Journal: Scientific Reports

Article Title: Tuning IL-2 signaling by ADP-ribosylation of CD25

doi: 10.1038/srep08959

Figure Lengend Snippet: Splenocytes from DEREG mice were incubated in the absence or presence of the indicated concentrations of NAD + before staining with fluorochrome-conjugated mAbs directed against CD4 and CD25 (mAb PC61 or mAb 7D4). Gating was performed on CD4 + cells. (a) Representative dot plots of cells incubated in the absence or presence of 12 μM NAD + . (b) Percentages of CD4 + GFP + cells staining with anti-CD25 mAbs plotted as a function of the concentration of added NAD + . Results are representative of two independent experiments.

Article Snippet: Alternatively, after erythrocyte lysis with Ack lysis buffer (Bio Whittaker) and depletion of B-cells with Dynabead-conjugated sheep anti-mouse IgG (Invitrogen), Tregs were positively selected by magnetic cell sorting using PE conjugated anti-CD25 (7D4) and magnetic bead-conjugated anti-PE antibodies according to the manufacturer's instructions (Miltenyi Biotec).

Techniques: Incubation, Staining, Concentration Assay

Journal: Arthritis Research & Therapy

Article Title: Therapeutic effect of a novel histone deacetylase 6 inhibitor, CKD-L, on collagen-induced arthritis in vivo and regulatory T cells in rheumatoid arthritis in vitro

doi: 10.1186/s13075-017-1357-2

Figure Lengend Snippet: Increased cytotoxic T-lymphocyte antigen-4 ( CTLA-4 ) expression by CKD-L in Treg cells from C57BL/6 mice ( n = 3). CD4 + CD25 – T cells were isolated from splenocytes of C57BL/6 mice. CD4 + CD25 – T cells were incubated with vehicle or HDAC6 inhibitors (1 to 10 μM) in the presence of antiCD3/CD28 beads and recombinant TGF-β2 in a 48-well plate for 6 days. CTLA-4 expression in CD4 + CD25 + Foxp3 + T cells was analyzed by mean fluorescence intensity ( MFI ; mean ± SEM) using flow cytometry. * p < 0.05, ** p < 0.001, vs vehicle

Article Snippet: Isolated CD4 + cells were incubated with microbead conjugated to a monoclonal antihuman CD25 antibody (MACS®; Miltenyi Biotec) for 15 min at 4 °C.

Techniques: Expressing, Isolation, Incubation, Recombinant, Fluorescence, Flow Cytometry

Journal: Cancer communications (London, England)

Article Title: IL-2Rα up-regulation is mediated by latent membrane protein 1 and promotes lymphomagenesis and chemotherapy resistance in natural killer/T-cell lymphoma.

doi: 10.1186/s40880-018-0334-8

Figure Lengend Snippet: Fig. 1 a–c Analysis of NK-92 and SNK-6 cell lines in terms of levels of a IL-2Rα mRNA by quantitative real-time PCR, b IL-2Rα protein by Western blot, and c soluble IL-2Rα protein in culture supernatant by ELISA. **P < 0.01 vs NK-92 cells. d, e Efficiency of NK-92 cell infection with d lentivirus encoding LMP1 or e negative control lentivirus at multiplicities of infection (MOIs) 100, 200, or 300. f–i Analysis of NK-92 cells (control) and NK-92 cells transduced with lentivirus encoding LMP1 (LMP1) or negative control lentivirus (NC) in terms of levels of f LMP1 and g IL-2Rα mRNA by quantitative real-time PCR, h LMP1 and IL-2Rα proteins by Western blot, and i soluble IL-2Rα protein in culture supernatant by ELISA. **P < 0.01 vs control or NC

Article Snippet: The membranes were incubated at room temperature for 1 h with one of the following primary antibodies: IL-2Rα, B-Raf, p-B-Raf, p38, p-p38, ERK, p-ERK (Abcam, Shanghai, China); LMP1, JNK, p-JNK (Santa Cruz, Shanghai, China); p65 and cyclins A1, A2, B1, and D (Boster); cyclin E (Proteintech, Wuhan, China); cyclin-dependent protein kinase (CDK) 1 (Abcam); and CDK2 and 4 (Boster).

