Traditionally, Toll-like receptor 9 (TLR9) signals through an MyD88-dependent cascade that results in proinflammatory gene transcription. Recently, it was reported that TLR9 also participates in a stress tolerance signaling cascade in nonimmune cells. In this noncanonical pathway, TLR9 binds to and inhibits sarcoplasmic/endoplasmic reticulum Ca 21 -ATPase 2 (SERCA2), modulating intracellular calcium handling, and subsequently resulting in the activation of 59-AMP–activated protein kinase a (AMPKa). We have previously reported that TLR9 causes increased contraction in isolated arteries; however, the mechanisms underlying this vascular dysfunction need to be further clarified. Therefore, we hypothesized that noncanonical TLR9 signaling was also present in vascular smooth muscle cells (VSMCs) and that it mediates enhanced contractile responses through SERCA2 inhibition. To test these hypotheses, aortic microsomes, aortic VSMCs, and isolated arteries from male Sprague-Dawley rats were incubated with vehicle or TLR9 agonist (ODN2395). Despite clear AMPKa activation after treatment with ODN2395, SERCA2 activity was unaffected. Alternatively, ODN2395 caused the phosphorylation of AMPKa via transforming growth factor b–activated kinase 1 (TAK1), a kinase involved in TLR9 inflammatory signaling. Downstream, we hypothesized that that TLR9 activation of AMPKa may be important in mediating actin cytoskeleton reorganization. ODN2395 significantly increased the filamentous-to-globular actin ratio, as well as indices of RhoA /Rho-associated protein kinase (ROCK) activation, with the latter being prevented by AMPKa inhibition. In conclusion, AMPKa phosphorylation after TLR9 activation in VSMCs appears to be an extension of traditional inflammatory signaling via TAK1, as opposed to SERCA2 inhibition and the noncanonical pathway. Nonetheless, TLR9-AMPKa signaling can mediate VSMC function via RhoA /ROCK activation and actin polymerization.
|Original language||English (US)|
|Number of pages||12|
|Journal||Journal of Pharmacology and Experimental Therapeutics|
|State||Published - Apr 2018|
ASJC Scopus subject areas
- Molecular Medicine