Journal: Nature Communications

Article Title: NF-κB inhibition rescues cardiac function by remodeling calcium genes in a Duchenne muscular dystrophy model

doi: 10.1038/s41467-018-05910-1

Figure Lengend Snippet: NF-κB causes heart dysfunction in mdx mice. a EMSA performed on wild-type (wt) and mdx hearts (left panel). Supershift EMSA performed on mdx hearts using specific antibodies for p65 and p50, and IgG as a control (Right panel). Arrowheads indicate shifted bands. b Western blots performed on whole heart lysates and probed for phosphorylated p65-ser536 (phospho-p65), p65, p50, and α-tubulin (used as a loading control). c Representative images of H&E and phosho-p65 staining prepared from 1-year old heart sections. Boxed regions appear as magnified images in neighboring panels. Scale bar = 50 μm. d cardiac output, e stroke volume, f ejection fraction ( p = 0.686), g end diastolic diameter, and h end diastolic volume assessed by echocardiogram on 13–14-month-old mice ( n = 4 wt; 8 mdx IKKβf/f ; 5 mdx HRTΔIKKβ ). i End-diastolic pressure volume relationship (EDPVR) assessed by ventricular pressure-volume relationship analysis on 13–14-month old mice ( n = 7 wt; 18 mdx IKKβf/f ; 12 mdx HRTΔIKKβ ). j Developed force measured from isolated multicellular cardiac muscles of 7-month old mice in response to β-adrenergic stimulation with isoproterenol ( n = 7 wt; 7 mdx IKKβf/f ; 9 mdx HRTΔIKKβ ; 3 NBD treated ( mdx -NBD); p = 0.278). k Relaxation time (RT 90 ) after isoproterenol stimulation in multicellular cardiac muscles ( n = same as J). l, m Ventricular pressure-volume relationship measurements after dobutamine administration on 13–14-month-old mice. l Maximal heart rate (HR) ( n = 5 wt; 8 mdx IKKβf/f ; 7 mdx HRTΔIKKβ ) and ( m ) Tau (isovolumetric relaxation) ( n = 5 wt; 7 mdx IKKβf/f ; 4 mdx HRTΔIKKβ ; p = 0.027 but multiple comparisons test did not detect differences between groups). Data expressed as means ± SEM with bars and plungers and individual data points with dots. * p < 0.05 relative to wt and # p < 0.05 relative to mdx IKKβf/f by d – i , k – m 1-way ANOVA followed by Tukey multiple comparison test where appropriate, j 2-way repeated measures ANOVA. Main effects for genotype/treatment

Article Snippet: For supershifts, extracts were incubated with the following antibodies: 4 μl p50 (114 Santa Cruz) or IgG or 0.5 μl p65 (Rockland) prior to loading on the gel.

Techniques: Control, Western Blot, Staining, Isolation, Muscles, Comparison

Journal: Nature Communications

Article Title: NF-κB inhibition rescues cardiac function by remodeling calcium genes in a Duchenne muscular dystrophy model

doi: 10.1038/s41467-018-05910-1

Figure Lengend Snippet: Cardiomyocyte NF-κB ablation normalizes calcium handling and increases gene expression. a Statistically significant gene categories from microarray analysis identified using Gene Set Enrichment Analysis. Heatmaps represent genes identified in annotations. b Calcium transient amplitude measured from cardiomyocytes isolated from 7–8-month old mice ( n = 38 wt; 43 mdx IKKβf/f ; 35 mdx HRTΔIKKβ cardiomyocytes). c Depiction of individual microarray genes that were up- and down-regulated in mdx HRTΔIKKβ relative to mdx IKKβf/f hearts. Genes shown in red are ≥ 1.5-fold upregulated and those in blue are ≥ 1.5-fold downregulated. d – g qPCR analysis of Slc8a1 expression. RNA isolated from d 6–7-month-old hearts ( n = 5 wt; 5 mdx IKKβf/f ; 4 mdx HRTΔIKKβ ), e mouse embryonic fibroblasts (MEFs) that were wt ( p65 +/+ ) or null ( p65 −/− ) for p65 ( n = 5). f C2C12 myotubes expressing empty vector as control (CT) or IκBα super repressor (SR) ( n = 6 CT; 8 SR), and g MEFs untreated (CT) or treated with TNF ( n = 4). Data expressed as means ± SEM with bars and plungers and individual data points with dots. * p < 0.05 relative to respective control and # p < 0.05 relative to mdx IKKβf/f by b, d 1-way ANOVA followed by Tukey multiple comparison test and e – g 2-tailed Student’s t test

