Journal: Hepatology (Baltimore, Md.)

Article Title: Tuning T-Cell Receptor Affinity to Optimize Clinical Risk-Benefit When Targeting Alpha-Fetoprotein-Positive Liver Cancer.

doi: 10.1002/hep.30477

Figure Lengend Snippet: FIG. 3. Functional analysis of AFP TCR mutant panels by IFN-γ ELISpot. Bars represent mean numbers of responding T cells in at least 6 donors, each tested in triplicate, with the standard error for interdonor variation. Red: HLA-A*02:01+AFP+ HCC cell lines (HepG2, JHH5.A2, and HuH6). Blue: HLA-A*02-AFP+ HCC cell lines (JHH5, Hep3B). Green: normal primary hepatocytes (HEP2, HEP3). TCRs are arranged by increasing affinity from left to right. (A) wt TCR AFPc239 and initial panel of mutants. Data are representative of 2 donors, showing mean ± SEM in triplicate wells. (B) Refined panel of enhanced affinity TCRs against HCC cell lines and normal liver cells. Abbreviation: ntd, nontransduced T cells.

Article Snippet: K8002 and K8008; DAKO, Ely, UK), according to protocols supplied by the manufacturer. taRget Cell lINeS The HCC lines, HuH6, JHH-5, and JHH-4, were purchased from the Japanese Collection of Research Bioresources Cell Bank (Osaka, Japan), and Hep3B cells were obtained from Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH (Braunschweig, Germany).

Techniques: Functional Assay, Mutagenesis, Enzyme-linked Immunospot

Journal: Cell Death & Disease

Article Title: NUPR1, a new target in liver cancer: implication in controlling cell growth, migration, invasion and sorafenib resistance

doi: 10.1038/cddis.2016.175

Figure Lengend Snippet: NUPR1 regulates cell viability, growth, migration and invasion of HCC cells. ( a ) Cell viability of HCC cells transfected with siNUPR1 (siNUPR1 #1) and siNC was assessed by MTS assay after treatment with the indicated concentrations of sorafenib for 48 h. Data are expressed as the percentage of control cells and are the means±S.D. of three separate experiments, each performed in triplicate. * P <0.05, ** P <0.01. ( b ) Representative images of clonogenic assay of HCC cells transfected with siNUPR1 (siNUPR1 #1) and siNC. The experiment continued for 14 days. Surviving colonies were stained and counted. Data are expressed as the percentage of colonies and are the means±S.D. of three separate experiments, each performed in duplicate. ( c ) Representative images of wound-healing assay after NUPR1 siRNA-mediated gene silencing (shNUPR1 #1) in PLC/PRF/5. The experiment was conducted for 24 h. Data are reported as the percentage of cell migration and represent the average±S.D. of three experiments, each performed in duplicate. * P <0.05. ( d ) Representative images of transwell migration assay of Hep3B shNUPR1 (shNUPR1 #1) or Hep3B pSilencer cells. Data are reported as the percentage of migrated cells compared with control (siNC) and are the means±S.D. of three separate experiments, each performed in duplicate (* P <0.05). ( e ) Matrigel invasion assay in Hep3B shNUPR1 cells (shNUPR #1) compared with pSilencer as control. Data are reported as the percentage of invaded cells compared with control (pSilencer) and are the means±S.D. of three separate experiments, each performed in duplicate. * P <0.05

Article Snippet: A total of 10 × 10 6 Hep3B cells stably expressing shRNA against NUPR1 or control shRNA (pSilencer) were implanted subcutaneously in female nude athymic mice (Fox1 nu/nu) (6 weeks old, male; n =12) obtained from Envigo (Udine, Italy).

