Journal: Respiratory Research

Article Title: RAGE inhibition alleviates lipopolysaccharides-induced lung injury via directly suppressing autophagic apoptosis of type II alveolar epithelial cells

doi: 10.1186/s12931-023-02332-6

Figure Lengend Snippet: LPS-induced autophagic death in A549 cells depended on RAGE activation. A Treated A549 cells with siRAGE and detected the RAGE mRNA expression. The expression of Ager genes was decreased by 50%. B Detected the immunoblots of RAGE as well as Beclin1, LC3 II/I and cleaved Caspase 3 in A549 cells, each experiment was repeated more than three times. C The cell survival rate of A549 cells in response to siRAGE pre-treatment followed by LPS stimulation. Data were obtained from CCK-8 experiments and performed more than three times. D Immunofluorescence of cleaved Caspase3 to detect the apoptosis of A549 cells, spinning disk confocal microscopy at × 40 magnification (scale bar 50 μm). E Detected the immunoblots of STAT3 and phosphorylation STAT3 (p-STAT3) in A549 cells, each experiment was repeated more than three times. *Indicates the significant difference compared with the control group. # Indicates the significant difference compared with LPS group. P < 0.05, differences in characteristics between groups were analyzed using the Kruskal–Wallis test with Dunn’s post hoc tests

Article Snippet: Anti-RAGE (1:1000, #6996S; Cell Signaling Technology, Inc., MA, USA), anti-STAT3 (1:1000, #12640; Cell Signaling Technology, Inc., MA, USA), anti-phosphorylated STAT3 (1:2000, #9145; Cell Signaling Technology, Inc., MA, USA), anti-cleaved caspase 3 (1:1000, #9664; Cell Signaling Technology, Inc., MA, USA), anti-LC3II/I (1:1000, #4108; Cell Signaling Technology, Inc., MA, USA), anti-Beclin1 (1:1000, #3495; Cell Signaling Technology, Inc., MA, USA) and anti-GADPH (1:1000, #2118; Cell Signaling Technology, Inc., MA, USA) were used as primary antibodies.

Techniques: Activation Assay, Expressing, Western Blot, CCK-8 Assay, Immunofluorescence, Confocal Microscopy

Journal: Molecular Vision

Article Title: Expression of high-mobility groups box-1/receptor for advanced glycation end products/osteopontin/early growth response-1 pathway in proliferative vitreoretinal epiretinal membranes

doi:

Figure Lengend Snippet: Specificity of the Antibodies used in the study.

Article Snippet: HMGB1 binding to RAGE activates key cell-signaling pathways such as mitogen-activated protein kinases and NF-κB [ - ].

Techniques: Sequencing

Journal: Molecular Vision

Article Title: Expression of high-mobility groups box-1/receptor for advanced glycation end products/osteopontin/early growth response-1 pathway in proliferative vitreoretinal epiretinal membranes

doi:

Figure Lengend Snippet: Proliferative diabetic retinopathy epiretinal membranes. A : Immunohistochemical staining of panendothelial cell marker CD34 shows blood vessels positive for CD34 (arrows; original magnification 40×). Immunohistochemical staining of high-mobility group box −1 (HMGB1). Low power ( B ; original magnification 40×) and high power ( C ; original magnification 100×) showing vascular endothelial (arrows) and stromal (arrowheads) cells expressing strong immunoreactivity to HMGB1. Immunohistochemical staining of receptor for advanced glycation end products (RAGE). Low-power ( D ; original magnification 40×) and high-power ( E ; original magnification 100×) showing vascular endothelial (arrows) and stromal (arrowheads) cells expressing strong immunoreactivity to RAGE. Immunohistochemical staining of osteopontin (OPN) showing vascular endothelial cells (arrows; F ) and stromal cells (arrowheads; G ) expressing strong immunoreactivity to OPN (original magnification 100×). Immunohistochemical staining of early growth response-1 showing immunoreactivity in vascular endothelial (arrows) and stromal (arrowheads) cells ( H ; original magnification 100×).

Article Snippet: HMGB1 binding to RAGE activates key cell-signaling pathways such as mitogen-activated protein kinases and NF-κB [ - ].

