Deficiency of CCAAT/enhancer-binding protein homologous protein (CHOP) prevents diet-induced aortic valve calcification in vivo.

Aortic valve (AoV) calcification is common in aged populations. Its subsequent aortic stenosis has been linked with increased morbidity, but still has no effective pharmacological intervention. Our previous data show endoplasmic reticulum (ER) stress is involved in AoV calcification. Here, we investigated whether deficiency of ER stress downstream effector CCAAT/enhancer-binding protein homology protein (CHOP) may prevent development of AoV calcification. AoV calcification was evaluated in Apoe-/- mice (n = 10) or in mice with dual deficiencies of ApoE and CHOP (Apoe-/- CHOP-/- , n = 10) fed with Western diet for 24 weeks. Histological and echocardiographic analysis showed that genetic ablation of CHOP attenuated AoV calcification, pro-calcification signaling activation, and apoptosis in the leaflets of Apoe-/- mice. In cultured human aortic valvular interstitial cells (VIC), we found oxidized low-density lipoprotein (oxLDL) promoted apoptosis and osteoblastic differentiation of VIC via CHOP activation. Using conditioned media (CM) from oxLDL-treated VIC, we further identified that oxLDL triggered osteoblastic differentiation of VIC via paracrine pathway, while depletion of apoptotic bodies (ABs) in CM suppressed the effect. CM from oxLDL-exposed CHOP-silenced cells prevented osteoblastic differentiation of VIC, while depletion of ABs did not further enhance this protective effect. Overall, our study indicates that CHOP deficiency protects against Western diet-induced AoV calcification in Apoe-/- mice. CHOP deficiency prevents oxLDL-induced VIC osteoblastic differentiation via preventing VIC-derived ABs releasing.

Cai Z ，Liu B ，Wei J ，Fu Z ，Wang Y ，Wang Y ，Shen J ，Jia L ，Su S ，Wang X ，Lin X ，Chen H ，Li F ，Wang J ，Xiang M ... -  《-》

Endoplasmic reticulum stress participates in aortic valve calcification in hypercholesterolemic animals.

Cai Z ，Li F ，Gong W ，Liu W ，Duan Q ，Chen C ，Ni L ，Xia Y ，Cianflone K ，Dong N ，Wang DW ... -  《-》

RAGE deficiency alleviates aortic valve calcification in ApoE(-/-) mice via the inhibition of endoplasmic reticulum stress.

Receptor for advanced glycation end products (RAGE) and endoplasmic reticulum (ER) stress have been shown to be involved in calcific aortic valve disease (CAVD). However, the association between RAGE and ER stress remains unknown in the pathogenesis of CAVD. The current study aims to test the hypothesis that RAGE deficiency alleviates aortic valve calcification via the inhibition of ER stress. Up-regulation of RAGE and ER stress markers in calcified human aortic valves were confirmed by immunoblotting. Aortic valve calcification was evaluated in atherosclerotic prone ApoE-/- mice or in mice with dual deficiencies of ApoE and RAGE (ApoE-/-RAGE-/-) fed with high cholesterol diet for 24weeks. Echocardiography and histological examination show that genetic deficiency of RAGE attenuates aortic valve calcification in ApoE-/- mice. Meanwhile, RAGE deficiency inhibited the osteogenic signaling and ER stress activation as well as suppressed macrophage infiltration in vivo. Cultured human aortic valve interstitial cells (AVICs) were treated with high molecular group box 1 protein (HMGB1) as in vitro model. We found that HMGB1 induced osteoblastic differentiation and calcification through RAGE/ER stress. Furthermore, Sox9 up-regulation and intranuclear translocation mediated the pro-osteogenic effect of HMGB1 on AVICs. RAGE or ER stress knockdown reduced the up-regulation of monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-α (TNF-α) in human AVICs exposed to HMGB1.These novel findings demonstrate that RAGE deficiency protects against aortic valve calcification in high cholesterol diet-fed ApoE-/- mice via inhibition of ER stress. HMGB1 induces AVIC osteoblastic differentiation and calcification through RAGE/ER stress/Sox9 pathway.

Wang B ，Cai Z ，Liu B ，Liu Z ，Zhou X ，Dong N ，Li F ... -  《-》

Transforming growth factor-β1 promotes fibrosis but attenuates calcification of valvular tissue applied as a three-dimensional calcific aortic valve disease model.

Jenke A ，Kistner J ，Saradar S ，Chekhoeva A ，Yazdanyar M ，Bergmann AK ，Rötepohl MV ，Lichtenberg A ，Akhyari P ... -  《-》

Histone deacetylase 6 reduction promotes aortic valve calcification via an endoplasmic reticulum stress-mediated osteogenic pathway.

Aortic valve (AoV) calcification occurs via a pathophysiologic process that includes osteoblastic differentiation of valvular interstitial cells (VICs). Histone deacetylases (HDACs) have been shown to be involved in the pathogenesis of vascular diseases. Here, we investigated the role of HDAC6 in AoV calcification. AoV cusps from patients with aortic stenosis (n = 7) and normal controls (n = 7) were subjected to determination of calcified nodules and HDAC6 expression. Human VICs were cultured in osteogenic media and treated with 10 uM tubacin or HDAC6 small interfering RNA silencing to inhibit HDAC6. Treatment with 100 uM tauroursodeoxycholic acid was used to suppress endoplasmic reticulum stress. Activating transcription factor 4 (ATF4) small interfering RNA was used to knock down ATF4. Alizarin red staining was used to evaluate calcified nodules formation of VICs cultured with osteogenic media for 14 days. HDAC6 expression was significantly reduced in AoV tissue of patients with aortic stenosis compared with controls. Tubacin treatment or HDAC6 silencing markedly promoted osteoblastic differentiation accompanied by endoplasmic reticulum stress activation in VICs. The HDAC6 inhibition-induced osteogenic pathway was mediated by endoplasmic reticulum stress/ATF4 pathway as indicated by tauroursodeoxycholic acid pretreatment or ATF4 silencing. Finally, alizarin red staining showed that HDAC6 inhibition promoted osteoblastic differentiation of VICs, which could be suppressed by tauroursodeoxycholic acid. HDAC6 inhibition promotes AoV calcification via an endoplasmic reticulum stress/ATF4-mediated osteogenic pathway. HDAC6 may be a novel target for AoV calcification prevention and treatment.

Fu Z ，Li F ，Jia L ，Su S ，Wang Y ，Cai Z ，Xiang M ... -  《-》