• Home
  • Search Results
  • Extracellular HSP70 Activates ERK1/2, NF-kB and Pro-Inflammatory Gene Transcription Through Binding with RAGE in A549 Human Lung Cancer Cells.

Extracellular HSP70 Activates ERK1/2, NF-kB and Pro-Inflammatory Gene Transcription Through Binding with RAGE in A549 Human Lung Cancer Cells.

Cellular physiology and biochemistry : international journal of experimental cellular physiology, biochemistry, and pharmacology (2017-08-30)
Nauana Somensi, Pedro Ozorio Brum, Vitor de Miranda Ramos, Juciano Gasparotto, Alfeu Zanotto-Filho, Diana Carolina Rostirolla, Maurilio da Silva Morrone, José Claudio Fonseca Moreira, Daniel Pens Gelain
ABSTRACT

Heat shock protein 70 (HSP70) has been recently described with extracellular actions, where it is actively released in inflammatory conditions. Acting as DAMPs (damage associated molecular pattern), extracellular HSP70 (eHSP70) interacts with membrane receptors and activates inflammatory pathways. At this context, the receptor for advanced glycation endproducts (RAGE) emerges as a possible candidate for interaction with eHSP70. RAGE is a pattern-recognition receptor and its expression is increased in several diseases related to a chronic pro-inflammatory state. One of the main consequences of RAGE ligand-binding is the ERK1/2 (extracellular signal-regulated kinases)-dependent activation of NF-kB (nuclear factor kappa B), which leads to expression of TNF-α (tumor necrosis factor alpha) and other cytokines. The purpose of this work is to elucidate if eHSP70 is able to evoke RAGE-dependent signaling using A549 human lung cancer cells, which constitutively express RAGE. Immunoprecipitation and protein proximity assay were utilized to demonstrate the linkage between RAGE and eHSP70. To investigate RAGE relevance on cell response to eHSP70, siRNA was used to knockdown the receptor expression. Signaling pathways activation were evaluated by western blotting, gene reporter luciferase and real time quantitative PCR. Protein eHSP70 shown to be interacting physically with the receptor RAGE in our cell model. Treatment with eHSP70 caused ERK1/2 activation and NF-κB transactivation impaired by RAGE knockdown. Moreover, the stimulation of pro-inflammatory cytokines expression by eHSP70 was inhibited in RAGE-silenced cells. Finally, conditioned medium of eHSP70-treated A549 cells caused differential effects in monocytes cytokine expression when A549 RAGE expression is inhibited. Our results evidence eHSP70 as a novel RAGE agonist capable of influence the cross-talk between cancer and immune system cells.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Duolink® In Situ Red Starter Kit Mouse/Rabbit
Sigma-Aldrich
Duolink® In Situ Detection Reagents Red
Sigma-Aldrich
Duolink® In Situ PLA® Probe Anti-Mouse MINUS, Affinity purified Donkey anti-Mouse IgG (H+L)
Sigma-Aldrich
Duolink® In Situ PLA® Probe Anti-Rabbit PLUS, Affinity purified Donkey anti-Rabbit IgG (H+L)
Sigma-Aldrich
Duolink® In Situ Orange Starter Kit Mouse/Rabbit
Sigma-Aldrich
Duolink® In Situ PLA® Probe Anti-Mouse PLUS
Sigma-Aldrich
Duolink® In Situ PLA® Probe Anti-Rabbit MINUS
Sigma-Aldrich
Duolink® In Situ Detection Reagents Orange
Sigma-Aldrich
Duolink® In Situ Wash Buffers, Fluorescence
Sigma-Aldrich
Duolink® In Situ Detection Reagents Green
Sigma-Aldrich
Duolink® In Situ Detection Reagents FarRed
Sigma-Aldrich
Duolink® In Situ Mounting Medium with DAPI
Sigma-Aldrich
Duolink® In Situ Probemaker MINUS
Sigma-Aldrich
Duolink® In Situ Probemaker PLUS
Sigma-Aldrich
Duolink® In Situ Detection Reagents Brightfield
Sigma-Aldrich
Duolink® flowPLA Detection Kit - Green, Duolink® PLA kit for Flow Cytometry with Green Detection
Sigma-Aldrich
Duolink® In Situ PLA® Probe Anti-Goat PLUS
Sigma-Aldrich
Duolink® flowPLA Detection Kit - FarRed, Duolink® PLA kit for Flow Cytometry with FarRed Detection
Sigma-Aldrich
Duolink® PLA Control Kit - PPI, Control Kit to reliably detect Protein-Protein Interactions (PPI) with Duolink® PLA
Sigma-Aldrich
Duolink® In Situ PLA® Probe Anti-Goat MINUS, Affinity purified Donkey anti-Goat IgG (H+L)
Sigma-Aldrich
Duolink® In Situ Wash Buffer, Brightfield
Sigma-Aldrich
Duolink® flowPLA Detection Kit - Red, DUOLINK® : PLA kit for Flow Cytometry with Red Detection
Sigma-Aldrich
Duolink® In Situ Red Starter Kit Mouse/Goat
Sigma-Aldrich
Duolink® In Situ Red Starter Kit Goat/Rabbit
Sigma-Aldrich
Duolink® In Situ Microplate Nuclear Stain, Anti-Fade
Sigma-Aldrich
Duolink® flowPLA Detection Kit - Orange, Duolink® PLA kit for Flow Cytometry with Orange Detection
Sigma-Aldrich
Duolink® In Situ Orange Starter Kit Goat/Rabbit
Sigma-Aldrich
HMG-1 human, lyophilized powder, ≥90% (SDS-PAGE), Histidine-tagged, recombinant, expressed in E. coli
Sigma-Aldrich
Duolink® In Situ Orange Starter Kit Mouse/Goat
Sigma-Aldrich
Duolink® In Situ Microplate Heat Transfer Block