Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Response of esophageal cancer cells to epigenetic inhibitors is mediated via altered thioredoxin activity.

doi: 10.1038/labinvest.2015.148

Figure Lengend Snippet: Figure 1 Esophageal cancer cells show loss of nuclear Trx and higher TXNRD expression. (a) The panels show representative serial sections of stained case-matched normal esophageal epithelium and ESCC or EAC tissue specimens for Trx, TXNIP and TXNRD. Refer to Table 1 for results of all cases. (b) Please note loss of nuclear Trx in transition zones of esophageal cancers. Bars represent 200 μm.

Article Snippet: Immunoblotting was performed as described before.22 Following antibodies and dilutions were used: Trx: 1:5000 from abcam/ab133524, TXNIP: 1:1000 from Novus Biologicals/clone JY2 and TXNRD: 1:10 000 from Abcam/ ab124954.

Techniques: Expressing, Staining

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Response of esophageal cancer cells to epigenetic inhibitors is mediated via altered thioredoxin activity.

doi: 10.1038/labinvest.2015.148

Figure Lengend Snippet: Figure 2 Characterization of esophageal cell lines for Trx, TXNIP and TXNRD. (a) Subcellular localization of Trx and its associated proteins TXNIP and TXNRD was analyzed by immunofluorescence stainings. Shown are 3D image stacks and, for Trx, one central image through the nucleus. Note cytoplasmic and nuclear Trx expression in Het-1A cells, but loss of nuclear Trx in both cancer cell lines OE21 and OE33 (see inserts of nuclear sections). Bar represents 20 μm. (b and c) Protein levels of Trx, TXNIP and TNXRD levels were compared by immunoblotting with subsequent (b) quantification by ImageJ. (c) Shown is one representative immunoblot of three independent experiments. Note slight Trx increase and strong TXNIP decrease in both cancer cell lines. (d) Quantification of baseline Trx activity with higher Trx activity in OE21 and, to a lesser extent, also in OE33 compared with non- neoplastic Het-1A cells. Shown is the mean ± s.e.m. for three independent experiments. Significance levels are represented as *0.05–0.01 and *** ≤0.001.

Article Snippet: Immunoblotting was performed as described before.22 Following antibodies and dilutions were used: Trx: 1:5000 from abcam/ab133524, TXNIP: 1:1000 from Novus Biologicals/clone JY2 and TXNRD: 1:10 000 from Abcam/ ab124954.

Techniques: Immunofluorescence, Expressing, Western Blot, Activity Assay

Journal: Laboratory investigation; a journal of technical methods and pathology

Article Title: Response of esophageal cancer cells to epigenetic inhibitors is mediated via altered thioredoxin activity.

doi: 10.1038/labinvest.2015.148

Figure Lengend Snippet: Figure 4 TXNIP is increased in OE21 cells after MS-275/AZA treatment. For analysis of Trx, TXNIP and TXNRD levels after MS-275 and/or AZA treatment, proteins were isolated 6 and 24 h after addition of inhibitors and evaluated by (a) immunoblotting with (b) quantification by densitometry. For Het-1A and OE33 cells, no differences for Trx, TXNIP and TXNRD were detectable. However, OE21 cells displayed an increase of TXNIP, especially for combination of MS-275 and AZA after 24 h. Shown is one representative immunoblot for the target and control protein of each the same protein lysate. The bar diagrams represent the mean ± s.e.m. for three independent experiments.

Article Snippet: Immunoblotting was performed as described before.22 Following antibodies and dilutions were used: Trx: 1:5000 from abcam/ab133524, TXNIP: 1:1000 from Novus Biologicals/clone JY2 and TXNRD: 1:10 000 from Abcam/ ab124954.

