Atorvastatin and pravastatin stimulate nitric oxide and reactive oxygen species generation, affect mitochondrial network architecture and elevate nicotinamide N‐methyltransferase level in endothelial cells

Abstract

Statins belong to the most often prescribed medications, which efficiently normalise hyperlipidaemia and prevent cardiovascular complications in obese and diabetic patients. However, beside expected therapeutic results based on the inhibition of 3‐hydroxyl‐3‐methylglutaryl‐CoA reductase, these drugs exert multiple side effects of poorly understood characteristic. In this study, side effects of pravastatin and atorvastatin on EA.hy926 endothelial cell line were investigated. It was found that both statins activate proinflammatory response, elevate nitric oxide and reactive oxygen species (ROS) generation and stimulate antioxidative response in these cells. Moreover, only slight stimulation of the mitochondrial biogenesis and significant changes in the mitochondrial network organisation have been noted. Although biochemical bases behind these effects are not clear, they may partially be explained as an elevation of AMP‐activated protein kinase (AMPK) activity and an increased activating phosphorylation of sirtuin 1 (Sirt1), which were observed in statins‐treated cells. In addition, both statins increased nicotinamide N‐methyltransferase (NNMT) protein level that may explain a reduced fraction of methylated histone H3. Interestingly, a substantial reduction of the total level of histone H3 in cells treated with pravastatin but not atorvastatin was also observed. These results indicate a potential additional biochemical target for statins related to reduced histone H3 methylation due to increased NNMT protein level. Thus, NNMT may directly modify gene activity.

Coronary microvascular injury in myocardial infarction: perception and knowledge for mitochondrial quality control

“…For example, exercising rats following MI was shown to overtly promote EC proliferation, VEGF level, angiogenesis, eNOS activity, and endothelium-dependent vasodilation 165 . In addition to exercise, mitochondrial biogenesis in ECs may be stimulated by various medications including statins 166 , resveratrol 167 , liraglutide 168 , cannabidiol 169 , salidroside 170 , and red wine polyphenols 171 . Nonetheless, only metformin 172 and resveratrol 173 augment mitochondrial biogenesis in ECs, improved cardiac function and decreased infarct size in MI or myocardial I/R injury models.…”

Effect of lipopolysaccharide (LPS) on HAEC cells. Does nicotinamide N-methyltransferase sensitize HAEC cells to LPS?

“…Keeping in mind earlier findings published by other authors [Akar et al, 2020] and results obtained in our laboratory [Dymkowska et al, 2021] indicating that some pathological stimuli may elevate NNMT content in a variety of cells and that activity of this enzyme may be related to cellular energy metabolism [Mistry et al, 2020] we tested the putative effect of LPS on NNMT level in HAECs (Fig. 8).…”

“…However, the physiological role of this enzyme in vascular endothelial cells is still obscure. Experimental data concerning NNMT clearly show that cells challenged by some proinflammatory stimuli exhibit an elevated level of this protein [Dymkowska et al, 2021]. This may suggest that NNMT may play a role in the adaptive stress response of cells.…”

“…Measurement of cytosolic ROS level. Cells were loaded with a 5 µM DHE probe (dihydroethidium, Molecular Probes) as described previously [Dymkowska et al, 2021]. The fluorescence was measured at the excitation and emission wavelengths of 535 nm and 610 nm, respectively.…”

“…For visualization of the mitochondrial network architecture, confluent cells grown on the collagen-coated coverslips (ϕ12 mm) were loaded with 100 nM MitoTracker Red CMXRos (Molecular Probes) as previously described [Dymkowska et al, 2021]. After gentle rinsing, the cells were fixed with 4% paraformaldehyde, rinsed with PBS supplemented with 5% BSA, permeabilized with 0.1% Triton X-100 in 5% BSA and rinsed overnight with 1% BSA containing PBS at 4 • C. Finally cells were stained with Actin-Stain 488 Phalloidin (1:1000; Cytoskeleton) and with 2 µg/ml Hoechst 33342 (ThermoFisher Scientific) Finally, cells were sealed in VECTASHIELD Mounting Medium (VECTOR Laboratories).…”