Twist-related protein 1 negatively regulated osteoblastic transdifferentiation of human aortic valve interstitial cells by directly inhibiting runt-related transcription factor 2.
OBJECTIVE
Valve calcification involves transdifferentiation of valve interstitial cells (VICs) into osteoblasts. Twist-related protein 1 (TWIST1) has been established as a negative regulator of osteoblast differentiation in both mouse and human mesenchymal stem cells, but its function in human aortic VICs is unknown. In our study, we determined the mechanism of TWIST1 action in regulating osteoblastic transdifferentiation of human aortic VICs.


METHODS
Human calcified and noncalcified aortic valves were examined for TWIST1 expression. Human aortic VICs were isolated and cultured.


RESULTS
The data showed that calcified aortic valves express lower levels of TWIST1. In vitro experiments showed that TWIST1 overexpression inhibited the transdifferentiation of VICs into osteoblasts by decreasing the expression of runt-related transcription factor 2 (RUNX2) and its downstream osteoblastic markers. Through chromatin immunoprecipitation and dual luciferase assays, we found that TWIST1 repressed the expression of RUNX2 by directly binding to an E-box located at -820 bp of the RUNX2 P2 promoter region and inhibiting its activity.


CONCLUSIONS
Our study results suggest that TWIST1 could play an important role in preventing human aortic valve calcification by negatively regulating osteoblastic transdifferentiation of human aortic VICs through direct inhibition of RUNX2.
Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai, China.
Xi-Wu
XW
Zhang
Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai, China.
Bo-Yao
BY
Zhang
Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai, China; Department of Cardiothoracic Surgery, No. 153 Hospital, Zhengzhou, China.
Shu-Wei
SW
Wang
Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai, China.
De-Jun
DJ
Gong
Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai, China.
Lin
L
Han
Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai, China. Electronic address: zhiyunx@hotmail.com.
Zhi-Yun
ZY
Xu
Department of Cardiothoracic Surgery, Changhai Hospital, Second Military Medical University, Shanghai, China.
Xiao-Hong
XH
Liu
D000328:Adult; D001021:Aortic Valve / Q000166:cytology* / Q000378:metabolism / Q000473:pathology*; D001024:Aortic Valve Stenosis / Q000378:metabolism*; D015153:Blotting, Western; D002114:Calcinosis / Q000378:metabolism*; D054338:Cell Transdifferentiation; D047369:Chromatin Immunoprecipitation; D050659:Core Binding Factor Alpha 1 Subunit / Q000378:metabolism*; D005260:Female; D005434:Flow Cytometry; D006801:Humans; D007124:Immunoenzyme Techniques; D066298:In Vitro Techniques; D008156:Luciferases; D008297:Male; D008875:Middle Aged; D009687:Nuclear Proteins / Q000378:metabolism*; D010006:Osteoblasts / Q000166:cytology* / Q000378:metabolism; D011401:Promoter Regions, Genetic; D060888:Real-Time Polymerase Chain Reaction; D015398:Signal Transduction; D051879:Twist-Related Protein 1 / Q000378:metabolism*
D050659:Core Binding Factor Alpha 1 Subunit; D009687:Nuclear Proteins; C493709:RUNX2 protein, human; C103173:TWIST1 protein, human; D051879:Twist-Related Protein 1; D008156:Luciferases