Background:
We showed that injection of recombinant heat shock protein 27 (rHSP27) into ApoE
-/-
mice reduces atherosclerosis lesions and total serum cholesterol levels. Extracellular HSP27 also interacts with scavenger receptor (SR)-A and inhibits acLDL uptake in macrophages, but the role of SR-A in HSP27 atheroprotection remains unclear.
Purpose:
To elucidate the mechanism of HSP27 atheroprotection by exploring if the presence of SR-A is required to facilitate its salutary effects.
Methods/Results:
rHSP27 treatment of bone-marrow derived macrophages from ApoE
-/-
mice reduced acLDL uptake (38%, p<0.05) using flow cytometry - effects not observed (p=0.171) in cells from ApoE
-/-
SR-A
-/-
mice. rHSP27 reduced acLDL binding (38%, p<0.01) and uptake (34%, p<0.01) in THP-1 macrophages associated with downregulation of SR-A transcript (46%, p<0.01) and total protein levels (63%, p<0.001). Flow cytometry and confocal microscopy revealed reduction in SR-A expression at the cell surface. To explore the signaling involved, NF-kB activation was assessed in THP-1 cells stably transfected with an NF-kB reporter. rHSP27 increased NF-kB activity (7 fold, p<0.001) and induced translocation of the NF-kB p65 subunit from the cytosol to the nucleus - effects that were inhibited (p<0.05) by the NF-kB antagonist, BAY. Addition of BAY inhibited the effect of rHSP27 on SR-A gene and protein expression and acLDL uptake (p<0.05). To explore the importance of SR-A in HSP27 atheroprotection, rHSP25 (mouse homologue) or PBS was administered to ApoE
-/-
and ApoE
-/-
SR-A
-/-
mice for 3 weeks on a high fat diet. Treatment of ApoE
-/-
mice revealed reductions of en face aortic lesions (39%, p<0.001), SR-A expression in the intima, and total serum cholesterol levels (32%, p<0.01). In the absence of SR-A, HSP27 atheroprotection was abolished (p=0.79) despite similar decreases in serum cholesterol levels (25%, p<0.05).
Conclusions:
Extracellular HSP27 inhibits foam cells by downregulating SR-A expression via an NF-κB-dependent mechanism. HSP27 atheroprotection requires the presence of SR-A expression in vivo. In the absence of SR-A, loss of atheroprotection occurs despite reductions in total serum cholesterol levels, suggesting SR-A as a novel target for HSP27 in the vessel wall.
Extracellular Release of the Atheroprotective Heat Shock Protein 27 Is Mediated by Estrogen and Competitively Inhibits acLDL Binding to Scavenger Receptor-A