Journal: Theranostics
Article Title: CD4+ T cell-mimicking nanoparticles encapsulating DIABLO/SMAC mimetics broadly neutralize HIV-1 and selectively kill HIV-1-infected cells
doi: 10.7150/thno.59728
Figure Lengend Snippet: DIABLO/SMAC mimetic loaded TNP are more effective than free drug at killing HIV-1-infected cells. HIV-1-infected T CM (HIV-T CM ) or macrophages (HIV-Mφ) were exposed to vehicle, DIABLO/SMAC mimetic loaded TNP (LCL-161/LCL or AT-406/AT), or free drug for 4 h, washed three times with PBS, and then incubated for a further 24 h in fresh media. HIV-T CM were treated with 100 nM of each drug/TNP and HIV-Mφ with 1 µM drug/TNP. n = 4. (A) HIV-T CM were surface stained for exposed phosphatidylserine (Phosphatidylserine PS [phycoerythrin]), fixed, washed then harvested and analyzed by flow cytometry. A representative donor is shown. (B) HIV-Mφ were surface stained for exposed phosphatidylserine, fixed, permeabilized, then stained for HIV-1 p55+p24+p17 (HIV-1 Ag AF488 [Alexa Fluor 488]), washed, then harvested and analyzed by flow cytometry. A representative donor is shown. (C) Left , representative western blots of PARP1, cleaved PARP1 (cPARP1), RIPK1, cleaved RIPK1 (cRIPK1), caspase 8 (CASP8) and β-actin (ACTB). Right, d ensitometric analysis of blots is shown.
Article Snippet: Cell death was estimated by staining cells for HIV-1 antigens and exposed phosphatidylserine using anti-phosphatidylserine (Millipore Cat# 05-719, RRID:AB_309933), anti-HIV-1 p55+p24+p17 (Bioss Cat# BS-4942R, RRID:AB_2892128), Alexa Fluor 488 (AF488) conjugated goat anti-rabbit IgG (Cell Signaling Technology Cat# 4412, RRID:AB_1904025), and phycoerythrin (PE) conjugated goat anti-mouse IgG (Cell Signaling Technology Cat# 8887, RRID:AB_2797678) using a procedure as previously described .
Techniques: Infection, Incubation, Staining, Flow Cytometry, Western Blot