SMOOTH MUSCLE CELL

P = 0.041, P = 0.028, and P = 0.002, respectively). Expression was observed in macrophages, foam cells, endothelial cells, and smooth muscle cells. Moreover, unstable plaques from men demonstrated greater features of plaque instability compared to unstable plaques from women, including hemorrhage, large lipid core, abundance of inflammatory cells, and fibrous cap infiltration by inflamma- tory cells ( P < 0.05 for all). Conclusion: Our preliminary findings indicate a possible association between androgens and the androgen receptor pathway and plaque instability. With further investigations, our ongoing work will unravel the underlying mechanisms

Design and method: VSMC from resistance arteries from normotensive (NT) and hypertensive (HT) subjects were studied (n = 5). Proteins were labelled with isobaric tandem mass tags and identified by liquid chromatography tandem mass spectrometry. The oxidative proteome was assessed using stable isotopelabelled iodoacetamide to target cysteine thiols. Nox5 silencing was performed by siRNA. Protein expression was detected by western blotting. The inflammatory, pro-fibrotic and mitogenic phenotype of VSMCs was assessed by measuring proinflammatory cytokines (IL-6, IL-8), pro-collagen I in the culture media.

Conclusions:
Our study provides new insights into the proteomic changes related to vascular phenotype in hypertension and demonstrated that Nox5 plays an important role in VSMC phenotypic switching associated with vascular injury and remodelling in hypertension.

S-07-5 ASPROSIN INDUCES VASCULAR ENDOTHELIAL-TO-MESENCHYMAL TRANSITION IN DIABETIC LOWER EXTREMITY PERIPHERAL ARTERY DISEASE
Mei You 1 , Yushuang Liu 1 , Bowen Wang 1 , Zhencheng Yan 1 1 Daping Hospital, Army Medical University, China Objective: Altered adipokine secretion in dysfunctional adipose tissue facilitates the development of atherosclerotic diseases including lower extremity peripheral artery disease (PAD). Asprosin is a recently identified adipokine and displays potent regulatory role in metabolism, but the relationship between asprosin and lower extremity PAD remains uninvestigated. In this study, we investigated the effect of asprosin accounting for the lower extremity vascular injury in diabetic patients based on the metabolic profiles of PAD.
Design and method: 33 type 2 diabetes mellitus (T2DM) patients (DM), 51 T2DM patients with PAD (DM+PAD) and 30 healthy normal control (NC) volunteers were recruited and the blood samples were collected for detecting the circulatory asprosin level and metabolomic screening. RNA sequencing was performed using the aorta tissues from the type 2 diabetic db/db mice and its control db/m mice. Moreover, human umbilical vein endothelial cells (HUVECs) were treated with asprosin to determine its impact on the endothelial-to-mesenchymal transition (EndMT).

Results:
The circulating levels of asprosin in DM+PAD group were significantly higher than that of NC group and the DM group. Circulating asprosin level was remarkably negatively correlated with ankle-brachial index (ABI), even after adjusting for age, sex, body mass index (BMI) and other traditional risk factors of PAD (P = 0.049). Logistic regression analysis revealed that asprosin is an independent risk factor for PAD (OR:3.944, 95%CI:1.656 -9.393, P = 0.002) and receiveroperator characteristic (ROC) curve determined a cut-off value at 188.70 ng/ml with a good sensitivity (74.5%) and specificity (74.6%) of asprosin to distinguish PAD (Area under curve:0.788, 95%CI:0.694 -0.863, P < 0.001). Data from metabolomics displayed a typical characteristics of de novo amino acid synthesis in collagen protein production by myofibroblasts in which TGF-beta signaling pathway might plays a critical role in patients with PAD. In addition, activation of TGF-beta signaling pathway also appeared in the aortic tissue of db/db mice, suggesting a causal relationship between TGF-beta signaling pathway and artery injury. Asprosin directly induces EndMT in HUVECs in a TGF-beta-dependent manner as TGF-beta signaling pathway inhibitor SB431542 erased the promotional effect of asprosin on EndMT.

Conclusions:
Elevated circulatory asprosin level is an independent risk factor of lower extremity PAD and might serve as a diagnostic marker. Mechanistically, asprosin directly induces EndMT that participates in vascular injury via activation of TGF-beta signaling pathway.

S-07-6 NEUTROPHIL EXTRACELLULAR TRAPS AS MEDIATORS OF ENDOTHELIAL DAMAGE IN HYPERTENSION AND DIABETES
Chloe Landry 1,2 , Fengxia Xiao 2 , Jean Francois Thibodeau 2 , Chet E Holterman 2 , Lihua Xhu 2 , Alex Gutsol 2 , Christopher RJ Kennedy 1,2 , Dylan Burger 1,2 1 University of Ottawa, Canada, 2 Ottawa Hospital Research Institute, Canada Objective: Endothelial dysfunction (ED) plays a key role in the pathogenesis of hypertension and diabetes, yet its molecular determinants are poorly understood. Concomitant hypertension and diabetes can synergistically increase damage to the vasculature, with increasing evidence suggesting that resulting immune dysregulation may contribute to endothelial injury. Neutrophil extracellular traps (NETs), networks of chromatin and protein-containing extracellular fibers released to fight infection, are generated from neutrophils following activation of the citrullinating enzyme peptidylarginine deiminase 4 (PAD4). PAD4 has been shown to be upregulated in atherosclerosis, vasculitis, and various inflammatory autoimmune diseases, leading to NET-induced endothelial injury. Whether NETs could contribute to hypertension-and diabetes-induced ED has not been established. The objective of the present study is to investigate the impact of hypertension and diabetes on PAD4 activity and the contribution of NETs to resulting vascular injury.

Design and Methods:
Promyelocytic HL-60 cells were differentiated to neutrophillike cells and treated with angiotensin II (10 -7 M) and/or high glucose (25 mM) for 24 hours, with or without subsequent NET-inducing PMA stimulation (500 nM). NETs were isolated from the culture medium by differential centrifugation and assessed by Western blot analysis of citrullinated histone H3 (citH3, a primary NET component). Isolated NETs were used to treat Human Umbilical Vein Endothelial Cells (HUVECs) at 0.5-500 ng/ml for 24 hours, after which cell viability (XTT cell assay) and proliferation (BrdU assay) were measured. We also assessed NETosis in mice expressing human renin in the liver (TTRhRen) intercrossed with OVE26 diabetic mice [i.e. hypertensive-diabetic, HD mice], compared with wild-type (WT) FVBN/J mice. We examined kidney levels of citH3, neutrophil elastase (NE), and PAD4 expression by immunohistochemistry.