Institute of Cardiovascular Diseases, Medical School of Jiaotong University, Shanghai, People's Republic of China.
Institute of Cardiovascular Diseases, Medical School of Jiaotong University, Shanghai, People's Republic of China ; Department of Cardiology, Rui Jin Hospital, Medical School of Jiaotong University, Shanghai, People's Republic of China.
PLoS One. 2013 Dec 27;8(12):e83853. doi: 10.1371/journal.pone.0083853. eCollection 2013.
This study aimed to identify major proteins in the pathogenesis of coronary artery in-stent restenosis (ISR) in diabetic minipigs with sirolimus-eluting stenting, and to investigate the roles of key candidate molecules, particularly ADAM10, in human arterial smooth muscle cells (HASMCs).
The stents were implanted in the coronary arteries of 15 diabetic and 26 non-diabetic minipigs, and angiography was repeated at six months. The intima of one vascular segment with significant ISR and one with non-ISR in diabetic minipigs were isolated and cultured in conditioned medium (CM). The CM was analyzed by LC-MS/MS to uncover proteins whose levels were significantly increased (≥ 1.5-fold) in ISR than in non-ISR tissues. After literature searching, we focused on the identified proteins, whose biological functions were most potentially related to ISR pathophysiology. Among them, ADAM10 was significantly increased in diabetic and non-diabetic ISR tissues as compared with non-ISR controls. In cell experiments, retrovirus-mediated overexpression of ADAM10 promoted growth and migration of HASMCs. The effects of ADAM10 were more remarkable in high-glucose culture than in low-glucose culture. Using shRNA and an inhibitor of γ-secretase (GSI), we found that the influences of ADAM10 were in part mediated by Notch1 and notch 3 pathway, which up-regulated Notch downstream genes and enhanced nuclear translocation of the small intracellular component of Notch1 and Notch3.
This study has identified significantly increased expression of ADAM10 in the ISR versus non-ISR segment in diabetic minipigs and implicates ADAM10 in the enhanced neointimal formation observed in diabetes after vascular injury.
本研究旨在鉴定雷帕霉素洗脱支架置入后糖尿病小型猪冠状动脉支架内再狭窄（ISR）发病机制中的主要蛋白，并研究关键候选分子，特别是 ADAM10，在人动脉平滑肌细胞（HASMCs）中的作用。
在 15 只糖尿病和 26 只非糖尿病小型猪的冠状动脉中植入支架，6 个月时重复进行血管造影。从糖尿病小型猪的具有显著 ISR 和非 ISR 的一段血管中分离和培养内膜，并在条件培养基（CM）中进行分析。通过 LC-MS/MS 分析 CM，以发现 ISR 组织中水平显著升高（≥1.5 倍）的蛋白。在文献检索后，我们专注于鉴定出的蛋白，其生物学功能最有可能与 ISR 病理生理学相关。其中，ADAM10 在糖尿病和非糖尿病 ISR 组织中的表达明显高于非 ISR 对照组。在细胞实验中，逆转录病毒介导的 ADAM10 过表达促进 HASMCs 的生长和迁移。在高糖培养中，ADAM10 的作用比在低糖培养中更为显著。使用 shRNA 和 γ-分泌酶抑制剂（GSI），我们发现 ADAM10 的影响部分是由 Notch1 和 notch3 通路介导的，该通路上调了 Notch 下游基因，并增强了 Notch1 和 Notch3 的小细胞内成分的核转位。
本研究鉴定了糖尿病小型猪 ISR 与非 ISR 节段中 ADAM10 的表达明显增加，并提示 ADAM10 参与了血管损伤后糖尿病中观察到的新生内膜形成增强。