Techniques: Migration, Transfection, MTS Assay, Control, Clonogenic Assay, Staining, Wound Healing Assay, Transwell Migration Assay, Invasion Assay

Journal: Cell Death & Disease

Article Title: NUPR1, a new target in liver cancer: implication in controlling cell growth, migration, invasion and sorafenib resistance

doi: 10.1038/cddis.2016.175

Figure Lengend Snippet: NUPR1 regulates expression of RELB and IER3 genes, and RELB and IER3 regulate cell viability and the colony-formation capacity of HCC cells. ( a ) Gene expression analysis by qPCR in HCC cells after NUPR1 gene silencing (siNUPR1 #1). Data are reported as the percentage of gene expression inhibition of each gene and are the means±S.D. of three separate experiments, each performed in triplicate. ( b ) Gene expression analysis, by qPCR, in Hep3B shNUPR1 (shNUPR1 #1) cells compared with pSilencer, as control. Data are expressed as reported in panel ( a ). ( c ) Western blotting analysis of P -ERK1/2 (Thr202/Tyr204) and total ERK1/2 in Hep3B shNUPR1 (shNUPR #1) and in control cells (pSilencer). ( d ) Gene expression analyses by qPCR after RELB and IER3 gene silencing in Hep3B cells. Data are expressed as reported in panel ( a ). ( e ) Cell viability of Hep3B cells transfected with siRELB, siIER3 and siNC was assessed by MTS assay after treatment with the indicated sorafenib concentrations for 48 h. Data are expressed as reported in . * P <0.05. ( f ) Representative images of the clonogenic assay of Hep3B cells transfected with siRELB, siIER3 and siNC. The experiment continued for 14 days. Surviving colonies were stained and counted. Data are expressed as reported in . * P <0.05; ns=non-significant

Article Snippet: A total of 10 × 10 6 Hep3B cells stably expressing shRNA against NUPR1 or control shRNA (pSilencer) were implanted subcutaneously in female nude athymic mice (Fox1 nu/nu) (6 weeks old, male; n =12) obtained from Envigo (Udine, Italy).

Techniques: Expressing, Gene Expression, Inhibition, Control, Western Blot, Transfection, MTS Assay, Clonogenic Assay, Staining

Journal: Cell Death & Disease

Article Title: NUPR1, a new target in liver cancer: implication in controlling cell growth, migration, invasion and sorafenib resistance

doi: 10.1038/cddis.2016.175

Figure Lengend Snippet: NUPR1 knockdown inhibited tumor growth of Hep3B cells in nude mice. ( a ) Microphotography of 8 out of the 12 mice inoculated with stable Hep3B cells harboring NUPR1 shRNA (shNUPR1) (right flank) or non-specific shRNA (pSilencer) (left flank). ( b ) Microphotographs of tumors collected after 4 weeks of injection with Hpe3B pSilencer cells. ( c ) Tumor growth of Hep3B cells harboring pSilencer or shNUPR1. * P <0.05; ** P <0.01

Article Snippet: A total of 10 × 10 6 Hep3B cells stably expressing shRNA against NUPR1 or control shRNA (pSilencer) were implanted subcutaneously in female nude athymic mice (Fox1 nu/nu) (6 weeks old, male; n =12) obtained from Envigo (Udine, Italy).

Techniques: Knockdown, shRNA, Injection

Journal: Cell Death & Disease

Article Title: NUPR1, a new target in liver cancer: implication in controlling cell growth, migration, invasion and sorafenib resistance

doi: 10.1038/cddis.2016.175

Figure Lengend Snippet: Functional analysis for the data set of differentially expressed genes and network analysis of dynamic gene expression obtained following shRNA-mediated NUPR 1 knockdown in Hep3B cells. ( a and b ) IPA functional pathway analyses of genes differentially expressed (≥ 3-fold) in Hep3B cells upon NUPR1 suppression. Top functions that meet a P -value cutoff of 0.05 are displayed. The orange line represents the cutoff value for significance. ( a ) Genes that were upregulated and ( b ) genes downregulated. ( c ) The five top-scoring networks were merged and are displayed graphically as nodes (genes/gene products) and edges (the biological relationships between the nodes). Intensity of the node color indicates the degree of up regulation (red) or downregulation (green). Nodes are displayed using various shapes that represent the functional class of the gene product (rhomboid=transporter; square=cytokine; diamond=enzyme; vertical oval=transmembrane receptor; horizontal oval=transcription factor; rectangle=nuclear receptor; hexagon=translation factor; circle=other). Edges are displayed with various labels that describe the nature of the relationship between the nodes:→acts on; -— binding only. The length of an edge reflects the evidence supporting the specific node-to-node relationship, as edges supported by articles from the literature are shorter. Dotted edges represent indirect interaction

Article Snippet: A total of 10 × 10 6 Hep3B cells stably expressing shRNA against NUPR1 or control shRNA (pSilencer) were implanted subcutaneously in female nude athymic mice (Fox1 nu/nu) (6 weeks old, male; n =12) obtained from Envigo (Udine, Italy).