Article Snippet: For supershifts, extracts were incubated with the following antibodies: 4 μl p50 (114 Santa Cruz) or IgG or 0.5 μl p65 (Rockland) prior to loading on the gel.

Techniques: Gene Expression, Microarray, Isolation, Expressing, Plasmid Preparation, Control, Comparison

Journal: Nature Communications

Article Title: NF-κB inhibition rescues cardiac function by remodeling calcium genes in a Duchenne muscular dystrophy model

doi: 10.1038/s41467-018-05910-1

Figure Lengend Snippet: Cardiomyocyte NF-κB ablation causes global H3K27ac enrichment in mdx hearts. a Venn diagram pie chart depicting the H3K27ac ChIP-seq annotation within specified regions of the genome from mouse hearts. b Genome-wide distribution of H3K27ac binding loci relative to transcription start sites (TSS). c – d Genome-wide fragment density showing potential overlap of c . ChIP-seq histone marks across peaks from our ChIP-seq performed in mdx HRTΔIKKβ hearts and d our ChIP-seqs across peaks from p65 ChIP-seq. e Network showing the top 15 Gene Ontology clusters identified from differentially enriched genes in the mdx HRTΔIKKβ when compared to mdx IKKβf/f H3K27 regions. The most significantly enriched pathway for each cluster is labeled as the representative term for that group. f Gene expression analyzed by qPCR on total RNA isolated from hearts ( n = Rcan1 : 5 wt; 6 mdx IKKβf/f ; 6 mdx HRTΔIKKβ ; Cacna1h : 5 for all genotypes; Camk4 5 wt; 6 mdx IKKβf/f ; 5 mdx HRTΔIKKβ ). g – h ChIP performed with an H3K27ac antibody and qPCR analysis was used to detect enrichment on denoted genes. DNA extracted from g mouse hearts ( n = 3) or h control and TNF treated MEFs ( n = 4 Slc8a1 and Rcan1 and 3 Cacna1h and Camk4 ). Genes were expressed as a ratio. Dotted line represents level of enrichment equal to g mdx IKKβf/f hearts and h control MEFs. Bars represent ( g ) enrichment in hearts and h depletion in TNF treated MEFs. f Data expressed using box and whiskers plots. The central line in the boxes is the median value; the lower and upper boundaries of the boxes represent the lower and upper quartiles, respectively; the lower and upper whiskers represent the minimum and maximum values, respectively. Individual data points are plotted with dots. g – h Data expressed as means ± SEM with bars and plungers and individual data points with dots. f * p < 0.05 wt and # p < 0.05 mdx IKKβf/f , by 1-way ANOVA followed by Tukey multiple comparison test. g – h * p < 0.05 mdx IKKβf/f or untreated MEFs by 2-tailed Student’s t test

Article Snippet: For supershifts, extracts were incubated with the following antibodies: 4 μl p50 (114 Santa Cruz) or IgG or 0.5 μl p65 (Rockland) prior to loading on the gel.

