Death-associated protein kinase 3 mediates vascular structural remodelling via stimulating smooth muscle cell proliferation and migration

Abstract

Death-associated protein kinase 3 (DAPK3) also known as zipper-interacting kinase is a serine/threonine kinase that mainly regulates cell death and smooth muscle contraction. We have previously found that protein expression of DAPK3 increases in the mesenteric artery from spontaneously hypertensive rats (SHRs) and that DAPK3 mediates the development of hypertension in SHRs partly through promoting reactive oxygen species-dependent vascular inflammation. However, it remains to be clarified how DAPK3 controls smooth muscle cell (SMC) proliferation and migration, which are also important processes for hypertension development. We, therefore, sought to investigate whether DAPK3 affects SMC proliferation and migration. siRNA against DAPK3 significantly inhibited platelet-derived growth factor (PDGF)-BB-induced SMC proliferation and migration as determined by bromodeoxyuridine (BrdU) incorporation and a cell counting assay as well as a Boyden chamber assay respectively. DAPK3 siRNA or a pharmacological inhibitor of DAPK3 inhibited PDGF-BB-induced lamellipodia formation as determined by rhodamine-phalloidin staining. DAPK3 siRNA or the DAPK inhibitor significantly reduced PDGF-BB-induced activation of p38 and heat-shock protein 27 (HSP27) as determined by Western blotting. In ex vivo studies, PDGF-BB-induced SMC out-growth was significantly inhibited by the DAPK inhibitor. In vivo, the DAPK inhibitor significantly prevented carotid neointimal hyperplasia in a mouse ligation model. The present results, for the first time, revealed that DAPK3 mediates PDGF-BB-induced SMC proliferation and migration through activation of p38/HSP27 signals, which may lead to vascular structural remodelling including neointimal hyperplasia. The present study suggests DAPK3 as a novel pharmaceutical target for the prevention of hypertensive cardiovascular diseases.

A novel inhibitory effect of oxazol-5-one compounds on ROCKII signaling in human coronary artery vascular smooth muscle cells

“…The data presented herein reveal DI to possess similar efficacy toward both ZIPK and ROCKII activities; nevertheless, the DI compound has been used in two recent studies by Usui and colleagues to assess the role of ZIPK in cardiovascular disease3144. In the first study, the in vivo administration of DI to spontaneously-hypertensive rats was associated with decreases in blood pressure, reactive oxygen species (ROS) production, inflammatory responses, as well as vascular smooth muscle contractility and hypertrophy31.…”

“…In vitro treatment of human umbilical vein endothelial cells with DI could also suppress tumor necrosis factor (TNF)α-induced inflammatory responses via ROS-dependent mechanisms. In a second study, the DI compound was used in vitro to link ZIPK with PDGF-BB-induced vascular SMC proliferation and migration through p38MAPK and HSP27 pathways44. Additional in vivo treatments with DI were shown to attenuate vascular structural remodeling during carotid neointimal hyperplasia.…”

“…The relationship between ROCKII and COX-2 was previously investigated with results linking ROCKII with the stimulation of COX-2, and subsequent reduction of E-cadherin and α-catenin along with disruption of adherens junction formation and increased cellular motility62. Experimental approaches that applied both siRNA and DI treatments provided links between ZIPK and a group of downstream targets (HSP27, ERK1/2 and p38MAPK) involved in reorganization of the actin cytoskeleton during vascular smooth muscle remodeling44. But, there have been previously described linkages between PDGF-BB-induced SMC hypertrophy and ROCK signaling in studies employing Y2763263.…”

“…The in vitro characterization of the 2-phenyl-4-(3-pyridinylmethylene)-5(4H)-oxazolone inhibitor (also known as DI) suggested specificity and potency of this compound toward ZIPK4142. More recent applications of the DI compound in VSM revealed novel functions of ZIPK in (i) development of essential hypertension in mesenteric arteries of the spontaneously hypertensive rat31, and vascular inflammatory and hypertrophic remodeling responses44. Based on these data, we employed the DI compound in studies to define the functional targets of ZIPK in human coronary artery VSMCs, and confirmed that the DI compound possesses potent inhibitory action against ZIPK.…”

De‐novo‐Fragmententwurf für die Wirkstoffforschung und chemische Biologie

“…[19] Nachdem wir biochemisch bestätigen konnten, dass Verbindung 2 DAPK3 inhibiert, entwickelten wir ein Strukturmodell, um die biochemischen Daten besser zu verstehen und ein Hit-to-Lead-Programm zu initiieren. Die Blockade von DAPK3 führt zu vaskulärer Umstrukturierung, die bereits für die Unterdrückung der Neointimalhyperplasie in vivo gezeigt werden konnte.…”