Transfection of human glioblastoma astrocytoma U373-CD14 with siRNA using DreamFect Gold

Human U373-CD14 were transfected in 6-well plate with 20 nM siRNA using DreamFect Gold. Gene silencing was assessed 24 h after transfection.

This article demonstrates the high efficiency of DreamFect Gold from OZ Biosciences for siRNA transfection into human glioblastoma astrocytoma.

article reference: J Biol Chem. 2014 Apr 9. 

Modulation of TLR3, TLR4 and TLR7 mediated IFN-β, Rantes and TNFα production by HIVEP1.

Butt AQ, Miggin SM.

Abstract

Upon activation of Toll-like receptors (TLRs), recruitment of the adaptor proteins, MyD88, Mal, TRIF and TRAM is facilitated which then elicits downstream signaling cascades leading to the production of pro-inflammatory cytokines and pathogen elimination. A clearer understanding of the biological processes that modulate TLR functionality is vital towards deciphering their role in pathogen clearance. Herein, we identified HIVEP1 as a novel TLR adaptor interacting partner and a crucial member of the TLR signaling complex. Given the limited understanding of the biological function of HIVEP1, we sought to decipher its role in innate immunity. To this end, we show that HIVEP1 inhibits TLR3, TLR4 and TLR7 ligand-induced IFN-β and Rantes, but enhances TNFα production. We also show that HIVEP1 inhibits TLR3, TLR4 and TLR7 mediated IFN-β and CCL5 reporter gene activity, but enhances NF-κB reporter gene activity. Importantly, whilst HIVEP1 suppresses RV-16, VSV, EMCV and SV-mediated Rantes and Type I IFN induction, it enhances TNFα induction. Mechanistic analysis revealed that HIVEP1 precludes the phosphorylation of TBK1 and concomitant phosphorylation and nuclear translocation of IRF3 and IRF7. Intriguingly, our study shows that HIVEP1 plays an opposing role in TLR3, TLR4 and TLR7-mediated NF-κB activation whereby it is required for the phosphorylation of p65, IKKα and IKKβ and degradation of IκBα. Conclusively, our study shows for the first time that HIVEP1 plays a major regulatory role in antiviral immune responses through the modulation of TLR functionality and that HIVEP1 plays a functionally disparate role in TLR-driven NF-κB and IRF3/7 signaling.

DreamFect™Gold allows transfecting all types of nucleic acids with a very high efficiency. Due to its formulation, DreamFect™Gold delivers a large quantity of nucleic acids leading to higher protein expression compared to other transfection reagents. It is fully biodegradable and does not interfere with cellular mechanisms.