Techniques: ChIP-sequencing, Genome Wide, Binding Assay, Labeling, Gene Expression, Isolation, Control, Comparison

Journal: Nature Communications

Article Title: NF-κB inhibition rescues cardiac function by remodeling calcium genes in a Duchenne muscular dystrophy model

doi: 10.1038/s41467-018-05910-1

Figure Lengend Snippet: CTCF, SIN3A, and HDAC1 mediate a less permissive chromatin conformation on calcium genes upon NF-κB activation. a Motif analysis performed on genes identified as having both p65 ChIP-seq peaks and H3K27ac mdx HRTΔIKKβ ChIP-seq regions. b Pie graph representing the percentage of genes with CTCF motifs up- and downregulated in the microarray (relative to mdx hearts with intact NF-κB). c ChIP-seq data derived from genome-wide fragment density analysis showing potential overlap of CTCF peaks with p65 peaks. d The same analysis as in c except p65 peaks were split between two groups either containing or lacking an NF-κB consensus motif. e – f ChIP performed with a CTCF antibody and qPCR analysis was used to detect enrichment on denoted genes. DNA extracted from e mdx IKKβf/f and mdx HRTΔIKKβ hearts ( n = 4 Slc8a1 ; 5 Rcan1 ; 3 cacna1h ; 2 Camk4 ) and ( f ) control and TNF treated MEFs ( n = 4 except n = 2 Camk4 ), g – h The same analyses were performed as in e, f . DNA extracted from mdx IKKβf/f and mdx HRTΔIKKβ hearts and ChIP performed with a ( g ) SIN3A antibody ( n = 3 Slc8a1 and Cacna1h ( p = 0.7); n = 4 Rcan1 and Camk4 ) ( h ) HDAC1 antibody ( n = 3 except n = 4 Slc8a1 ). e – h Enrichment on different genes were plotted as a ratio. Dotted line represents level of enrichment equal to e, g – h mdx IKKβf/f hearts and ( f ) control MEFs. Bars represent e, g – h depletion in mdx HRTΔIKKβ hearts and f enrichment in TNF treated MEFs. i Gene expression analyzed by qPCR on total RNA isolated from HL-1 cardiomyocytes ( n = 4). j Heatmaps showing ChIP-seq fragment densities of SIN3A and HDAC1 surrounding p65 peaks, showing potential overlap. Left panel includes genes upregulated and right panel includes genes downregulated in the microarray. e – h Data expressed as means ± SEM with bars and plungers and individual data points with dots. ( i ) Data expressed using box and whiskers plots. The central line in the boxes is the median value; the lower and upper boundaries of the boxes represent the lower and upper quartiles, respectively; the lower and upper whiskers represent the minimum and maximum values, respectively. Individual data points are plotted with dots. e – h * p < 0.05 mdx IKKβf/f or untreated by 2-tailed Student’s t test. i * p < 0.05 CT and # p < 0.05 TSA by 1-way ANOVA followed by Tukey multiple comparison test

Article Snippet: For supershifts, extracts were incubated with the following antibodies: 4 μl p50 (114 Santa Cruz) or IgG or 0.5 μl p65 (Rockland) prior to loading on the gel.

Techniques: Activation Assay, ChIP-sequencing, Microarray, Derivative Assay, Genome Wide, Control, Gene Expression, Isolation, Comparison

Journal: JCI Insight

Article Title: NF- κ B represses retinoic acid receptor–mediated GPRC5A transactivation in lung epithelial cells to promote neoplasia

doi: 10.1172/jci.insight.153976

Figure Lengend Snippet: ( A ) Small interfering RNA (siRNA) targeting p65 and scramble control siRNA were transfected to Calu-1 cells, treated with or without TNF-α. The protein levels of GPRC5A and p65 were determined by Western blotting. ( B ) Calu-1 cells were transfected with plasmid overexpressing IκBα-AA mutant (S32A, S36A) or vector control, then treated with or without TNF-α; GPRC5A and IκBα protein levels were determined with specific antibodies through Western blotting. ( C ) Schematic representation of WT and truncation mutant of RelA/p65. ( D ) Calu-1 transfectants harboring vector control or inducible expression of FL and truncated p65 were established; cells were treated with doxycycline (300 ng/mL) for 24 hours, and GPRC5A protein levels were analyzed by Western blotting. These blots were run in separate gels performed in parallel with equal loading (please see uncropped/unedited gels in the supplement). ( E and F ) Calu-1 cells with inducible expression of WT p65 and serine 276A mutant were treated with doxycycline (300 ng/mL) for 24 hours; GPRC5A protein and mRNA levels were determined by Western blotting ( E ) and qPCR ( F ), respectively. Data are presented as mean ± SD from 3 independent experiments with duplicates and analyzed by 2-tailed Student’s t test, *** P < 0.001.

