cartilage of CFA groups, CFA with anti-HMGB1 antibody treatment groups at day 21, or in the control cartilage; scale bar: 100 μm. B) Rate of ADAMTS5, MMP13, IL-1β and IL-6 positive cells in the cartilage was markedly increased in the CFA groups, while markedly inhibited after treatment with anti-HMGB1 antibody; data are shown as the means with 95% CI and analyzed by one-way analysis of variance (ANOVA), n=5, *p<0.05, **p<0.01, ***p<0.001.

Article Snippet: After 2 weeks, 5 rats previously injected with CFA were intraperitoneally injected with 100 μg/kg/day anti HMGB1 antibody (MAB1690-100; R&D System) for 7 days.

Techniques: Inhibition, Expressing, Immunohistochemistry

Journal: European Journal of Histochemistry : EJH

Article Title: Inhibition of HMGB1 suppresses inflammation and catabolism in temporomandibular joint osteoarthritis via NF-κB signaling pathway

doi: 10.4081/ejh.2022.3357

Figure Lengend Snippet: Inhibition of HMGB1 suppresses the NF-κB signaling pathway in vivo. Expression of phospho-p65 in the synovium (A) and cartilage (B) of CFA groups, CFA with anti-HMGB1 antibody treatment groups at day 21 were detected by IF staining, or in the control groups; data are shown as the means with 95% CI and analyzed by one-way analysis of variance (ANOVA), n=5, *p<0.05, **p<0.01, ***p<0.001; scale bar: 100 μm. C) Schematic diagram of inhibition of HMGB1 suppresses inflammation and catabolism in temporomandibular joint osteoarthritis via NF-κB signaling pathway.

Article Snippet: After 2 weeks, 5 rats previously injected with CFA were intraperitoneally injected with 100 μg/kg/day anti HMGB1 antibody (MAB1690-100; R&D System) for 7 days.

Techniques: Inhibition, In Vivo, Expressing, Staining

Journal: International Journal of Molecular Sciences

Article Title: Anti-Septic Functions of Cornuside against HMGB1-Mediated Severe Inflammatory Responses

doi: 10.3390/ijms23042065

Figure Lengend Snippet: Effects of overexpression of HMGB1 on the anti-inflammatory functions of CN. Overexpression of human HMGB1 in HUVECs was achieved using the pCMV6-Ac-GFP vector (pCMV-HMGB1). ( A – F ) represent similar experiments as shown in A ( A ), A ( B ), B ( C ), C ( D ), A ( E ), and B ( F ), except that HUVECs were transfected with mock-vector (Mock-HMGB1) or pCMV6-Ac-GFP vector (pCMV-HMGB1). Data are expressed as the mean ± SD values of three independent experiments on different days. * p < 0.05 versus group treatment with LPS only.

Article Snippet: Overexpression of HMGB1 in HUVECs was achieved by incorporating human HMGB1 in the pCMV6-Ac-GFP vector (RG205918, OriGene Technologies, Inc., Rockville, MD, USA).

Techniques: Over Expression, Plasmid Preparation, Transfection

Journal: Journal of Neuroinflammation

Article Title: Role of high mobility group box protein 1 (HMGB1) in peripheral blood from patients with multiple sclerosis

doi: 10.1186/s12974-015-0269-9

Figure Lengend Snippet: Demographic and baseline clinical characteristics of MS patients and healthy controls included in the HMGB1 expression study

Article Snippet: HMGB1 transcripts were determined with TaqMan® gene expression assays (Hs01590761_g1; Applied Biosystems).

Techniques: Expressing

Journal: Journal of Neuroinflammation

Article Title: Role of high mobility group box protein 1 (HMGB1) in peripheral blood from patients with multiple sclerosis

doi: 10.1186/s12974-015-0269-9

Figure Lengend Snippet: Bar graphs comparing HMGB1 mRNA expression levels in PBMCs from MS patients and healthy controls (A) and between different clinical forms of MS (B). HMGB1 expression was determined by real-time PCR using GAPDH as endogenous control. Results are expressed as fold change in HMGB1 gene expression in MS patients relative to controls. Errors bars represent standard error of the mean. Number of individuals included in the study is shown in parentheses. HC: healthy controls. MS: whole group of multiple sclerosis patients. RR: relapsing-remitting MS. SP: secondary progressive MS. PP: primary progressive MS.

Article Snippet: HMGB1 transcripts were determined with TaqMan® gene expression assays (Hs01590761_g1; Applied Biosystems).

Techniques: Expressing, Real-time Polymerase Chain Reaction, Control, Gene Expression

Journal: Journal of Neuroinflammation

Article Title: Role of high mobility group box protein 1 (HMGB1) in peripheral blood from patients with multiple sclerosis

doi: 10.1186/s12974-015-0269-9

Figure Lengend Snippet: Box plots showing serum levels of HMGB1 in the whole MS group and controls (A) and in MS patients with different clinical forms of MS (B). Serum levels of HMGB1 were measured using a commercially available ELISA, as described in ‘ .’ For the sake of clarity, only significant P values are shown in the graphs. Number of individuals included in the study is shown in parentheses. Eleven patients with RRMS, five with SPMS, and eight with PPMS were also included in the HMGB1 gene expression study. HC: healthy controls. MS: whole group of multiple sclerosis patients. RR: relapsing-remitting MS. SP: secondary progressive MS. PP: primary progressive MS.

Article Snippet: HMGB1 transcripts were determined with TaqMan® gene expression assays (Hs01590761_g1; Applied Biosystems).

Techniques: Enzyme-linked Immunosorbent Assay, Gene Expression

Journal: Oncotarget

Article Title: Targeted gene silencing of CCL2 inhibits triple negative breast cancer progression by blocking cancer stem cell renewal and M2 macrophage recruitment

doi: 10.18632/oncotarget.9885

Figure Lengend Snippet: MDA-MB-231 breast tumor xenografts were injected with Ca-TAT peptides complexed to control siRNAs (Con), or CCL2 siRNAs (huCCL2si1 or huCCL2si2), and immunostained for expression of A. PCNA, B. Cleaved caspase-3, C. HMGB1, or D. LC3B. Scale=100 microns. Statistical analysis was performed by One Way ANOVA followed by Bonferonni post-hoc analysis. Statistical significance was determined by p-value less than 0.05. *p<0.05, ***p<0.001, n.s=not significant. Mean+SEM is shown. N=6 animals per group.

Article Snippet: Samples were incubated with the following antibodies at 1: 50 dilution, overnight on ice in PBS containing 2% BSA: CD24- PE (cat no.555428, BD Pharmingen), CD11b-APC-Cy7 (cat. no 557657, BD Pharmingen), murine CCL2 (cat no. 1784, Santa Cruz Biotechnology), human CCL2 (cat no. sc1304, Santa Cruz Biotechnology) Ki67 (cat no. Sc15402, Santa Cruz Biotechnology), HMGB1 (cat no. 6893, Cell Signaling Technology) or LC3B (cat no. L10382, Invitrogen).

Techniques: Injection, Control, Expressing