Techniques: Real-time Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay, Infection, Negative Control, Control, Transduction

Journal: Cancer communications (London, England)

Article Title: IL-2Rα up-regulation is mediated by latent membrane protein 1 and promotes lymphomagenesis and chemotherapy resistance in natural killer/T-cell lymphoma.

doi: 10.1186/s40880-018-0334-8

Figure Lengend Snippet: Fig. 3 Treating SNK-6 cells constitutively expressing LMP1 for 1 h with the following selective inhibitors of proteins in the MAPK/NF-κB pathway down-regulated IL-2Rα: a 0.1 μM SB590885 (B-Raf inhibitor), b 20 μM PD98059 (ERK inhibitor), c 10 μM SB203580 (p38 inhibitor), d 20 μM SP600125 (JNK inhibitor), or e 100 μM pyrrolidine dithiocarbamate (PDTC, NF-κB inhibitor)

Article Snippet: The membranes were incubated at room temperature for 1 h with one of the following primary antibodies: IL-2Rα, B-Raf, p-B-Raf, p38, p-p38, ERK, p-ERK (Abcam, Shanghai, China); LMP1, JNK, p-JNK (Santa Cruz, Shanghai, China); p65 and cyclins A1, A2, B1, and D (Boster); cyclin E (Proteintech, Wuhan, China); cyclin-dependent protein kinase (CDK) 1 (Abcam); and CDK2 and 4 (Boster).

Techniques: Expressing

Journal: Cancer communications (London, England)

Article Title: IL-2Rα up-regulation is mediated by latent membrane protein 1 and promotes lymphomagenesis and chemotherapy resistance in natural killer/T-cell lymphoma.

doi: 10.1186/s40880-018-0334-8

Figure Lengend Snippet: Fig. 2 LMP1 up-regulated IL-2Rα expression through the MAPK/NF-κB pathway in NK-92 cells. a NK-92 cells transduced with LMP1-encoding lentivirus led to higher levels of p-Raf-B, p-p38, p-JNK, p-ERK, and p65 than transduction with NC lentivirus. b–f NK-92 cells transduced with LMP1-encoding lentivirus were treated for 1 h with the following selective inhibitors of proteins in the MAPK/NF-κB pathway, leading to IL-2Rα down-regulation: b 0.1 μM SB590885 (B-Raf inhibitor), c 20 μM PD98059 (ERK inhibitor), d 10 μM SB203580 (p38 inhibitor), e 20 μM SP600125 (JNK inhibitor), and f 100 μM pyrrolidine dithiocarbamate (PDTC; NF-κB inhibitor)

Article Snippet: The membranes were incubated at room temperature for 1 h with one of the following primary antibodies: IL-2Rα, B-Raf, p-B-Raf, p38, p-p38, ERK, p-ERK (Abcam, Shanghai, China); LMP1, JNK, p-JNK (Santa Cruz, Shanghai, China); p65 and cyclins A1, A2, B1, and D (Boster); cyclin E (Proteintech, Wuhan, China); cyclin-dependent protein kinase (CDK) 1 (Abcam); and CDK2 and 4 (Boster).

Techniques: Expressing, Transduction

Journal: Cancer communications (London, England)

Article Title: IL-2Rα up-regulation is mediated by latent membrane protein 1 and promotes lymphomagenesis and chemotherapy resistance in natural killer/T-cell lymphoma.

doi: 10.1186/s40880-018-0334-8

Figure Lengend Snippet: Fig. 5 Growth curves of a NK-92 and b SNK-6 cells. Numbers of cells were determined at 24-h intervals after infection with lentivirus encoding IL-2Rα or negative control (NC) lentivirus. Data are mean ± SD. *P < 0.05, **P < 0.01 vs uninfected cells (control) or cells infected with NC lentivirus

Article Snippet: The membranes were incubated at room temperature for 1 h with one of the following primary antibodies: IL-2Rα, B-Raf, p-B-Raf, p38, p-p38, ERK, p-ERK (Abcam, Shanghai, China); LMP1, JNK, p-JNK (Santa Cruz, Shanghai, China); p65 and cyclins A1, A2, B1, and D (Boster); cyclin E (Proteintech, Wuhan, China); cyclin-dependent protein kinase (CDK) 1 (Abcam); and CDK2 and 4 (Boster).

Techniques: Infection, Negative Control, Control

Journal: Cancer communications (London, England)

Article Title: IL-2Rα up-regulation is mediated by latent membrane protein 1 and promotes lymphomagenesis and chemotherapy resistance in natural killer/T-cell lymphoma.

doi: 10.1186/s40880-018-0334-8

Figure Lengend Snippet: Fig. 4 Non-transduced NK-92 and SNK-6 cells (control) as well as the same cell lines transduced with lentivirus encoding IL-2Rα or negative control lentivirus (NC) were analyzed in terms of levels of a IL-2Rα mRNA by quantitative real-time PCR, b IL-2Rα protein by Western blot, and c soluble IL-2Rα protein in culture supernatant by ELISA. **P < 0.01 vs control or NC

Article Snippet: The membranes were incubated at room temperature for 1 h with one of the following primary antibodies: IL-2Rα, B-Raf, p-B-Raf, p38, p-p38, ERK, p-ERK (Abcam, Shanghai, China); LMP1, JNK, p-JNK (Santa Cruz, Shanghai, China); p65 and cyclins A1, A2, B1, and D (Boster); cyclin E (Proteintech, Wuhan, China); cyclin-dependent protein kinase (CDK) 1 (Abcam); and CDK2 and 4 (Boster).