Techniques: Immunohistochemical staining, Staining, Marker, Expressing

Journal: Molecular Vision

Article Title: Expression of high-mobility groups box-1/receptor for advanced glycation end products/osteopontin/early growth response-1 pathway in proliferative vitreoretinal epiretinal membranes

doi:

Figure Lengend Snippet: Proliferative vitreoretinopathy epiretinal membranes. Immunohistochemical staining of α-smooth muscle actin showing immunoreactivity in spindle-shaped myofibroblasts (arrows; A ) and ( B ; original magnification 100×). Immunohistochemical staining of high-mobility group box −1 (HMGB1). Low-power ( C ; original magnification 40×) and high-power ( D ; original magnification 100×) showing spindle-shaped cells expressing cytoplasmic immunoreactivity to HMGB1 (arrows). Immunohistochemical staining of the receptor for advanced glycation end products (RAGE) showing spindle-shaped cells expressing cytoplasmic immunoreactivity to RAGE (arrows; E ; original magnification 40×). Immunohistochemical staining of osteopontin showing strong cytoplasmic immunoreactivity in spindle-shaped cells (arrows; F ; original magnification 40×). Immunohistochemical staining of early growth response-1 showing spindle-shaped cells expressing cytoplasmic immunoreactivity (arrows; G ; original magnification 40×) and cells expressing nuclear immunoreactivity (arrows; H ; original magnification 100×).

Article Snippet: HMGB1 binding to RAGE activates key cell-signaling pathways such as mitogen-activated protein kinases and NF-κB [ - ].

Techniques: Immunohistochemical staining, Staining, Expressing

Journal: Molecular Vision

Article Title: Expression of high-mobility groups box-1/receptor for advanced glycation end products/osteopontin/early growth response-1 pathway in proliferative vitreoretinal epiretinal membranes

doi:

Figure Lengend Snippet: Mean numbers in relation to type of proliferative diabetic retinopathy (PDR).

Article Snippet: HMGB1 binding to RAGE activates key cell-signaling pathways such as mitogen-activated protein kinases and NF-κB [ - ].

Techniques: Expressing

Journal: PLoS ONE

Article Title: Soluble HMGB1 Is a Novel Adipokine Stimulating IL-6 Secretion through RAGE Receptor in SW872 Preadipocyte Cell Line: Contribution to Chronic Inflammation in Fat Tissue

doi: 10.1371/journal.pone.0076039

Figure Lengend Snippet: A) TLR2, TLR4, RAGE and HMGB1 mRNA expression was assessed by RT-PCR from total RNA obtained from SW872 cell after 12h in culture, with (+) and without (−) Reverse Transcriptase (RT). B) Cell surface expression of TLR2, TLR4 and RAGE were analyzed by FACS C) SW872 cells were treated with rHMGB1 1 µg.mL −1 for 5 h with control mouse IgG (CTL), TLR2 or TLR4 blocking antibody (n = 4). D) SW872 cells were treated as in (C) except that control goat IgG (CTL) or goat polyclonal anti-RAGE were used (n = 4). All values are expressed as means +/− SD and degrees of significance are indicated in the figure captions as follow, * p<0.05, ** p<0.01, *** p<0.001 and ns = not significant.

Article Snippet: This result was confirmed using another antibody against RAGE (rabbit anti-RAGE, 4679S, Cell Signaling Technology) (data not shown).

Techniques: Expressing, Reverse Transcription Polymerase Chain Reaction, Blocking Assay

Journal: Burns & Trauma

Article Title: Curcumin-incorporated 3D bioprinting gelatin methacryloyl hydrogel reduces reactive oxygen species-induced adipose-derived stem cell apoptosis and improves implanting survival in diabetic wounds

doi: 10.1093/burnst/tkac001

Figure Lengend Snippet: Cur suppresses AGEs/AGER axis-mediated ROS and ADSC apoptosis. ( a ) ADSCs pre-treated with AGEs were mixed with Cur or the p65 inhibitor PDTC (30 μM). Left: western blot demonstrates protein expression levels of AGER and phosphorylated and total p65. Right: comparison of AGER and p-p65 expression levels between the study groups. ( b ) ROS level was tested by fluorescence microscopy and total cell number was observed by bright field microscopy to calculate the percentage. ( c ) Flow cytometry showing ROS levels in ADSCs. ( d ) FITC/PI stained ADSCs analyzed by flow cytometry. Data are shown as means ± SD. Statistical analysis: * p <0.05, * * p <0.01 and * p <0.001. AGE Advanced glycation end product, AGER AGE receptor, ROS reactive oxygen species, ADSC adipose-derived stem cell, Cur curcumin, PDTC pyrrolidinedithiocarbamate