Article Snippet: LPS was purchased from MilliporeSigma; TNF-α was purchased from R&D Systems; doxycycline was purchased from Selleckchem; antibodies (catalog numbers in parentheses) against H3K9ac (ab4441) and myc-tag (ab9232) for ChIP were purchased from Abcam; antibodies against Gprc5a (sc-98885), IκBα (sc-371), GAPDH (sc-365062), RARα (C-20), RARβ (C-19), and GFP (sc-8334) were purchased from Santa Cruz Biotechnology; NF-κB p65 NLS antibody (for IHC, NBP2-24541) was purchased from Novus Biologicals; antibodies against p-p65 (S536; 3033), H3K27Ac (8173), and β-actin conjugated with HRP (12262) were purchased from Cell Signaling Technology; p-p65 (S276) antibody (D155005) was from BBI Life Sciences Corporation; RNA polymerase II antibody (05-623B) was from MilliporeSigma; antibodies against myc-tag (catalog 562-5) and Flag-tag (catalog PM020) were purchased from MBL International Corporation; and customized GPRC5A antibody was generated by Abmart.

Techniques: Small Interfering RNA, Control, Transfection, Western Blot, Plasmid Preparation, Mutagenesis, Expressing

Journal: JCI Insight

Article Title: NF- κ B represses retinoic acid receptor–mediated GPRC5A transactivation in lung epithelial cells to promote neoplasia

doi: 10.1172/jci.insight.153976

Figure Lengend Snippet: ( A and B ) Calu-1 cells were treated with TNF-α and CHX separately or in combination; IκBα protein and mRNA levels were determined by Western blotting and quantified with ImageJ (NIH) ( A ) and qPCR ( B ), respectively. Data are presented as the mean ± SD. ( C and D ) Calu-1 cells were treated with TNF-α and CHX separately or in combination; GPRC5A mRNA and protein levels were determined by qPCR ( C ) and Western blotting and quantified with ImageJ ( D ), respectively. Data are presented as the mean ± SD. ( E ) Three NF-κB binding sites (designated as letters A–C) on GPRC5A promoter–luc plasmid were mutated individually or in combination. The repression effect of p65 was determined by luciferase assay. ( F ) The RA response element (RARE) at the GPRC5A promoter–luc plasmid was mutated, and the repression effect of p65 was determined by luciferase assay. Data are presented as mean ± SD from 3 independent experiments with duplicates and analyzed by 2-tailed Student’s t test, * P < 0.05; ** P < 0.01; *** P < 0.001.

Article Snippet: LPS was purchased from MilliporeSigma; TNF-α was purchased from R&D Systems; doxycycline was purchased from Selleckchem; antibodies (catalog numbers in parentheses) against H3K9ac (ab4441) and myc-tag (ab9232) for ChIP were purchased from Abcam; antibodies against Gprc5a (sc-98885), IκBα (sc-371), GAPDH (sc-365062), RARα (C-20), RARβ (C-19), and GFP (sc-8334) were purchased from Santa Cruz Biotechnology; NF-κB p65 NLS antibody (for IHC, NBP2-24541) was purchased from Novus Biologicals; antibodies against p-p65 (S536; 3033), H3K27Ac (8173), and β-actin conjugated with HRP (12262) were purchased from Cell Signaling Technology; p-p65 (S276) antibody (D155005) was from BBI Life Sciences Corporation; RNA polymerase II antibody (05-623B) was from MilliporeSigma; antibodies against myc-tag (catalog 562-5) and Flag-tag (catalog PM020) were purchased from MBL International Corporation; and customized GPRC5A antibody was generated by Abmart.

