pwpi empty vector  (Addgene inc)

Journal: PLoS Pathogens

Article Title: HTLV-1 Tax-Mediated Inhibition of FOXO3a Activity Is Critical for the Persistence of Terminally Differentiated CD4 + T Cells

doi: 10.1371/journal.ppat.1004575

Figure Lengend Snippet: Briefly, activated T cells were transduced by lentiviral particles (LVP) expressing or not Tax. (A) Tax expression at 48 h on transduced T cells in response to LVP Tax concentrations (5–160 ng/10 6 cells; n = 3). (B) At 24 and 48 h post-transduction, total RNA was extracted, and subjected to Biomark analyses. Heatmaps of genes significantly modulated following Tax expression. (C) Table showing fold changes and P -values for several modulated FOXO3a targets genes on Tax-transduced T cells (n = 5; paired t test).

Article Snippet: The lentiviral vector pWPI (empty vector), packaging plasmid psPAX2 and envelope plasmid pMD2G were generously provided by VGTI-Florida, whereas pCLXSN-Tax vector was purchased from Addgene (ref: 44038; MA, USA).

Techniques: Expressing, Transduction

Journal: PLoS Pathogens

Article Title: HTLV-1 Tax-Mediated Inhibition of FOXO3a Activity Is Critical for the Persistence of Terminally Differentiated CD4 + T Cells

doi: 10.1371/journal.ppat.1004575

Figure Lengend Snippet: Briefly, activated T cells were transduced by lentiviral particles (LVP) expressing or not Tax for 2–28 days. (A) FOXO3a signaling on transduced T cells expressing or not Tax at 48 h determined by immunoblotting (n = 3). (B) Densitometric quantification of specific bands was performed using ImageJ software. Results shown represent the mean relative expression ± SD of 3 independent experiments. (C–E) Persistence and stepwise differentiation of activated CD4 + T cells transduced with LVP expressing Tax in the presence or absence of AKT i (n = 5). (C) Absolute numbers of total viable CD3 + T cells were determined by trypan blue exclusion. Results are expressed in log 2 scale. P values were determined based on the comparison with LVP empty -transduced cells. The underlined numbers represent the half-life of cultured LVP empty (black) and LVP Tax +AKT i (green) conditions. (D) Differentiation status of transduced T cells subsets at 7–28 dpt. Pie charts are representative of raw data from five independent experiments. (E) Correlation between the absolute numbers of viable transduced T cells at 28 days and the levels of expression of FOXO3a-related proteins at 48 h are also shown (n = 9; Spearman test).

Article Snippet: The lentiviral vector pWPI (empty vector), packaging plasmid psPAX2 and envelope plasmid pMD2G were generously provided by VGTI-Florida, whereas pCLXSN-Tax vector was purchased from Addgene (ref: 44038; MA, USA).

Techniques: Expressing, Western Blot, Software, Transduction, Cell Culture