Journal: Journal of Hepatocellular Carcinoma

Article Title: Inhibition of Annexin A10 Contributes to ZNF281 Mediated Aggressiveness of Hepatocellular Carcinoma

doi: 10.2147/JHC.S400989

Figure Lengend Snippet: ZNF281 was increased expression in hepatocellular carcinoma (HCC). ( A ) Immunohistochemistry of ZNF281 in HCC tissues in comparison with corresponding paracancerous tissues (left: 4×, right: 20× objective). ( B ) Quantification from A (n=3; **p < 0.01). ( C ) Comparison of mRNA level of ZNF281 from HCC and corresponding paracancerous tissues of 14 HCC patients. ( D ) Comparison of ZNF281 mRNA level between HCC patients and the non-cancer control in the TCGA liver cancer and the GTEx datasets. ( E ) Comparison of ZNF281 mRNA level between HCC patients and normal control of Chen’s liver in the Oncomine database. ( F ) Protein level of ZNF281 in commonly used normal liver or HCC cell lines.

Article Snippet: In addition, of the 25 HCC cell lines recorded in the CCLE (Broad Institute Cancer Cell Line Encyclopedia) dataset, 16 lines showed high levels for ZNF281, but low levels of ANXA10, including Huh7, HLE and HepG2 we used, also supporting negative regulation between them ( Figure S6 ).

Techniques: Expressing, Immunohistochemistry, Comparison, Control

Journal: Journal of Hepatocellular Carcinoma

Article Title: Inhibition of Annexin A10 Contributes to ZNF281 Mediated Aggressiveness of Hepatocellular Carcinoma

doi: 10.2147/JHC.S400989

Figure Lengend Snippet: Knockdown of ZNF281 inhibited migration and invasion in HCC cells with retardation of EMT. ( A ) Western blot of ZNF281 in shRNA (ZNF281.kd1 or ZNF281.kd2) stably infected HLE or Huh7 cells. ( B and C ) Wound healing assays (10×) in ZNF281 knockdown HLE or Huh7 cells. ( D and E ) Quantification from ( B ) and ( C ) were shown in mean ± SEM (n=3; **p < 0.01, and ***p < 0.001). ( F and G ) Matrigel transwell assays (20×) in ZNF281 knockdown HLE and Huh7 stable cells. ( H and I ) Quantification (n=3; **p < 0.01) from ( F and G ). ( J and K ) The mRNA levels of EMT markers were determined by RT-qPCR (n=3; *p < 0.05, **p < 0.01 and ***p < 0.001). ( L ) Representative phase contrast (P/C) pictures of HLE cells transfected with shRNAs against ZNF281. Bar= 100 μm. ( M ) Immunofluorescence observed under confocal microscopy of mesenchymal marker Vimentin and epithelial marker ZO-1 in HLE cells following ZNF281 depletion with ZNF281.kd1. (bar=10 μm).

Article Snippet: In addition, of the 25 HCC cell lines recorded in the CCLE (Broad Institute Cancer Cell Line Encyclopedia) dataset, 16 lines showed high levels for ZNF281, but low levels of ANXA10, including Huh7, HLE and HepG2 we used, also supporting negative regulation between them ( Figure S6 ).

Techniques: Knockdown, Migration, Western Blot, shRNA, Stable Transfection, Infection, Quantitative RT-PCR, Transfection, Immunofluorescence, Confocal Microscopy, Marker

Journal: Journal of Hepatocellular Carcinoma

Article Title: Inhibition of Annexin A10 Contributes to ZNF281 Mediated Aggressiveness of Hepatocellular Carcinoma

doi: 10.2147/JHC.S400989

Figure Lengend Snippet: RNA-seq screening for potential targets of ZNF281 in HCC cells. (A and B) Bioinformatic analyses of down-regulated ( B ) or up-regulated ( C ) genes upon ZNF281 with Metascape online tools. (C) Identification of ANXA10 from the heatmap including top 40 DEGs. ( D and E ) Validation by RT-qPCR of selected DEGs in stable shZNF281 expressing HLE or Huh7 cells. (F) Western blotting confirmation of ZNF281 and ANXA10 expression in HLE cells with stable depletion of ZNF281. (G) Immunofluorescence for ZNF281 and ANXA10in HLE or Huh7 cells following ZNF281 depletion (bar=25 μm).

Article Snippet: In addition, of the 25 HCC cell lines recorded in the CCLE (Broad Institute Cancer Cell Line Encyclopedia) dataset, 16 lines showed high levels for ZNF281, but low levels of ANXA10, including Huh7, HLE and HepG2 we used, also supporting negative regulation between them ( Figure S6 ).