Article Snippet: The protein was transferred to a PVDF membrane and the primary antibody was incubated at 4°C overnight, followed by incubation with the secondary antibody (BA1054, Boster, Wuhan, China) at room temperature for 2 h. The primary antibodies were rabbit AGER (#55222, 1:500; Cell Signaling Technology/CST, Beverly, MA, USA), p65 (#3033, 1:500; CST), p-p65 (#8242, 1:500; CST) and β-actin (ab8227, 1:1000; Abcam, Cambridge, UK).

Techniques: Western Blot, Expressing, Fluorescence, Microscopy, Flow Cytometry, Staining, Derivative Assay

Journal: Burns & Trauma

Article Title: Curcumin-incorporated 3D bioprinting gelatin methacryloyl hydrogel reduces reactive oxygen species-induced adipose-derived stem cell apoptosis and improves implanting survival in diabetic wounds

doi: 10.1093/burnst/tkac001

Figure Lengend Snippet: Role of NF-κB p65 signal in the inhibitory effect of Cur on ADSCs. ( a ) Left: time course analysis of AGER and phosphorylated and total p65 protein expression levels in ADSCs pre-treated with AGEs (800 μg/mL; 24 h) treated with Cur (20 μM). Right: comparison of AGER and p-p65 expression levels between the study groups. ( b) ADSCs pre-treated with AGEs (800 μg/mL; 24 h) with or without Cur (20 μM; 24 h). Translocation of p65 from the cytoplasm to the nucleus was quantified by cell immunofluorescence staining (red arrowheads). ( c ) V-mCherry/caspase-3 stained ADSCs analyzed by flow cytometry. Relative fluorescence intensity was used to calculate cell apoptosis and caspase-3 expression level. Data are shown as means ± SD. Statistical analysis: * p <0.05, * * p <0.01 and * p <0.001. ( * p vs 0 h of AGER, # p vs 0 h of p-p65). NF-κB nuclear factor-κB, Cur curcumin, ADSC adipose-derived stem cell, AGE advanced glycation end product, AGER AGE receptor

Article Snippet: The protein was transferred to a PVDF membrane and the primary antibody was incubated at 4°C overnight, followed by incubation with the secondary antibody (BA1054, Boster, Wuhan, China) at room temperature for 2 h. The primary antibodies were rabbit AGER (#55222, 1:500; Cell Signaling Technology/CST, Beverly, MA, USA), p65 (#3033, 1:500; CST), p-p65 (#8242, 1:500; CST) and β-actin (ab8227, 1:1000; Abcam, Cambridge, UK).

Techniques: Expressing, Translocation Assay, Immunofluorescence, Staining, Flow Cytometry, Fluorescence, Derivative Assay

Journal: Journal of Immunology Research

Article Title: HMGB1 Promotes Systemic Lupus Erythematosus by Enhancing Macrophage Inflammatory Response

doi: 10.1155/2015/946748

Figure Lengend Snippet: HMGB1-enhanced macrophage inflammatory response induced by ALD-DNA might be dependent on RAGE but not on TLR2 and TLR4. (a-b) RAW264.7 cells were transfected with pHMGB1, and then stimulated with ALD-DNA (50 μ g/mL) in the presence of OxPAPC (30 μ g/mL) or RAGE-Fc (10 μ g/mL) for 24 h. The supernatants were collected and assayed for the concentrations of TNF- α (a) and IL-6 (b) using ELISA. (c) Representative immunoblot of three independent experiments has shown the efficiency of RAGE knockdown. (d-e) RAW264.7 cells transfected with siRAGE and pHMGB1 were stimulated with 50 μ g/mL of ALD-DNA for 24 h. The supernatants were collected and assayed for the concentrations TNF- α (d) and IL-6 (e) using ELISA. ∗ P < 0.05.

Article Snippet: HMGB1 and RAGE antibody were obtained from Cell Signaling Technology and GAPDH antibody from Santa Cruz Biotechnology.

Techniques: Transfection, Enzyme-linked Immunosorbent Assay, Western Blot