Journal: Cell Death and Differentiation

Article Title: Suppressor of hepatocellular carcinoma RASSF1A activates autophagy initiation and maturation

doi: 10.1038/s41418-018-0211-7

Figure Lengend Snippet: RASSF1A interacts with HDAC6 to enhance acetylation of α-Tubulin and co-localizes with acetylated microtubules. a , b Representative immunoblot ( a ) and quantification ( b ) showing levels of acetylated α-tubulin (Ac-α-Tub) in liver tissues from 4-month-old wild-type and RASSF1A −/− mice. c , d Representative immunoblot ( c ) and quantification ( d ) showing levels of acetylated α-tubulin in hepatocytes isolated from wild-type and RASSF1A −/− mice. e , f Representative immunoblot ( e ) and quantification ( f ) showing levels of acetylated α-tubulin in HeLa cells transiently transfected with control plasmid, plasmid expressing HA-RASSF1A (HA-RA), and plasmid expressing HA-RASSF1A∆ (HA-RA∆). g , h Representative images ( g ) and quantification ( h ) showing the immunostaining intensities of acetylated α-tubulin in cells similar to those in e . Bar = 20 µm. Red, HA-RA or HA-RA∆; green, acetylated a-tubulin; and yellow, colocalization. i , j Representative immunoblot ( i ) and quantification ( j ) showing the impact of RASSF1A on levels of HDAC6 in cells similar to those in e . k , l Representative immunoblots showing levels of Flag-HDAC6 coimmunoprecipitated with endogenous RASSF1A from HeLa cells ( k ) or exogenous RASSF1A in 293T cells ( l ) transiently overexpressing Flag-HDAC6. m Representative immunoblots showing levels of RA-HA or RA-RA∆ coimmunoprecipitated with Flag-HDAC6 in 293T cells. n Representative immunoblot showing levels of GFP fused fragments of RASSF1A coimmunoprecipitated with HDAC6 from lysates of HeLa cells transiently expressing GFP fused RASSF1A constructs

Article Snippet: Plasmids encoding Myc-LC3 (#24919), HDAC6 (#30482) and P27 (#14049) were purchased from Addgene.

Techniques: Western Blot, Isolation, Transfection, Control, Plasmid Preparation, Expressing, Immunostaining, Construct

Journal: Cellular and Molecular Gastroenterology and Hepatology

Article Title: Continuous Activation of C/EBPβ Transcription Factor Prevents Fibrosis Resolution After Alcohol Cessation

doi: 10.1016/j.jcmgh.2025.101525

Figure Lengend Snippet: C/EBPβ inhibition promotes proresolving phenotype in liver macrophages. ( A ) WT or Cebpb KO hepatocytes were cocultured with WT liver macrophages in a Transwell coculture system. Macrophage collagen degradation activity, n = 4 per group. ∗∗ P < .01. ( B ) RNA-seq data of Cyp3a gene expression fold change in WT mice during resolution (Res/WDA) and Cebpb KO mice during resolution (KO_Res/WDA). ( C ) Relative mRNA of Cyp3a16 and Cyp3a41 ( Cyp3a41a+ Cyp3a41b ) in WT and KO mice at 4 weeks after stopping alcohol. ( D ) Schematic of Cyp3a gene cluster showing ATAC peaks position and gRNA position. ( Bottom ) Differentially accessible peaks in hepatocytes from resolution condition compared with control hepatocytes. ( E ) Primary hepatocytes were transfected with dCas9-p300 vector and single gRNA targeting Cyp3a region or scrambled control. Relative mRNA expression in hepatocytes. ( F ) After 48 hours of incubation hepatocyte conditioned media was used for HDL isolation. Liver macrophages were treated with isolated HDL from hepatocytes expressing scr control RNA or Cyp3a targeting RNA. Relative mRNA in macrophages, n = 4–8 per group. ∗ P < .05; ∗∗ P < .01. ( G ) Macrophages were tested in a collagen degradation assay. n = 5 per group. ∗ P < .05. ( H ) Macrophages were treated with 10 μM of corresponding oxysterols. ( Right ) Relative mRNA in macrophages, n = 4–8 per group. ∗∗ P < .01. ( Right ) Whole liver LXRα target gene expression log2Fold (KO/WT). ( Bottom ) Correlation between CYP3A7 and LRXα target genes in human liver (TCGA + GTEx) samples. ( I ) Macrophages were treated with 10 μM of corresponding oxysterols. After 24 hours of culture macrophages were tested in a collagen degradation assay. ( J ) Relative mRNA in macrophages, n = 4–8 per group. ∗ P < .05; ∗∗ P < .01. ( K ) Correlation between CYP3A7 and proresolving macrophage marker genes in human liver (TCGA + GTEx) samples.

Article Snippet: AAV-TBG-control and AAV-TBG-iCre were from VectorBiolabs (Malvern, PA) and were used at 1 x 10 11 genome copies per mouse (Cre/control). dCas9-p300 plasmid was from Addgene (pcDNA-dCas9-p300 Core, Cat# 61357).

Techniques: Inhibition, Activity Assay, RNA Sequencing, Gene Expression, Control, Transfection, Plasmid Preparation, Expressing, Incubation, Isolation, Degradation Assay, Targeted Gene Expression, Marker