Techniques: RNA Sequencing, Biomarker Discovery, Quantitative RT-PCR, Expressing, Western Blot, Immunofluorescence

Journal: Journal of Hepatocellular Carcinoma

Article Title: Inhibition of Annexin A10 Contributes to ZNF281 Mediated Aggressiveness of Hepatocellular Carcinoma

doi: 10.2147/JHC.S400989

Figure Lengend Snippet: Knockdown of ANXA10 attenuated the effect of ZNF281 depletion on inhibiting migration, invasion and metastasis of HCC cells. (A) Western blot for ANXA10 expression upon ANXA10 depletion in ZNF281kd.1 HCC cells. (B and C) Wound healing assays (10×) in ZNF281kd.1 HLE or Huh7 cells subjected to ANXA10 siRNA. (D and E) Quantification of ( B and C ) as mean ± SEM (n=3; *p < 0.05, ***p < 0.001). (F and G) Matrigel transwell assays (4×) in ZNF281 knockdown HLE and Huh7 cells transfected with scramble control or ANXA10 siRNA. The quantification was shown in the right panel (n=3; ***p < 0.001). (H) Expression of ANXA10 in HLE cells with different shRNA transfection as indicated. (I) Resected lungs for indicated groups. Arrows indicated the pulmonary metastatic nodules with macroscopic observation. (J) Statistical analysis of the pulmonary nodules in different groups as indicated (*p < 0.05, ***p < 0.001). (K and M) Representative images of HE staining for the resected lungs. Scale bars: 500 μm.

Article Snippet: In addition, of the 25 HCC cell lines recorded in the CCLE (Broad Institute Cancer Cell Line Encyclopedia) dataset, 16 lines showed high levels for ZNF281, but low levels of ANXA10, including Huh7, HLE and HepG2 we used, also supporting negative regulation between them ( Figure S6 ).

Techniques: Knockdown, Migration, Western Blot, Expressing, Transfection, Control, shRNA, Staining

Journal: Journal of Hepatocellular Carcinoma

Article Title: Inhibition of Annexin A10 Contributes to ZNF281 Mediated Aggressiveness of Hepatocellular Carcinoma

doi: 10.2147/JHC.S400989

Figure Lengend Snippet: ANXA10 inhibited the migration and invasion with suppressed expression of EMT markers in HCC cells. (A) ANXA10 protein levels in HLE cells transfected with siRNAs against ANXA10 (siANXA10.1 and siANXA10.2) or ANXA10 overexpression plasmid. (B and C) Microscopic fields (10×) from wound healing assays in ANXA10 knockdown or ANXA10 overexpression HLE cells. (D and E) Quantification of ( B and C ) plotted as mean ± SEM (n=3; **p < 0.01 and ***p < 0.001). (F and G) Matrigel transwell assays (4×) in HLE cells with ANXA10 knockdown or overexpression. (H and I) Quantification of ( F ) and ( G ) (n=3; **p < 0.01, and ***p < 0.001). (J and K) Normalized mRNA levels of EMT markers in ANXA10 knockdown or overexpression HLE cells by RT-qPCR (n=3; *p < 0.05, **p < 0.01, and ***p < 0.001). (l) Immunofluorescence of Vimentin in HLE with ANXA10 overexpression (bar=10 μm).

Article Snippet: In addition, of the 25 HCC cell lines recorded in the CCLE (Broad Institute Cancer Cell Line Encyclopedia) dataset, 16 lines showed high levels for ZNF281, but low levels of ANXA10, including Huh7, HLE and HepG2 we used, also supporting negative regulation between them ( Figure S6 ).

Techniques: Migration, Expressing, Transfection, Over Expression, Plasmid Preparation, Knockdown, Quantitative RT-PCR, Immunofluorescence

Journal: Aging Cell

Article Title: Liver osteopontin is required to prevent the progression of age‐related nonalcoholic fatty liver disease

doi: 10.1111/acel.13183

Figure Lengend Snippet: Liver OPN is p53 regulated. (a) Protein levels of p53 were evaluated by immunohistochemistry in liver sections of 3‐, 10‐, and 20‐month‐old (m) wild‐type (WT) mice ( n = 3–5). (b) OPN protein levels from liver homogenates were measured in WT and p53‐KO male and female mice fed a chow diet (CD) and a high‐fat diet (HFD) by immunoblotting using glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) as loading control ( n = 4–6). (c) Liver p53 and OPN protein levels were evaluated in WT mice fed a HFD injected with p53‐dominant positive adenovirus (adp53) and GFP (adGFP), using transferrin or GAPDH as a loading control ( n = 4–6). (d) Protein levels of p53 and OPN in siCtrl and sip53 HepG2 cells were measured by immunoblotting using GAPDH as loading control. Extracellular OPN was measured using an ELISA ( n = 4–5). (e) OPN protein levels from HepG2 cells silenced for p53 and Hep3B cells treated with palbociclib (Palbo) were measured by immunoblotting using GAPDH as loading control. OPN media levels from Hep3B cells treated either with vehicle (Veh) or with palbociclib (Palbo) were measured by ELISA ( n = 4–8). Values are means ± SEM . Significant differences are denoted by * p < 0.05, ** p < 0.01, and *** p < 0.001 (Student's t test)