Techniques: Western Blot, Binding Assay, Plasmid Preparation, Luciferase

Journal: JCI Insight

Article Title: NF- κ B represses retinoic acid receptor–mediated GPRC5A transactivation in lung epithelial cells to promote neoplasia

doi: 10.1172/jci.insight.153976

Figure Lengend Snippet: ( A ) RARβ in H157 cells was knocked down by siRNA followed by treatment with or without ATRA. The mRNA levels of RARβ and GPRC5A were determined by RT-PCR and quantified by ImageJ. ( B and C ) RARβ in H157 cells was knocked down by RARβ short hairpin (sh) RNA followed by treatment with or without ATRA. Protein and mRNA levels of RARβ and GPRC5A were determined by Western blotting ( B ) and RT-PCR and quantified by ImageJ ( C ), respectively. ( D and E ) Calu-1 cells were treated with TNF-α and ATRA separately or in combination, and the GPRC5A protein and mRNA levels were determined by Western blotting ( D ) and RT-PCR and quantified by ImageJ ( E ). ( F ) H157 cells was treated with TNF-α and ATRA separately or in combination. GPRC5A mRNA levels were determined by RT-PCR and quantified by ImageJ. ( G ) HEK293T cells were transfected with GPRC5A-luc and p65 plasmids and treated with or without ATRA. The p65 repression effect was determined by luciferase assay. Data are presented as mean ± SD from 3 independent experiments with duplicates and analyzed by 2-tailed Student’s t test, * P < 0.05; *** P < 0.001.

Article Snippet: LPS was purchased from MilliporeSigma; TNF-α was purchased from R&D Systems; doxycycline was purchased from Selleckchem; antibodies (catalog numbers in parentheses) against H3K9ac (ab4441) and myc-tag (ab9232) for ChIP were purchased from Abcam; antibodies against Gprc5a (sc-98885), IκBα (sc-371), GAPDH (sc-365062), RARα (C-20), RARβ (C-19), and GFP (sc-8334) were purchased from Santa Cruz Biotechnology; NF-κB p65 NLS antibody (for IHC, NBP2-24541) was purchased from Novus Biologicals; antibodies against p-p65 (S536; 3033), H3K27Ac (8173), and β-actin conjugated with HRP (12262) were purchased from Cell Signaling Technology; p-p65 (S276) antibody (D155005) was from BBI Life Sciences Corporation; RNA polymerase II antibody (05-623B) was from MilliporeSigma; antibodies against myc-tag (catalog 562-5) and Flag-tag (catalog PM020) were purchased from MBL International Corporation; and customized GPRC5A antibody was generated by Abmart.

Techniques: Reverse Transcription Polymerase Chain Reaction, Western Blot, Transfection, Luciferase

Journal: JCI Insight

Article Title: NF- κ B represses retinoic acid receptor–mediated GPRC5A transactivation in lung epithelial cells to promote neoplasia

doi: 10.1172/jci.insight.153976

Figure Lengend Snippet: ( A and B ) Calu-1 cells were treated with TNF-α (10 ng/mL) for various time points. The change of p65 and other proteins as indicated in the figure binding to the IκBα promoter ( A ) and GPRC5A promoter ( B ) was analyzed by chromatin immunoprecipitation (ChIP) assay using corresponding specific antibodies. Input as positive control and normal IgG (N IgG) as negative control. ( C and D ) The interaction of p65 with RARα ( C ) and RARβ2/β4 ( D ) was determined by immunoprecipitation (IP) assay. ( E ) RARα-Flag–expressing plasmid was cotransfected with GFP/GFP-p65/GFP-p65-S276A mutant to HEK293T cells. After 48 hours, cells were lysed by RIPA, and IP assay was performed to detect the interaction between RARα and WT RelA/p65 or serine 276A mutant. ( F ) Representative images of IF analysis of Calu-1 cells transfected with RARα-Flag plus GFP-p65 or GFP-p65-S276A as indicated. Scale bar: 10 μm.

