International Journal of Nanomedicine (May 2021)
Nano-Sized Hydroxyapatite Induces Apoptosis and Osteogenic Differentiation of Vascular Smooth Muscle Cells via JNK/c-JUN Pathway
Abstract
Qi Liu,1,* Pingping Xiang,1,* Mingyao Chen,1 Yi Luo,1 Yun Zhao,1,2 Jinyun Zhu,1 Wangwei Jing,1 Hong Yu1 1Department of Cardiology, Cardiovascular Key Laboratory of Zhejiang Province, The Second Affiliated Hospital, Zhejiang University School of Medicine, Hangzhou, Zhejiang Province, 310009, People’s Republic of China; 2The Affiliated Cardiovascular Hospital of Qingdao University, Qingdao, Shandong Province, 266071, People’s Republic of China*These authors contributed equally to this workCorrespondence: Hong YuDepartment of Cardiology, Cardiovascular Key Laboratory of Zhejiang Province, The Second Affiliated Hospital, Zhejiang University School of Medicine, 88 JieFang Road, Hangzhou, Zhejiang Province, 310009, People’s Republic of ChinaTel +86-571-87783992Fax +86-571-88002709Email [email protected]: The deposition of hydroxyapatite (HAp) crystals plays an important role in the development of vascular calcification (VC). This study aimed to demonstrate the effects of nanosized HAp (nHAp) on vascular smooth muscle cells (VSMCs) and VC progression.Methods: Transmission electron microscopy (TEM) was used to examine cellular uptake of nHAp. Cell viability was determined using CCK-8 assay kit. Mitochondrial impairment and reactive oxygen species were detected by TEM and fluorescence dye staining, respectively. Cell apoptosis was detected by Western blot analysis and Annexin V staining. Mouse model of VC was built via applying nHAp on the surface of abdominal aorta. Calcification was visualized by Alizarin red and von Kossa staining.Results: We found that nHAp could promote osteogenic transformation of VSMCs by elevating expression of runt-related factor 2 (Runx2), osteopontin (OPN) and alkaline phosphatase (ALP), impairing function and morphology of mitochondria and inducing apoptosis of VSMCs. More phosphorylation of c-Jun N-terminal protein kinase/c-JUN (JNK/c-JUN) in VSMCs was detected after mixing nHAp with VSMCs. HAp-induced osteogenic transformation of VSMCs was blocked by JNK inhibitor SP600125, resulted in decreased ALP activity, less Runx2 and OPN expressions. SP600125 also inhibited apoptosis of VSMCs. Application of nHAp to outside of aorta induced osteogenic transformation and apoptosis of VSMCs, and significant deposition of calcium on the vessel walls of mice, which can be effectively attenuated by SP600125.Conclusion: JNK/c-JUN signaling pathway is critical for nHAp-induced calcification, which could be a potential therapeutic target for controlling the progression of VC.Keywords: vascular calcification, hydroxyapatite, apoptosis, mitochondria, osteogenic transformation, JNK