Article Snippet: Data obtained from the Cell Line Encyclopedia (© 2019 The Broad Institute of MIT & Harvard) showed that the cell line with the least p53 expression, the Hep3B, also had the lowest OPN expression.

Techniques: Immunohistochemistry, Western Blot, Control, Injection, Enzyme-linked Immunosorbent Assay

Journal: British Journal of Cancer

Article Title: Inhibiting CBX4 efficiently protects hepatocellular carcinoma cells against sorafenib resistance

doi: 10.1038/s41416-020-01240-6

Figure Lengend Snippet: a The diagram illustrates how miR424 was identified from the miRNA sequencing data and predicted by Venn screening from miRNA database websites. b Sequence alignment of the human miR424 seed sequence with the 3’-UTR of CBX4. The mutated sequence in the matched binding sites for the gene that was used to create the firefly luciferase reporter constructs is shown at the bottom of the gene set. A luciferase reporter assay demonstrated that miR424 inhibited the transcription of the wild-type but not the mutant 3’-UTRs of CBX4. c The expression of endogenous CBX4 was inhibited in miR424-overexpressed Huh7-SR and PLC-SR cells. d In contrast, CBX4 levels were increased in Huh7 and PLC cells with miR424-TUD. All data were compared with the respective controls, and the mRNA level was detected by qRT-PCR. CBX4 mRNA expression was normalised to that of GAPDH mRNA; and three independent experiments were conducted. e Relative miR424 expression in HCC tissues and matched adjacent normal tissues as assessed by qRT-PCR. f , g Relative expression data of miR424 in HCC cases were further analysed. The negative relationship between miR424 expression and liver cirrhosis ( f ) and size ( g ). h , i Kaplan–Meier curves of disease-free survival (DFS) ( h ) and overall survival (OS) ( i ). Survival of the high and low miR424 expression groups assessed using log-rank (Mantel–Cox) test in HCC, which were divided according to a cut-off of 2.5, the median value of CBX4 mRNA expression relative to GAPDH mRNA. j From the TCGA database, tumour purity was highly negatively correlated with miR424 expression. k Linear regression and correlation between the miR424 and CBX4 mRNA levels in 341 HCC tissues from the TCGA database. l The negative linear regression and correlation analysis for the relation of the mRNA levels of CBX4 and those of miR424 in 106 HCC patients by qRT-PCR.

Article Snippet: The human HCC cell lines PLC (catalogue number: CRL-8024TM) was obtained from the Global Bioresource Center of American Type Culture Collection (ATCC) and Huh7 (catalogue number: JCRB0403) was purchased from Health Science Research Resources Bank (Osaka, Japan).

Techniques: Sequencing, Binding Assay, Luciferase, Construct, Reporter Assay, Mutagenesis, Expressing, Quantitative RT-PCR

Journal: Molecular Cancer

Article Title: Upregulation of microRNA-122 by farnesoid X receptor suppresses the growth of hepatocellular carcinoma cells

doi: 10.1186/s12943-015-0427-9

Figure Lengend Snippet: The level of FXR is positively correlated with that of miR-122. a and b The expression of FXR mRNA ( a ) and mature miR-122 ( b ) in 20 human HCC tissues and the corresponding adjacent noncancerous tissues was detected by qRT-PCR. c The correlation between the levels of FXR and miR-122 in HCC tissues was analyzed using Pearson’s test ( R 2 = 0.61, P < 0.01). d The expression of FXR mRNA and mature miR-122 in HCC cell lines (HepG2, Hep3B, Huh7, PLC, SMMC-7721, MHCC97L and MHCC97H) and hepatic cell line L02 was assayed by qRT-PCR. e The correlation between the levels of FXR and miR-122 in HCC cell lines was analyzed using Pearson’s test ( R 2 = 0.95, P < 0.01). β-actin was used as a control for FXR examination, while U6 snRNA as a control for miR-122 detection

Article Snippet: The other HCC cell lines including Huh7, PLC, SMMC-7721, MHCC97L and MHCC97H, and hepatic cell line L02 were purchased from China Center for Type Culture Collection (Wuhan, China).

Techniques: Expressing, Quantitative RT-PCR, Control