Techniques: Isolation, Western Blot, Control

Journal: Experimental and therapeutic medicine

Article Title: 3-Bromopyruvic acid regulates glucose metabolism by targeting the c-Myc/TXNIP axis and induces mitochondria-mediated apoptosis in TNBC cells.

doi: 10.3892/etm.2022.11447

Figure Lengend Snippet: Figure 4. 3‑BrPA regulates expression of c‑Myc, TXNIP and HK2. (A) Expression levels of c‑Myc, TXNIP, HK2 and GLUT1 were detected through western blot analysis. Grayscale values of the bands were determined using software. (B) 3‑BrPA downregulated c‑Myc and HK2 protein expression, whereas it upregulated TXNIP protein expression in TNBC cells (*P<0.05 vs. control group, the same cell line with 0 µM 3‑BrPA). (C) However, in non‑TNBC cells, the expression levels of c‑Myc, TXNIP, HK2 and GLUT1 were not significantly affected. The expression levels of GLUT1 were not significantly affected in both TNBC and non‑TNBC cells. TXNIP, thioredoxin‑interacting protein; HK2, hexokinase 2; GLUT1, glucose transporter 1; 3‑BrPA, 3‑bromopyruvic acid; TNBC, triple‑negative breast cancer.

Article Snippet: Then the membrane was cultured in QuickBlock Blocking Buffer for Western Blot (Beyotime Institute of Biotechnology) at room temperature for 1 h. The membrane was incubated with antibodies against GLUT1 (1:1,000; cat. no. TA312796; OriGene Technologies, Inc.), c‐Myc (1:1,000; cat. no. TA150121; OriGene Technologies, Inc.), TXNIP (1:1,000; cat. no. TA349090; OriGene Technologies, Inc.), HK2 (1:1,000; cat. no. TA500856; OriGene Technologies, Inc.), Bcl‐2 (1:2,000; cat. no. TA806591; OriGene Technologies, Inc.), Bax (1:2,000; cat. no. TA810334; OriGene Technologies, Inc.), cytochrome c (Cyt‐C; 1:5,000; cat. no. ab13575; Abcam), Caspase‐3 (1:1,000; cat. no. TA301776; OriGene Technologies, Inc.) and β‐actin (1:1,000; cat. no. TA811000; OriGene Technologies, Inc.) at room temperature over‐ night.

Techniques: Expressing, Western Blot, Software, Control

Journal: PLoS ONE

Article Title: Evaluating the antidiabetic effects of R-verapamil in type 1 and type 2 diabetes mellitus mouse models

doi: 10.1371/journal.pone.0255405

Figure Lengend Snippet: (A) STZ-induced mice administered 100 mg/kg/day racemic or R-Vera exhibited significantly reduced serum IL-6 expression compared to vehicle mice whereas those administered 50 mg/kg/day R-Vera did not. ** P < 0.01 indicates significant difference from vehicle using one-way ANOVA. N = 8 in each group. (B) STZ-induced mice showed highly upregulated islet tissue TXNIP compared to normal C57BL/6J mice. TXNIP was downregulated after racemic or R-Vera administration compared to vehicle. # P < 0.05 indicated significant difference between normal and vehicle; * P < 0.05 indicated significant difference to vehicle using one-way ANOVA. N = 8 in each group. (C) β-cell apoptosis assessed using TUNEL staining. Deep staining spots in images are apoptotic β-cells (200×). (D) TUNEL-positive cell counts show that STZ-induced mice administered 100 mg/kg/day R-Vera had significantly reduce β-cell apoptosis. * P < 0.05 indicated a significant difference between vehicle and RV-100 using one-way ANOVA. The sample numbers of 5, 7, 5, and 9 indicate the section numbers from the Vehicle, V-100, RV-100, and RV-50 groups, respectively (rather than the number of mice). Each data point represents the percentage of TUNEL–positive cells per slide in each group.

Article Snippet: The reagents were mouse anti-TXNIP antibody (1:500; NBP1-54578; Novus Biologicals, Centennial CO, USA), mouse anti-β-actin (1:2,000; No. 3700; Cell Signaling Technology, Danvers, MA, USA), and a Wes TM Simple Western system (ProteinSimple, San Jose, CA, USA).

Techniques: Expressing, TUNEL Assay, Staining