Catalpol inhibits HHcy-induced EndMT in endothelial cells by modulating ROS/NF-κB signaling

Abstract

AbstractBackgroundHyperhomocysteinemia (HHcy) is an independent risk factor for atherosclerosis (AS), the molecular mechanisms of its pathogenesis are not fully understood. Endothelial dysfunction is the key initiating link in AS. However, whether endothelial-mesenchymal transition (EndMT) is involved in the regulation of HHcy-induced endothelial dysfunction and the role of catalpol in it remain unexplored.Methods and ResultsIn vitroHHcy-treated primary human umbilical vein endothelial cells (HUVECs) were used to construct a model of endothelial dysfunction, and the antioxidants N-acetylcysteine (NAC) and catalase alcohol were administered. The endothelial dysfunction model was constructed by observing cell morphological changes, measuring intracellular reactive oxygen species (ROS) content, quantifying EndMT marker proteins (VE-cadherin, α-SMA, N-cadherin) and p-p65 and p65 protein expression by western blot, and detecting VE-cadherin and α-SMA fluorescence intensity by immunofluorescence.In vivoC57BL/6N mice were given a diet fed with 4.4% high methionine chow to construct a HHcy mice model and were treated with catalpol. The model was further validated by measuring the concentration of homocysteinemia (Hcy) in mice, HE and Masson staining to observe the pathological changes in the endothelium of aortic roots, immunohistochemistry to detect the expression of EndMT marker protein, immunofluorescence to detect the fluorescence intensity of p-p65, and fluorescent probe method to detect the content of ROS in the endothelium of aorta. The results showed that catalpol significantly inhibited HHcy-induced endothelial cell morphological transformation, reduced HHcy-induced increase in intracellular ROS content and α-SMA, N-cadherin, p-p65 protein expression, increased HHcy-induced decrease in VE-cadherin, CD31 protein expression, and was able to protect against endothelial pathological changes in the aortic root and reduce aortic endothelial ROS content.ConclusionsCatalpol inhibits HHcy-induced EndMT, and the underlying mechanism may be related to the ROS/NF-κB signaling pathway. Catalpol may be a potential drug for the treatment of HHcy-related AS.