Article Snippet: LPS was purchased from MilliporeSigma; TNF-α was purchased from R&D Systems; doxycycline was purchased from Selleckchem; antibodies (catalog numbers in parentheses) against H3K9ac (ab4441) and myc-tag (ab9232) for ChIP were purchased from Abcam; antibodies against Gprc5a (sc-98885), IκBα (sc-371), GAPDH (sc-365062), RARα (C-20), RARβ (C-19), and GFP (sc-8334) were purchased from Santa Cruz Biotechnology; NF-κB p65 NLS antibody (for IHC, NBP2-24541) was purchased from Novus Biologicals; antibodies against p-p65 (S536; 3033), H3K27Ac (8173), and β-actin conjugated with HRP (12262) were purchased from Cell Signaling Technology; p-p65 (S276) antibody (D155005) was from BBI Life Sciences Corporation; RNA polymerase II antibody (05-623B) was from MilliporeSigma; antibodies against myc-tag (catalog 562-5) and Flag-tag (catalog PM020) were purchased from MBL International Corporation; and customized GPRC5A antibody was generated by Abmart.

Techniques: Binding Assay, Chromatin Immunoprecipitation, Positive Control, Negative Control, Immunoprecipitation, Expressing, Plasmid Preparation, Mutagenesis, Transfection

Journal: JCI Insight

Article Title: NF- κ B represses retinoic acid receptor–mediated GPRC5A transactivation in lung epithelial cells to promote neoplasia

doi: 10.1172/jci.insight.153976

Figure Lengend Snippet: ( A ) GPRC5A protein expression level in multiple human NSCLC cell lines and normal human bronchial epithelial cell line (16HBE) was analyzed by Western blotting. ( B ) Calu-1 cells treated with or without TNF-α (10 ng/mL) for 12 hours. Binding of RNA polymerase II and the histone modification at the GPRC5A promoter were analyzed by ChIP using specific antibodies. ( C and D ) Calu-1 cells with inducible expression of WT and serine 276A mutant p65 were treated with doxycycline (300 ng/mL) for 12 hours; the change of RNA polymerase II binding ( C ) and histone modification (H3K9ac) at the GPRC5A promoter ( D ) were analyzed by ChIP. Input as positive control and normal IgG (N IgG) as negative control. ( E – H ) A549, H1975, and Calu-1 cells were treated with 5-Aza-dc (1 μM, 4 days) or SAHA (2.5 μM, 24 hours) individually or in combination; GPRC5A protein ( E – G ) and mRNA levels ( H ) were analyzed via Western blotting and qPCR. Data are presented as the mean ± SD. ( I ) Calu-1 cells were treated with or without SAHA (2.5 μM) for 12 hours; RNA polymerase II binding at the GPRC5A promoter was analyzed by ChIP. ( J ) Normal human bronchial epithelial cell line (16HBE) and multiple human NSCLC cell lines were treated with or without SAHA (2.5 μM, 24 hours); GPRC5A protein levels were analyzed by Western blotting. ( K ) Calu-1 cells were pretreated with DMSO (as vehicle control), 5-Aza-dc (1 μM, 3 days) or SAHA (2.5 μM, 3 hours) followed by TNF-α (10 ng/mL) treatment for an additional 24 hours. GPRC5A protein levels were analyzed by Western blotting. All data are presented as mean ± SD from 3 independent experiments with duplicates and analyzed by 2-tailed Student’s t test, ** P < 0.01; *** P < 0.001. H3K9ac, acetylated histone H3K9.

