Journal: Endocrinology

Article Title: Periovulatory expression of hydrogen peroxide-induced sulfiredoxin and peroxiredoxin 2 in the rat ovary: gonadotropin regulation and potential modification.

doi: 10.1210/en.2012-1414

Figure Lengend Snippet: FIG. 6. Regulation of ovarian PRDX2 expression by gonadotropins. A, Aliquots of total RNA (20 g) isolated from ovaries at the indicated time intervals after eCG/hCG stimulation were assayed for PRDX2 mRNA levels by Northern blotting using a rat PRDX2 cDNA probe. The expression of 28S ribosomal RNA was used as an internal standard. The PRDX2 transcript was quantified using a phosphorimager and normalized for 28S RNA levels from three independently performed experiments (mean SEM). B, In situ localization of ovarian PRDX2 mRNA. Ovarian sections were hybridized with a 35S-labeled PRDX2 cRNA probe. Photomicrographs were taken under bright-field (a, c, and e) and corresponding dark-field (b, d, and f) illumination. Arrowhead, Oocyte; arrow, theca cell; asterisk, growing follicle; P, preovulatory follicle; CL, corpus luteum, 20. C, Western blotting analysis of PRDX2 protein in the ovary. Ovarian lysates were separated on a 12% SDS-PAGE gel in the presence (reducing) or absence (nonreducing) of 10% 2-mercaptoethanol. The quantified PRDX2 protein levels are expressed as a mean SEM of two or three independently performed experiments.

Article Snippet: Blots were incubated with primary antibodies against Srx, PRDX2, -PRDX-SO2/3, p-Akt (Cell Signaling Technology, Beverly, MA), Akt (Cell Signaling Technology), pPTEN (Cell Signaling Technology), or PTEN (Cell Signaling Technology).

Techniques: Expressing, Isolation, Northern Blot, In Situ, Labeling, Western Blot, SDS Page

Journal: Endocrinology

Article Title: Periovulatory expression of hydrogen peroxide-induced sulfiredoxin and peroxiredoxin 2 in the rat ovary: gonadotropin regulation and potential modification.

doi: 10.1210/en.2012-1414

Figure Lengend Snippet: FIG. 7. Modification of PRDX2 in preovulatory follicles after gonadotropin treatment. Preovulatory follicles were dissected from ovaries obtained at different times after eCG/hCG treatment. Cell lysates of preovulatory follicles were analyzed by SDS-PAGE (A) or two- dimensional PAGE (B) followed by immunoblot analysis with antibodies specific to Srx, sulfinic/sulfonic 2-Cys PRDXs (PRDX-SO2/3), PRDX2, and GAPDH. Lysates of HeLa cells exposed to 1 mM H2O2 for 10 min were used as a positive control for the expression of hyperoxidized 2-Cys PRDXs. The region on the two-dimensional immunoblots corresponds to a molecular mass of 22–28 kDa (vertical) and isoelectric points of 5.2–5.6 (horizontal). The band intensities of each sample were measured and the ratio of hyperoxidized (Ox) to reduced (Re) PRDX2 was used to plot the graph (B, lower panel). Each point on the graph represents mean SEM of three independently performed experiments. *, P 0.05 vs. 0 h.

Article Snippet: Blots were incubated with primary antibodies against Srx, PRDX2, -PRDX-SO2/3, p-Akt (Cell Signaling Technology, Beverly, MA), Akt (Cell Signaling Technology), pPTEN (Cell Signaling Technology), or PTEN (Cell Signaling Technology).