Techniques: Functional Assay, Gene Expression, shRNA, Knockdown, Binding Assay

Journal: Cell Death & Disease

Article Title: NUPR1, a new target in liver cancer: implication in controlling cell growth, migration, invasion and sorafenib resistance

doi: 10.1038/cddis.2016.175

Figure Lengend Snippet: RUNX2 regulates cell viability, growth, migration and the expression of NUPR1 , RELB and IER3 genes in Hep3B cells and is expressed in human HCC samples. ( a ) Gene expression analysis by qPCR after NUPR1 gene silencing (shNUPR1 #1) in Hep3B cells. ( b ) Colony assay after RUNX2 siRNA-mediated gene knockdown. Data are expressed as reported in . ( c ) Representative images of transwell migration assay after RUNX2 gene silencing in Hep3B cells. Data are expressed as reported in .** P <0.01. ( d ) Cell viability of Hep3B cells transfected with siRUNX2 (siRUNX2 #1) and siNC was assessed by MTS assay after treatment with the indicated concentrations of sorafenib for 48 h. Data are expressed as reported in . ( e ) Gene expression analysis after RUNX2 gene silencing (siRUNX2 #1) in Hep3B cells performed by qPCR. Data are expressed as reported in . ( f ) RUNX2 gene expression analysis in 12 HCC tissues performed by qPCR. Data are indicated as reported in . ( g ) RUNX2 protein expression levels were examined by immunohistochemistry in the NL ( A ) and HCC tissues ( B ). Magnification= × 20, insert magnification= × 40. Scale bar=100 μ m

Article Snippet: A total of 10 × 10 6 Hep3B cells stably expressing shRNA against NUPR1 or control shRNA (pSilencer) were implanted subcutaneously in female nude athymic mice (Fox1 nu/nu) (6 weeks old, male; n =12) obtained from Envigo (Udine, Italy).

Techniques: Migration, Expressing, Gene Expression, Colony Assay, Knockdown, Transwell Migration Assay, Transfection, MTS Assay, Immunohistochemistry

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: Discover Oncology

Article Title: BIRC5 knockdown ameliorates hepatocellular carcinoma progression via regulating PPARγ pathway and cuproptosis

doi: 10.1007/s12672-024-01592-y

Figure Lengend Snippet: BIRC5 knockdown suppresses proliferation, invasion, migration and promotes apoptosis of HCC cells. A QRT-PCR assay was used to verify the knockdown efficiency of BIRC5 expression in both SK-Hep-1 and Hep-3B cells. B Cell viability of SK-Hep-1 and Hep-3B cells with or without BIRC5 knockdown was detected by CCK-8 assay. C Cell migration in si-NC and si-BIRC5 groups was measured by scratch assay. Scale bar: 100 μm. D The transwell assay was utilized to detect cell invasion in si-NC and si-BIRC5 groups. Scale bar: 100 μm. E Flow cytometry was performed to detect cell apoptosis of BIRC5 silencing SK-Hep-1 and Hep-3B cells. ** p < 0.01, *** p < 0.001 vs. si-NC

Article Snippet: The normal human liver cell line (L-O2) and human HCC lines (Hep-3B and SK-Hep-1) were obtained from iCell (Shanghai, China), then were cultured in DMEM medium (Hyclone, Logan, UT, USA) containing 10% heat-inactivated FBS, 100 U/mL penicillin and 100 μg/mL streptomycin (Hyclone, Logan, UT, USA) at 37 °C in a humidified atmosphere of 5% CO 2 concentration conditions.

