Background: Acute aortic dissection (AAD) is a life-threatening cardiovascular disease with a high mortality rate. Macrophage infiltration and vascular smooth muscle cell (VSMC) apoptosis are crucial in AAD pathogenesis. However, Annexin A1 (ANXA1) may protect against AAD by mitigating aortic damage and cell apoptosis. Therefore, this study assesses the protective effects of ANXA1 in an Angiotensin II (Ang II)-induced AAD mouse model, focusing on survival rates, aortic structural integrity, and the mechanism underlying apoptosis in human aortic vascular smooth muscle cells (HAVSMCs) through regulation of M1 macrophages.

Methods: C57BL/6J mice (n = 40) were divided into four groups: the control group, ANXA1-treated group, Ang II-induced AAD group, and Ang II+ANXA1-treated group. The survival rate was monitored in AAD mice over a 28-day time point, and the diameters of the ascending aorta and aortic arch were evaluated by histological analysis. Immunohistochemistry (IHC) was used to examine cleaved caspase-3 levels in the thoracic aorta to assess VSMC apoptosis. In vitro, M1 macrophages were co-cultured with HAVSMCs. Immunofluorescence was performed to determine macrophage infiltration and oxidative stress by measuring F4/80 levels, Tetramethyl Rhodamine Ethyl Ester (TMRE) mitochondrial membrane potential, TdT-mediated dUTP Nick-End Labeling (TUNEL)-positive cells, and reactive oxygen species (ROS) levels in tissues and cells. Additionally, cleaved caspase-3 expression was evaluated both in vivo and in vitro using Western blot analysis to further assess VSMC apoptosis.

Results: ANXA1 exhibited significant protective effects in the Ang II-induced AAD mouse model. The survival rate in the Ang II+ANXA1 group was markedly higher than in the Ang II group (p < 0.001). The Ang II group had significantly dilated ascending aorta (AA) and aortic arch (Arch) compared to the Ang II+ANXA1 group (p < 0.001), whereas ANXA1 treatment did not alter aortic diameter in healthy mice. ANXA1 treatment substantially alleviated aortic dissection, intramural hematoma, and elastic fiber rupture in AAD mice. The Ang II+ANXA1 group has significantly reduced cleaved caspase-3 expression compared to the Ang II group (p < 0.001), indicating decreased VSMCs apoptosis. Furthermore, ANXA1 treatment reduced macrophage infiltration (F4/80 expression) and the number of Neutrophil Cytosolic Factor 1 (NCF1)-positive macrophages (p < 0.001). In vitro co-culture of HAVSMCs with M1 macrophages showed that ANXA1 substantially alleviated NCF1-positive cells, mitochondrial dysfunction, and apoptosis (p < 0.001). However, NCF1 upregulation in M1 macrophages effectively counteracted these effects.

Conclusions: In summary, ANXA1 protects against Ang II-induced AAD by improving survival rates, reducing vascular abnormalities, and preserving aortic structural integrity. It also inhibits vascular smooth muscle cell apoptosis, macrophage infiltration, and NCF1 expression. Mechanistically, ANXA1 reduces mitochondrial-dependent smooth muscle cell apoptosis by suppressing M1 macrophage-derived NCF1. These findings suggest ANXA1 has a potential therapeutic agent for AAD by targeting both inflammation and oxidative stress.