Uremic toxic substances are essential elements for enhancing carotid artery stenosis after balloon-induced endothelial denudation: worsening role of the adventitial layer

This study tested the hypothesis that uremic-toxic substances play a crucial role in enhancing left-common carotid artery (LCCA) stenosis after balloon-denudation of LCCA endothelium (BDLCCAE), and that the adventitial layer plays a complementary role in worsening LCCA stenosis. In vitro results showed the protein expressions of inflammation (IL-1β/TNF-α/IL-6), apoptosis (mitochondrial-Bax/cleaved-caspase-3/cleaved-PARP) and autophagy (beclin/Atg5/LC3B-II to LC3B-I ratio) as well as protein (NOX-1/NOX-2/p22phox/oxidized-protein), total cellular (H2DCFDA) and mitochondrial (Mitosox) levels of oxidative stress were significantly increased in p-Cresol-treated umbilical vein endothelial cells (HUVECs) as compared with control, whereas angiogenesis capacity (i.e., Matrigel-assay for HUVECs) exhibited an opposite pattern to inflammation between the two groups (all P < 0.001). Animals (n = 60) were categorized into group 1 (sham-operated control), group 2 (BDLCCAE), group 3 [BDLCCAE + ESRD patient’s serum (1 cc/injection into deprived CA adventitia)], group 4 [BDLCCAE + ESRD patient’s serum (1 cc/injection from peri-adventitia)], and group 5 [BDLCCAE + ESRD patient’s serum (2 cc/by intravenous injection at days 1/3/7/10/14 after BDLCCADE)] and LCCA was harvested by day-21 after BDLCCAE procedure. Nitric-oxide release from LCCA and the LCCA cross-section area significantly and progressively reduced, whereas intimal and medial layers of LCCA significantly and progressively increased from groups 1 to 5 (all P < 0.001). The cellular expressions of inflammation (CD14+) and DNA-damage biomarker (γ-H2AX+) were significantly and progressively increased, whereas endothelial surface markers (CXCR4/vWF+) were significantly and progressively reduced from groups 1 to 5 (all P < 0.0001). Uremic toxins played an essential role in LCCA remodeling and obstruction. LCCA adventitia facilitated the initiation and propagation of LCCA proliferative obstruction.