Article Snippet: LPS was purchased from MilliporeSigma; TNF-α was purchased from R&D Systems; doxycycline was purchased from Selleckchem; antibodies (catalog numbers in parentheses) against H3K9ac (ab4441) and myc-tag (ab9232) for ChIP were purchased from Abcam; antibodies against Gprc5a (sc-98885), IκBα (sc-371), GAPDH (sc-365062), RARα (C-20), RARβ (C-19), and GFP (sc-8334) were purchased from Santa Cruz Biotechnology; NF-κB p65 NLS antibody (for IHC, NBP2-24541) was purchased from Novus Biologicals; antibodies against p-p65 (S536; 3033), H3K27Ac (8173), and β-actin conjugated with HRP (12262) were purchased from Cell Signaling Technology; p-p65 (S276) antibody (D155005) was from BBI Life Sciences Corporation; RNA polymerase II antibody (05-623B) was from MilliporeSigma; antibodies against myc-tag (catalog 562-5) and Flag-tag (catalog PM020) were purchased from MBL International Corporation; and customized GPRC5A antibody was generated by Abmart.

Techniques: Expressing, Western Blot, Binding Assay, Modification, Mutagenesis, Positive Control, Negative Control, Control

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Microbiota-driven therapeutic efficacy of Hyperoside in ulcerative colitis and associated anxiety

doi: 10.3389/fcimb.2026.1734356

Figure Lengend Snippet: Network pharmacology and molecular docking elucidate potential mechanisms of HYP in UC treatment. (A) Venn diagram of HYP and UC target genes. (B) PPI network of common targets. (C) KEGG pathway analysis. (D) The top 20 gene targets ranked by degree value. Molecular docking binding modes and affinities of HYP with (E) MAPK3, (F) AKT1, and (G) NFκB1. (H) Protein expression levels of p38MAPK, p-p38MAPK, AKT, p-AKT, NF-κB p65, and p-NF-κB p65 in the colon tissues.

Article Snippet: The following antibodies were used in this study: anti-ZO-1 antibody (AF5145, Affinity), anti-mucin-2 antibody (GB11344, Serbicebio), anti-BDNF antibody (GB11559, Serbicebio), anti-GFAP antibody (GB11096, Serbicebio), anti-Iba-1 antibody ( GB154490 , Serbicebio), NF-κB p65 (Proteintech, 10745-1-AP), Phospho-NF-κB p65 (Affinity, AF2006), AKT (Proteintech, 10176-2-AP), Phospho-AKT (Proteintech, 28731-1-AP), p38MAPK (Proteintech, 51115-1-AP), Phospho-p38MAPK (Affinity, AF4001).

Techniques: Binding Assay, Expressing

Journal: Frontiers in Cellular and Infection Microbiology

Article Title: Microbiota-driven therapeutic efficacy of Hyperoside in ulcerative colitis and associated anxiety

doi: 10.3389/fcimb.2026.1734356

Figure Lengend Snippet: Schematic of the mechanism by which HYP ameliorates DSS-induced UC and associated anxiety-like behaviors. HYP alleviates UC and comorbid anxiety through gut microbiota-dependent mechanisms. It remodels the gut microbial community, thereby enhancing host arginine metabolism and directly modulating the MAPK/PI3K-Akt/NF-κB signaling pathways. This integrated action suppresses systemic and neuroinflammation, restores intestinal barrier function, and ultimately ameliorates both colonic pathology and anxiety-like behaviors.

Article Snippet: The following antibodies were used in this study: anti-ZO-1 antibody (AF5145, Affinity), anti-mucin-2 antibody (GB11344, Serbicebio), anti-BDNF antibody (GB11559, Serbicebio), anti-GFAP antibody (GB11096, Serbicebio), anti-Iba-1 antibody ( GB154490 , Serbicebio), NF-κB p65 (Proteintech, 10745-1-AP), Phospho-NF-κB p65 (Affinity, AF2006), AKT (Proteintech, 10176-2-AP), Phospho-AKT (Proteintech, 28731-1-AP), p38MAPK (Proteintech, 51115-1-AP), Phospho-p38MAPK (Affinity, AF4001).

