Journal: Cancer Research

Article Title: MEN1 Promotes Ferroptosis by Disrupting CD44 Alternative Splicing to Suppress Lung Cancer

doi: 10.1158/0008-5472.CAN-25-0021

Figure Lengend Snippet: MEN1 -modulated ferroptosis is required for suppressing lung tumor growth and metastasis. A–D, Cell viability and representative invasion and migration images of WT and KO NCI-H460 cells; invaded cells and migration rate were quantified at the indicated time points. E, Schematic of experimental lung metastasis model in M-NSG mice. F, Representative IHC (Ki67 and HMGB1) and H&E staining images of lung tissue sections from M-NSG mice xenografted with WT/KO NCI-H460 cells. Scale bars, 1 mm or 50 μm. G and H, Tumor burden ( G ) and quantification of Ki67/HMGB1 IHC staining in lung tissues ( H ) from xenografted M-NSG mice. I, Schematic of the primary mouse lung cancer study ( n = 7, 9, and 8 for WT, KS, and KMS cohorts). J–M, Representative lung pictures ( J ), H&E staining and Ki67/HMGB1 IHC images of lung tissues ( K ), tumor burden and tumor number ( L ), and quantification of Ki67/HMGB1 IHC staining ( M ). Scale bars, 1 mm or 50 μm. Black arrows, tumor nodules. N, Kaplan–Meier survival analysis of WT, KS, and KMS mice. O, Lipid peroxidation (LPO) levels in lungs from WT, KS, and KMS mice. a.u, arbitrary unit. P and Q, Representative RNA FISH images ( P ) and quantification of GPX4 probe numbers in WT lung tissues and KS/KMS lung tumor tissues ( Q ). Scale bars, 20 or 50 μm. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, not significant.

Article Snippet: The paraffin-embedded tissue sections after deparaffinization and rehydration were subjected to antigen retrieval in 0.1 mol/L citrate buffer (pH 6.0) in a steam pot for 15 minutes, blocked with 5% BSA for 1 hour, and incubated with primary antibodies including menin (Bethyl Laboratories, #A300-105A, RRID: AB_2143306), Ki67 (Cell Signaling Technology, #9129, RRID: AB_2687446), HMGB1 (Proteintech, #10829-1-AP, RRID: AB_2232989), and SFTPC (Abclonal, #A1835, RRID: AB_2763872).

Techniques: Migration, Staining, Immunohistochemistry

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a Prediction of candidate HMGB1 crosstalk RNAs using the ceRDB database ( https://www.oncomir.umn.edu/cefinder ). The top 12 candidate RNAs and scores were shown. b The RNA expression levels of HMGB1 and the 12 candidate RNAs were analyzed in early stage HCC (BCLC stages 0 and A HCC) tissues by qRT-PCR. HBV + HCC (above) n = 26; HBV − HCC (below), n = 9. The results are means ± SD. Names marked red and black represent significantly ( p < 0.05) and nonsignificant genes, respectively. c Pearson correlation coefficient analysis between expression levels of HMGB1 and the 12 candidate RNAs, respectively, in early stage HCC tumor tissues

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: RNA Expression, Quantitative RT-PCR, Expressing

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a The mRNA expression of HMGB1 and RICTOR in the 11 indicated cell lines as determined by qRT-PCR. * p < 0.05. b Pearson correlation coefficient analysis between the expression level of HMGB1 and RICTOR in the cell lines from ( a ). c Immunohistochemical staining of HMGB1 and RICTOR in liver tissues in the DEN + CCl4-induced liver cancer mouse model. d , e The expression levels of RICTOR and HMGB1 mRNA (left) and protein (right) after the interference of HMGB1 or RICTOR in the HCCLM3 ( d ) and PLC/PRF/5 ( e ) cell lines (abbreviated as “KD-HMGB1 or KD-RICTOR”, respectively). f The expression levels of RICTOR and HMGB1 mRNA (left) and protein (right) after overexpression of HMGB1 or RICTOR 3′UTR in the QSG-7701 cell line (abbreviated as “OE-HMGB1 or OE-RICTOR”, respectively). The mRNA levels (left) were determined by qRT-PCR and protein level (right) was determined by Western blot. * p < 0.05, *** p < 0.001

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: Expressing, Quantitative RT-PCR, Immunohistochemical staining, Staining, Over Expression, Western Blot

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a Luciferase assays with reporter constructs containing the wild-type or mutant HMGB1/RICTOR 3′UTR downstream of the luciferase gene were performed after co-transfection with miR-200a/200b/429 mimics in HEK293T cells. * p < 0.05, ** p < 0.01, *** p < 0.001. b The mRNA levels of HMGB1 and RICTOR were determined by qRT-PCR 48 h after transfection of 100 nM miR-200a/200b/429 mimics in indicated cells. ** p < 0.01, *** p < 0.001. c RNA FISH assays detecting the cellular localization of HMGB1/RICTOR and miR-429 in indicated cells

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: Luciferase, Construct, Mutagenesis, Cotransfection, Quantitative RT-PCR, Transfection

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a Determination of spheroid formation after 3000 cells were seeded in low-adhesion plates for 10 days (left). The number of tumor spheroids was quantitated (right). *** p < 0.001. b Cell proliferation was evaluated using CCK8 assays. The results are means ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001. c Male nude mice ( n = 5) were subcutaneously injected with 1 × 10 4 HMGB1/RICTOR 3′UTR over-expressed liver cells (abbreviated as “OE-HMGB1 or OE-RICTOR”, respectively) or HMGB1/RICTOR mRNA interference HCC cells (abbreviated as “KD-HMGB1 or KD-RICTOR”, respectively). Tumorigenesis was assessed after 48 days as shown

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: Injection

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a PMBC killing assay scheme. HMGB1/RICTOR mRNA interference HCC cells (abbreviated as “KD-HMGB1 or KD-RICTOR”, respectively) were cocultured with activated PBMCs and treated with or without Atezolizumab for 48 h before apoptosis detection. b Cell apoptosis in treated HCC cells was evaluated by TUNEL assay (above). The apoptotic cell ratios were shown (below). c Exosomes isolated from cell culture supernatants in treated HCC cells were determined by flow cytometry using PE-fluorescein-conjugated anti-PD-L1 antibody (red). Isotype matched antibody (PE-fluorescein-conjugated anti-IgG antibody) was used as gating controls (black). Percentages of PD-L1 + Exo resulted from the relative fluorescence values gated by isotype controls. Exo: exosomes. ** p < 0.01, *** p < 0.001

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: TUNEL Assay, Isolation, Cell Culture, Flow Cytometry, Fluorescence

Journal: Signal Transduction and Targeted Therapy

Article Title: An RNA–RNA crosstalk network involving HMGB1 and RICTOR facilitates hepatocellular carcinoma tumorigenesis by promoting glutamine metabolism and impedes immunotherapy by PD-L1+ exosomes activity

doi: 10.1038/s41392-021-00801-2

Figure Lengend Snippet: a A model of the RNA–RNA crosstalk network involving HMGB1 and RICTOR acting in the early stage HCC

Article Snippet: The frozen sections were examined based on in situ hybridization using digoxin-labeled HMGB1, RICTOR, and miR-429 probes and digoxin detection kit (Boster biological technology, Guangzhou, China) according to the suggested procedures.

Techniques: