SUMMARYObjective: MicroRNAs (miRNAs) are noncoding small RNAs that control gene expression at the posttranscriptional level. Some dysregulated miRNAs have been shown to play important roles in epileptogenesis. The aim of this study was to determine if miR199a-5p regulates seizures and seizure damage by targeting the antiapoptotic protein silent information regulator 1 (SIRT1). Methods: Hippocampal expression levels of miR-199a-5p, SIRT1, and acetylated p53 were quantified by quantitative real-time polymerase chain reaction (RT-PCR) and Western blotting in the acute, latent, and chronic stages of epilepsy in a rat lithiumpilocarpine epilepsy model. Silencing of miR-199a-5p expression in vivo was achieved by intracerebroventricular injection of antagomirs. The effects of targeting miR-199a-5p and SIRT1 protein on seizure and epileptic damage post-status epilepticus were assessed by electroencephalography (EEG) and immunohistochemistry, respectively. Results: miR-199a-5p expression was up-regulated, SIRT1 levels were decreased, and neuron loss and apoptosis were induced in epilepsy model rats compared with normal controls, as determined by up-regulation of acetylated p53 and cleaved caspase-3 expression. In vivo knockdown of miR-199a-5p by an antagomir alleviated the seizurelike EEG findings and protected against neuron damage, in accordance with up-regulation of SIRT1 and subsequent deacetylation of p53. Furthermore, the seizure-suppressing effect of the antagomir was partly SIRT1 dependent. Significance: The results of this study suggest that silencing of miR-199a-5p exerts a seizure-suppressing effect in rats, and that SIRT1 is a direct target of miR-199a-5p in the hippocampus. The effect of miR-199a-5p on seizures and seizure damage is mediated via down-regulation of SIRT1. The miR-199a-5p/SIRT1 pathway may thus represent a potential target for the prevention and treatment of epilepsy and epileptic damage.

Catalpol, an iridoid glucoside, has been found present in large quantities in the root of Rehmannia glutinosa L. and showed a strong antioxidant capacity in the previous study. In the present work, the protective effect of catalpol against AS via inhibiting oxidative stress, DNA damage, and telomere shortening was found in LDLr−/− mice. This study also shows that activation of the peroxisome proliferator-activated receptor-γ coactivator-1α (PGC-1α)/telomerase reverse transcriptase (TERT) pathway, which is the new link between mitochondria and telomere, was involved in the protective effects of catalpol. Further, by using PGC-1α or TERT siRNA in oxLDL-treated macrophages, it is proved that catalpol reduced oxidative stress, telomere function, and related DNA damage at least partly through activating the PGC-1α/TERT pathway. Moreover, dual luciferase activity assay-validated catalpol directly enhanced PGC-1α promoter activity. In conclusion, our study revealed that the PGC-1α/TERT pathway might be a possible therapeutic target in AS and catalpol has highly favorable characteristics for the treatment of AS via modulating this pathway.

Hepatic steatosis and oxidative stress are considered to be the sequential steps in the development of nonalcoholic fatty liver disease (NAFLD). We previously found that catalpol, an iridoid glucoside extracted from the root of Romania glutinosa L, protected against diabetes-induced hepatic oxidative stress. Here, we found that the increased expression of p66shc was observed in NAFLD models and catalpol could inhibit p66shc expression to ameliorate NAFLD effectively. However, the underlying mechanisms remained unknown. The aim of the present study was to investigate the p66shc-targeting miRNAs in regulating oxidative stress and hepatic steatosis, also the mechanisms of catalpol inhibiting NAFLD. We found that the effects of catalpol inhibiting hepatic oxidative stress and steasis are dependent on inhibiting P66Shc expression. In addition, miR-96-5p was able to suppress p66shc/cytochrome C cascade via targeting p66shc mRNA 3'UTR, and catalpol could lead to suppression of NAFLD via upregulating miR-96-5p level. Thus, catalpol was effective in ameliorating NAFLD, and miR-96-5p/p66shc/cytochrome C cascade might be a potential target.

Oxidative stress and autophagy dysfunction are critical factors in the pathogenesis of atherosclerosis. Adenosine 50 monophosphate-activated protein kinase (AMPK) plays an important role in inhibiting oxidative stress and promoting autophagy. Catalpol, an iridoid glucoside extracted from the root of Rehmanniae glutinosa L, was reported to produce a potent antioxidant effect. However, the mechanism of catalpol inhibiting oxidative stress and its effect on AMPK remain unclear. This study aims to investigate the potential role of catalpol in regulating oxidative stress and autophagy in tumor necrosis factor-a (TNF-a)-treated human aorta epithelial cells (HAECs), and the important role of AMPK involved in catalpol's effects. In the present study, effects of catalpol on inhibiting oxidative stress and the related adhesion, apoptosis and promotion of autophagy were demonstrated. Catalpol also produces a potent effect on activating AMPK in TNF-a-treated HAECs. Mechanistically, the effects of catalpol on inhibiting oxidative stress and increasing the autophagy level were partly blocked by a pharmacological AMPK inhibitor, compound C or AMPK small interfering RNA, indicating that catalpol performed such protective effects by activating AMPK. In summary, this study demonstrated that AMPK is a key treatment target for endothelial cell injury and catalpol confers protection on TNF-a-treated HAECs by up-regulating AMPK activity. Catalpol has highly favorable characteristics for the treatment of atherosclerosis.

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