Techniques: Protein-Protein interactions

Journal: The Journal of allergy and clinical immunology

Article Title: ANKRD1 regulates innate immune responses against herpes simplex virus 1: A potential role in eczema herpeticum

doi: 10.1016/j.jaci.2018.01.001

Figure Lengend Snippet: The indicated expression plasmids were transfected into 293 FT cells and incubated overnight. The cells were then harvested and co-IP experiments were performed. Flag/DDK1-tagged protein expression and HA-tagged ANKRD1 protein expression are shown in IP samples and lysates. Data presented as one of three independent experiments.

Article Snippet: Myc-DDK1-tagged RELA (RC220780), Myc-DDK1-tagged NFKB1 (RC208384), Myc-DDK1-tagged IRF7 (RC217934), Myc-DDK1-tagged ANKRD1 (RC205609) and Myc-DDK1-tagged STING were purchased from OriGene (Rockville, MD).

Techniques: Expressing, Transfection, Incubation, Co-Immunoprecipitation Assay

Journal: Clinical, Cosmetic and Investigational Dermatology

Article Title: A Topical Chinese Herbal Alleviates Psoriasis by Regulating Keratinocytes Pyroptosis Through Inhibition of NLRP3 Inflammasome Activation

doi: 10.2147/CCID.S559712

Figure Lengend Snippet: ( A – C ) mRNA levels of NLRP3, Caspase-1, GSDMD in NHEK cells, n = 3. ( D – I ) Protein levels of NF-κB, P65, NLRP3, caspase-1, GSDMD in NHEK cells, n = 3.*P < 0.05, **P < 0.01, ***P < 0.001 compared with Control; # P < 0.05, ## P < 0.01, ### P < 0.001 compared with Model; & P < 0.05, && P < 0.01, &&& P < 0.001 compared with QS; ^P < 0.05, ^^P < 0.01 compared with Mcc950.

Article Snippet: The sections were then incubated overnight at 4°C with the following primary antibodies: NF-κB p65 Monoclonal antibody (66535-1-Ig, Proteintech, China), NLRP3 Monoclonal antibody (68102-1-Ig-100, Proteintech, China), Caspase-1 antibody (sc-56036, Santa Cruz, USA), and GSDMD Polyclonal antibody (20770-1-AP, Proteintech, China).

Techniques: Control

Journal: International Journal of Molecular Sciences

Article Title: Resveratrol Modulates Chemosensitisation to 5-FU via β1-Integrin/HIF-1α Axis in CRC Tumor Microenvironment

doi: 10.3390/ijms24054988

Figure Lengend Snippet: Resveratrol’s reduction of inflammation, vascularisation as well as cancer stemness and elevation of apoptosis via β1-integrin receptors in HCT-116/HCT-116R cells shown by Western blot analysis. X -axis: HCT-116 ( A ) and HCT-116R ( B ) cells in alginate drops were left untreated alone (Co.) or in TME, where they were left untreated or were treated with 2 nM 5-FU, 5 µM resveratrol, 0.5 µM β1-SO, 0.5 µM β1-ASO or combinations thereof. Samples were immunoblotted with antibodies against NF-kB (unphosphorylated NF-kB), p-NF-kB (phosphorylated NF-kB), cleaved-caspase-3, HIF-1α, VEGF, CD44, CD133, ALDH1 and β-actin (loading control). Y -axis shows densitometric units. Relative to TME control, values were p < 0.05 (⋆) and p < 0.01 (⋆⋆).

Article Snippet: The monoclonal antibodies against phospho-p65-NF-kB (#MAB7226), p65-NF-kB (#MAB5078) as well as polyclonal anti-cleaved-caspase-3 (#AF835) were acquired from R&D Systems (Heidelberg, Germany), while monoclonal antibody against β1-integrin (#14-0299-82) was from Thermo Fisher Scientific (Langenselbold, Germany).

Techniques: Western Blot, Control