Techniques: Modification, SDS Page, Western Blot, Positive Control, Expressing

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Compound kushen injection suppresses human acute myeloid leukaemia by regulating the Prdxs/ROS/Trx1 signalling pathway

doi: 10.1186/s13046-018-0948-3

Figure Lengend Snippet: Prdx3 was identified by LC-MS/MS for CKI against AML, and CKI regulated Prdxs/Trx1 in AML. a Data analysis. b GO annotation with 288 differential expressed proteins. c Heat map showed the changes in 11 functions in 54 differential expressed proteins after CKI treatment. d The interaction between PRDX3 and other selected proteins by STRING. e Validation of Prdx3, Prdx2, and Trx1 protein expression in healthy and AML cells by Western blot. f The protein expression changes to Prdx3, Prdx2, and Trx1 after treatment of AML cells with CKI and their detection by Western blot. g The expression and interaction of Prdx3, Prdx2, and Trx1 after CKI treatment in AML cells and their observation using laser confocal microscopy

Article Snippet: The blot was probed with rabbit polyclonal antibodies against Prdx3 (1:500, USCNK) and Trx1 (1 μg/ml, Abcam), and mouse monoclonal antibodies against Prdx2 (1:2000, Protein-tech), as primary antibodies, overnight at 4 °C.

Techniques: Liquid Chromatography with Mass Spectroscopy, Biomarker Discovery, Expressing, Western Blot, Confocal Microscopy

Journal: Journal of Experimental & Clinical Cancer Research : CR

Article Title: Compound kushen injection suppresses human acute myeloid leukaemia by regulating the Prdxs/ROS/Trx1 signalling pathway

doi: 10.1186/s13046-018-0948-3

Figure Lengend Snippet: Therapeutic efficacy of CKI on the patient-derived xenograft model of acute myeloid leukaemia. a The schematic diagram for the AML PDX model. b Analysis of bone marrow smear. At day 20, the bone marrow smear was stained and leukaemia cells were observed. Magnification fold: × 100 under an oil immersion lens. c The changes in body weight. d The survival analysis. e The remission state of bone marrow in the CKI group. f The immunophenotype of leukaemia cells in peripheral blood by flow cytometry analysis. g Statistics for the ratio of leukaemia cells. h The tissue weight index. i Testing the T-AOC vitality, GSH content, H 2 O 2 concentration and CAT activity in plasma from B-NSG mice. j Detection of Prdx3, Prdx2, and Trx1 expression by IHC in the PDX model. k The model for the anti-leukaemic effects of CKI

Article Snippet: The blot was probed with rabbit polyclonal antibodies against Prdx3 (1:500, USCNK) and Trx1 (1 μg/ml, Abcam), and mouse monoclonal antibodies against Prdx2 (1:2000, Protein-tech), as primary antibodies, overnight at 4 °C.

Techniques: Drug discovery, Derivative Assay, Staining, Flow Cytometry, Concentration Assay, Activity Assay, Clinical Proteomics, Expressing

Journal: Oncotarget

Article Title: Dimeric peroxiredoxins are druggable targets in human Burkitt lymphoma

doi:

Figure Lengend Snippet: A. Analysis of the expression of PRDX1-4, TRX1, TRXR1, in the data set E-GEOD-12453 deposited in the ArrayExpress database. The expression is analyzed in 5 sub-populations of normal B cells (left-hand) and in malignant B-cell derived DLBCL and BL cells (right-hand). B. IHC staining of lymph nodes (LNs) with reactive follicular hyperplasia (left panel) and representative BL cells for PRDX1 and PRDX2 (EnVision stain, 20x). In the right panel, insertions in the upper right corners have higher magnification (60x) of BL tissue stained with anti-PRDX1 and PRDX2, respectively. C. qRT-PCR analysis of the PRDX1 and PRDX2 gene expression in B cell-derived tumor cell lines and four CD19+ control B cells. qPCR data were normalized to housekeeping genes, RPL29 and β-2-microglobulin. The fold change is displayed on a log2 scale. Mean values from two independent experiments are shown, the bars indicate the standard error of the mean (SEM). D. Representative Western blot for the PRDX1 and PRDX2 protein levels in B cell-derived tumor cell lines and CD19+ control B cells obtained from 4 healthy donors.

Article Snippet: Immunohistochemistry was performed using the following antibodies: PRDX1 (Sigma-Aldrich, dilution 1:150, pH = 9.0) and PRDX2 (GeneTex, clone EPR5154, dilution 1:200, pH = 9.0).