Techniques: Knockdown, Migration, Quantitative RT-PCR, Expressing, CCK-8 Assay, Wound Healing Assay, Transwell Assay, Flow Cytometry

Journal: Discover Oncology

Article Title: BIRC5 knockdown ameliorates hepatocellular carcinoma progression via regulating PPARγ pathway and cuproptosis

doi: 10.1007/s12672-024-01592-y

Figure Lengend Snippet: Knockdown of BIRC5 affects the cuproptosis of HCC cells. A The supernatant contents of pyruvic acid and α-ketoglutaric acid in BIRC5-silenced SK-Hep-1 and Hep-3B cells were detected using assay kits. B The cellular copper ion (Cu 2+ ) level in SK-Hep-1 and Hep-3B cells characterized by knockdown of BIRC5 was measured using inductively coupled plasma-Optical Emission Spectrometer (ICP-OES). C Expressions of FDX1 and DLAT in BIRC5-silenced SK-Hep-1 and Hep-3B cells were detected by Western blot. ** p < 0.01, *** p < 0.001 vs. si-NC

Article Snippet: The normal human liver cell line (L-O2) and human HCC lines (Hep-3B and SK-Hep-1) were obtained from iCell (Shanghai, China), then were cultured in DMEM medium (Hyclone, Logan, UT, USA) containing 10% heat-inactivated FBS, 100 U/mL penicillin and 100 μg/mL streptomycin (Hyclone, Logan, UT, USA) at 37 °C in a humidified atmosphere of 5% CO 2 concentration conditions.

Techniques: Knockdown, Clinical Proteomics, Western Blot

Journal: Discover Oncology

Article Title: BIRC5 knockdown ameliorates hepatocellular carcinoma progression via regulating PPARγ pathway and cuproptosis

doi: 10.1007/s12672-024-01592-y

Figure Lengend Snippet: BIRC5 regulates the PPAR pathway in HCC cells. Protein levels of PPAR-γ and FABPP5 were verified by western blot in SK-Hep-1 and Hep-3B cells between si-NC, si-BIRC5 group. * p < 0.05, *** p < 0.001 vs. si-NC

Article Snippet: The normal human liver cell line (L-O2) and human HCC lines (Hep-3B and SK-Hep-1) were obtained from iCell (Shanghai, China), then were cultured in DMEM medium (Hyclone, Logan, UT, USA) containing 10% heat-inactivated FBS, 100 U/mL penicillin and 100 μg/mL streptomycin (Hyclone, Logan, UT, USA) at 37 °C in a humidified atmosphere of 5% CO 2 concentration conditions.

Techniques: Western Blot

Journal: Biomimetics

Article Title: Anticancer Activity of Thiophene Carboxamide Derivatives as CA-4 Biomimetics: Synthesis, Biological Potency, 3D Spheroid Model, and Molecular Dynamics Simulation

doi: 10.3390/biomimetics7040247

Figure Lengend Snippet: IC 50 (µM) of phenyl-thiophene-carboxamide compounds ( 2a – 2e ) on several cell lines.

Article Snippet: Our results showed that the PSA of the most synthesized structures was biomimetic to CA-4, and similar chemical and biological properties were observed against Hep3B cancer cell line.

Techniques:

Journal: Biomimetics

Article Title: Anticancer Activity of Thiophene Carboxamide Derivatives as CA-4 Biomimetics: Synthesis, Biological Potency, 3D Spheroid Model, and Molecular Dynamics Simulation

doi: 10.3390/biomimetics7040247

Figure Lengend Snippet: RHA-3 ( 2b ) and RHA-6 ( 2e ) perturb 3D hepatocellular spheroids’ formation. Images of cluster/s formed by Hep3B hepatocellular carcinoma in presence of 2b (17 µg/mL) or 2e (17 µg/mL) after 24 h of treatment compared to controls ( A ). Cluster percentage of occupied area relative to the negative control ( B ), cluster circularity ( C ), and cluster count ( D ). The non-treated cells are referred to as a negative control. At 100 μg/mL, DOX was utilized as a positive control. The scale bar represents a distance of 10 μm. Circularity scale: a value of 1 represents a perfect circle (ns: p > 0.05, **: p ≤ 0.01, ***: p ≤ 0.001, and ****: p ≤ 0.0001).

Article Snippet: Our results showed that the PSA of the most synthesized structures was biomimetic to CA-4, and similar chemical and biological properties were observed against Hep3B cancer cell line.

Techniques: Negative Control, Positive Control