Premium
Inhibition of endogenous heat shock protein 70 attenuates inducible nitric oxide synthase induction via disruption of heat shock protein 70/Na + /H + exchanger 1‐Ca 2+ ‐calcium‐calmodulin‐dependent protein kinase II/transforming growth factor β‐activated kinase 1‐nuclear factor‐κB signals in BV‐2 microglia
Author(s) -
Huang Chao,
Lu Xu,
Wang Jia,
Tong Lijuan,
Jiang Bo,
Zhang Wei
Publication year - 2015
Publication title -
journal of neuroscience research
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.72
H-Index - 160
eISSN - 1097-4547
pISSN - 0360-4012
DOI - 10.1002/jnr.23571
Subject(s) - hsp70 , nitric oxide synthase , iκb kinase , phosphorylation , nitric oxide , protein kinase a , chemistry , kinase , heat shock protein , iκbα , microbiology and biotechnology , nf κb , signal transduction , biochemistry , biology , organic chemistry , gene
Inducible nitric oxide synthase (iNOS) critically contributes to inflammation and host defense. The inhibition of heat shock protein 70 (Hsp70) prevents iNOS induction in lipopolysaccharide (LPS)‐stimulated macrophages. However, the role and mechanism of endogenous Hsp70 in iNOS induction in microglia remains unclear. This study addresses this issue in BV‐2 microglia, showing that Hsp70 inhibition or knockdown prevents LPS‐induced iNOS protein expression and nitric oxide production. Real‐time PCR experiments showed that LPS‐induced iNOS mRNA transcription was blocked by Hsp70 inhibition. Further studies revealed that the inhibition of Hsp70 attenuated LPS‐stimulated nuclear translocation and phosphorylation of nuclear factor (NF)‐κB as well as the degradation of inhibitor of κB (IκB)‐α and phosphorylation of IκB kinase β (IKKβ). This prevention effect of Hsp70 inhibition on IKKβ–NF‐κB activation was found to be dependent on the Ca 2+ /calcium–calmodulin‐dependent protein kinase II (CaMKII)/transforming growth factor β‐activated kinase 1 (TAK1) signals based on the following observations: 1) chelation of intracellular Ca 2+ or inhibition of CaMKII reduced LPS‐induced increases in TAK1 phosphorylation and 2) Hsp70 inhibition reduced LPS‐induced increases in CaMKII/TAK1 phosphorylation, intracellular pH value, [Ca 2+ ] i , and CaMKII/TAK1 association. Mechanistic studies showed that Hsp70 inhibition disrupted the association between Hsp70 and Na + /H + exchanger 1 (NHE1), which is an important exchanger responsible for Ca 2+ influx in LPS‐stimulated cells. These studies demonstrate that the inhibition of endogenous Hsp70 attenuates the induction of iNOS, which likely occurs through the disruption of NHE1/Hsp70‐Ca 2+ ‐CaMKII/TAK1–NF‐κB signals in BV‐2 microglia, providing further insight into the functions of Hsp70 in the CNS. © 2015 Wiley Periodicals, Inc.