Techniques: Expressing, Derivative Assay, Immunohistochemistry, Staining, Quantitative RT-PCR, Gene Expression, Control, Western Blot

Journal: Oncotarget

Article Title: Dimeric peroxiredoxins are druggable targets in human Burkitt lymphoma

doi:

Figure Lengend Snippet: A. Raji-sub cells were treated with lentiviruses encoding two different PRDX1-specific shRNAs or control, non-targeting shRNA. After puromycin selection, the number of viable cells was evaluated in a hemocytometer for three consecutive days. At days 1 (d1) and 3 (d3) cells were collected, and the levels of PRDX1, PRDX2 were evaluated by immunoblotting. B. The level of cleaved PARP (clPARP) was assessed by immunoblotting. C. Raji cells were subjected to transduction with lentiviruses carrying PRDX2-specific shRNA and hygromycin resistance gene. After 6 days of antibiotic selection, the cells were infected with lentiviruses encoding PRDX1-targeting shRNA2 and puromycin resistance genes. Following 3 days of puromycin selection, the number of viable cells was assessed with flow cytometry, by counting the PI-negative cells. Three days after puromycin selection, cells were collected and the levels of PRDX1 and PRDX2 were analyzed by immunoblotting. D. Levels of clPARP, P-ERK1/2, p21 were assessed at day 3 post selection by immunoblotting. DNA synthesis was assessed in Raji cells expressing PRDX1-specific shRNA2 (shPRDX1) and control cells expressing non-targeting shRNA (control shRNA) with the Click-iT EdU (5-ethynyl-2′-deoxyuridine) incorporation assay. Data show the % of EdU-positive cells, mean values from two independent repeats ± SD, * P < 0.05. The cell cycle distributions in Raji cells expressing PRDX1-specific shRNA2 (shPRDX1) and control cells expressing non-targeting shRNA (control shRNA) were evaluated with a propidium iodide flow cytometry-based assay. The error bars indicate the SD ( n = 2), * P < 0.05. E. Namalwa cells were subjected to sequential lentiviral transductions to downregulate PRDX1 and PRDX2, as described in C. and the number of viable cells was assessed in a hemocytometer for three consecutive days. The degree of PRDX1 and PRDX2 knockdown was assessed by immunoblotting in cells collected 3 days after puromycin selection.

Article Snippet: Immunohistochemistry was performed using the following antibodies: PRDX1 (Sigma-Aldrich, dilution 1:150, pH = 9.0) and PRDX2 (GeneTex, clone EPR5154, dilution 1:200, pH = 9.0).

Techniques: Control, shRNA, Selection, Western Blot, Transduction, Infection, Flow Cytometry, DNA Synthesis, Expressing, Knockdown

Journal: Oncotarget

Article Title: Dimeric peroxiredoxins are druggable targets in human Burkitt lymphoma

doi:

Figure Lengend Snippet: Cells were cultured with SK053, biotinylated SK053 (SK-bio) or its inactive counterpart (SK-in), harvested and analyzed by immunoblotting using specific antibodies, as indicated on the left side of each blot. Bands labeled M migrated at ∼ 22 kDa, a mass corresponding to the molecular weight of the peroxiredoxin monomer, and bands labeled D migrated at ∼ 44 kDa, a mass corresponding to the molecular weight of the peroxiredoxin dimer. A. The concentration of SK-bio and SK-in was 55 μM (=LC 80 ). B. Cells were cultured with the indicated concentrations of SK053 for 24 h. C. 10 μM of SK053 was used, which is close to the LC 50 .

Article Snippet: Immunohistochemistry was performed using the following antibodies: PRDX1 (Sigma-Aldrich, dilution 1:150, pH = 9.0) and PRDX2 (GeneTex, clone EPR5154, dilution 1:200, pH = 9.0).

Techniques: Cell Culture, Western Blot, Labeling, Molecular Weight, Concentration Assay