Protective Effect of Spironolactone on Endothelial-to-Mesenchymal Transition in HUVECs via Notch Pathway

Background/Aims: The present study investigated whether the transient receptor potential melastatin 4 (TRPM4) channel plays a role in high salt diet (HSD)-induced endothelial injuries. Methods: Western blotting and immunofluorescence were used to examine TRPM4 expression in the mesenteric endothelium of Dahl salt-sensitive (SS) rats fed a HSD. The MTT, TUNEL, and transwell assays were used to evaluate the cell viability, cell apoptosis, and cell migration, respectively, of human umbilical vein endothelial cells (HUVECs). Enzyme-linked immunosorbent assays were used to determine the concentrations of intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion protein 1 (VCAM-1), and E-selectin. Carboxy-H2DCFDA, a membrane-permeable reactive oxygen species (ROS)-sensitive fluorescent probe, was used to detect intracellular ROS levels. Results: TRPM4 was mainly expressed near the plasma membrane of mesenteric artery endothelial cells, and its expression level increased in SS hypertensive rats fed a HSD. Its protein expression was significantly upregulated upon treatment with exogenous hydrogen peroxide (H₂O₂) and aldosterone in cultured HUVECs. Cell viability decreased upon treatment with both agents in a concentration-dependent manner, which could be partially reversed by 9-phenanthrol, a specific TRPM4 inhibitor. Exogenous H₂O₂ induced apoptosis, enhanced cell migration, and increased the release of adhesion molecules, including ICAM-1, VCAM-1, and E-selectin, all of which were significantly attenuated upon treatment with 9-phenanthrol. Aldosterone and H₂O₂ induced the accumulation of intracellular ROS, which was significantly inhibited by 9-phenanthrol, suggesting that oxidative stress is one of the mechanisms underlying aldosterone-induced endothelial injury. Conclusions: Given the fact that oxidative stress and high levels of circulating aldosterone are present in hypertensive patients, we suggest that the upregulation of TRPM4 in the vascular endothelium may be involved in endothelial injuries caused by these stimuli.

Section: Discussionsupporting

confidence: 57%

Transient Receptor Potential Melastatin 4 (TRPM4) Contributes to High Salt Diet-Mediated Early-Stage Endothelial Injury

Ding

Ban

Liu

et al. 2017

“…EndMT facilitates cell migration and the generation of new tissue types during development and contributes to the pathogenesis of fibrosis [52]. Endothelial cells are normally closely packed with each other, which limits their potential for migration and dissociation from the matrix.…”

Section: Discussionmentioning

confidence: 99%

Low-Intensity Pulsed Ultrasound Prevents the Oxidative Stress Induced Endothelial-Mesenchymal Transition in Human Aortic Endothelial Cells

Zhang

Ren

et al. 2018

Background/Aims: Endothelial-mesenchymal transition (EndMT) has been shown to take part in the generation and progression of diverse diseases, involving a series of changes leading to a loss of their endothelial characteristics and an acquirement of properties typical of mesenchymal cells. Low-intensity pulsed ultrasound (LIPUS) is a new therapeutic option that has been successfully used in fracture healing. However, whether LIPUS can inhibit oxidative stress-induced endothelial cell damages through inhibiting EndMT remained unknown. This study aimed to investigate the protective effects of LIPUS against oxidative stress-induced endothelial cell damages and the underlying mechanisms. Methods: EndMT was induced by H₂O₂ (100 µm for seven days). Human aortic endothelial cells (HAECs) were exposed to H₂O₂ with or without LIPUS treatment for seven days. The expression of EndMT markers (CD31, VE-cadherin, FSP1 and α-SMA) were analyzed. The levels of total and phosphorylated PI3K and AKT proteins were detected by Western Blot analysis. Cell chemotaxis was determined by wound healing and transwell assay. Results: LIPUS relieved EndMT by decreasing ROS accumulation and increasing activation of the PI3K signaling cascade. LIPUS alleviated the migration of EndMT-derived mesenchymal-like cells through reducing extracellular matrix (ECM) deposition that is associated with matrix metallopeptidase (MMP) proteolytic activity and collagen production. Conclusion: LIPUS produces cytoprotective effects against oxidative injuries to endothelial cells through suppressing the oxidative stress-induced EndMT, activating the PI3K/AKT pathway under oxidative stress, and limiting cell migration and excessive ECM deposition.

“…Supernatant protein content was quantified by the Bradford method. Samples were separated on a polyacrylamide gel and then transferred to a nitrocellulose membrane [40,41]. After blockage, the membrane was incubated with primary antibody.…”

Section: Methodsmentioning

confidence: 99%

Early Spironolactone Treatment Attenuates Heart Failure Development by Improving Myocardial Function and Reducing Fibrosis in Spontaneously Hypertensive Rats

Cezar

Damatto

Pagan

et al. 2015

Background: We evaluated the role of the aldosterone blocker spironolactone in attenuating long-term pressure overload-induced cardiac remodeling and heart failure (HF) in spontaneously hypertensive rats (SHR). Methods and Results: Thirteen month-old male SHR were assigned to control (SHR-C, n=20) or spironolactone (SHR-SPR, 20 mg/kg/day, n=24) groups for six months. Normotensive Wistar-Kyoto rats (WKY, n=15) were used as controls. Systolic blood pressure was higher in SHR groups and unchanged by spironolactone. Right ventricular hypertrophy, which characterizes HF in SHR, was less frequent in SHR-SPR than SHR-C. Echocardiographic parameters did not differ between SHR groups. Myocardial function was improved in SHR-SPR compared to SHR-C [developed tension: WKY 4.85±0.68; SHR-C 5.22±1.64; SHR-SPR 6.80±1.49 g/mm²; -dT/dt: WKY 18.0 (16.0-19.0); SHR-C 20.8 (18.4-25.1); SHR-SPR 28.9 (24.2-34.6) g/mm²/s]. Cardiomyocyte cross-sectional area and total collagen concentration (WKY 1.06±0.34; SHR-C 1.85±0.63; SHR-SPR 1.28±0.39 µg/mg wet tissue) were greater in SHR-C than WKY and SHR-SPR. Type 3 collagen expression was lower in SHR-C than WKY and unchanged by spironolactone. Soluble collagen, type I collagen, and lysyl oxidase did not differ between groups. Conclusion: Early spironolactone treatment decreases heart failure development frequency by improving myocardial systolic and diastolic function and attenuating hypertrophy and fibrosis in spontaneously hypertensive rats.