Techniques: Control, Transduction, Negative Control, Real-time Polymerase Chain Reaction, Western Blot, Enzyme-linked Immunosorbent Assay

Journal: Cancer communications (London, England)

Article Title: IL-2Rα up-regulation is mediated by latent membrane protein 1 and promotes lymphomagenesis and chemotherapy resistance in natural killer/T-cell lymphoma.

doi: 10.1186/s40880-018-0334-8

Figure Lengend Snippet: Fig. 6 Apoptosis assay of NK-92 and SNK-6 cells transduced with lentivirus-encoded IL-2Rα or negative control (NC) lentivirus vector. Cells were collected and resuspended in binding buffer containing Annexin V-PE and 7-AAD, then processed for flow cytometry. In each box, the left lower region indicates viable cells negative for 7-AAD and Annexin V-PE; the left upper region indicates damaged cells; the right lower region indicates early-stage apoptotic cells positive for Annexin V-PE but negative for 7-AAD; and the right upper region indicates late-stage apoptotic or dead cells positive for Annexin V-PE and 7-AAD. Numbers indicate the percentage of cells in each region. Results are representative of three independent experiments

Article Snippet: The membranes were incubated at room temperature for 1 h with one of the following primary antibodies: IL-2Rα, B-Raf, p-B-Raf, p38, p-p38, ERK, p-ERK (Abcam, Shanghai, China); LMP1, JNK, p-JNK (Santa Cruz, Shanghai, China); p65 and cyclins A1, A2, B1, and D (Boster); cyclin E (Proteintech, Wuhan, China); cyclin-dependent protein kinase (CDK) 1 (Abcam); and CDK2 and 4 (Boster).

Techniques: Apoptosis Assay, Transduction, Negative Control, Plasmid Preparation, Binding Assay, Flow Cytometry

Journal: Cancer communications (London, England)

Article Title: IL-2Rα up-regulation is mediated by latent membrane protein 1 and promotes lymphomagenesis and chemotherapy resistance in natural killer/T-cell lymphoma.

doi: 10.1186/s40880-018-0334-8

Figure Lengend Snippet: Fig. 8 Influence of IL-2Rα and its antibody on sensitivity to a, b gemcitabine, c, d doxorubicin, and e, f l-asparaginase (l-Asp) in SNK-6 cells. Cells overexpressed with IL-2Rα were treated with anti-IL-2Rα antibody (0.2 μg/ml) and different concentrations of one of the three drugs. Cell viability was determined by CCK-8 assay. IC50 values are presented as mean (95% confidence interval)

Article Snippet: The membranes were incubated at room temperature for 1 h with one of the following primary antibodies: IL-2Rα, B-Raf, p-B-Raf, p38, p-p38, ERK, p-ERK (Abcam, Shanghai, China); LMP1, JNK, p-JNK (Santa Cruz, Shanghai, China); p65 and cyclins A1, A2, B1, and D (Boster); cyclin E (Proteintech, Wuhan, China); cyclin-dependent protein kinase (CDK) 1 (Abcam); and CDK2 and 4 (Boster).

Techniques: CCK-8 Assay

Journal: Cancer communications (London, England)

Article Title: IL-2Rα up-regulation is mediated by latent membrane protein 1 and promotes lymphomagenesis and chemotherapy resistance in natural killer/T-cell lymphoma.

doi: 10.1186/s40880-018-0334-8

Figure Lengend Snippet: Fig. 7 Cell cycle distribution of a NK-92 and b SNK-6 cells expressing lentivirus-encoded IL-2Rα. **P < 0.01 vs uninfected cells (control) or cells infected with NC lentivirus

Article Snippet: The membranes were incubated at room temperature for 1 h with one of the following primary antibodies: IL-2Rα, B-Raf, p-B-Raf, p38, p-p38, ERK, p-ERK (Abcam, Shanghai, China); LMP1, JNK, p-JNK (Santa Cruz, Shanghai, China); p65 and cyclins A1, A2, B1, and D (Boster); cyclin E (Proteintech, Wuhan, China); cyclin-dependent protein kinase (CDK) 1 (Abcam); and CDK2 and 4 (Boster).

Techniques: Expressing, Control, Infection