Pyridostigmine attenuated high-fat-diet induced liver injury by the reduction of mitochondrial damage and oxidative stress via α7nAChR and M3AChR.

Obesity is a major cause of nonalcohol fatty liver disease (NAFLD), which is characterized by hepatic fibrosis, lipotoxicity, inflammation, and apoptosis. Previous studies have shown that an imbalance in the autonomic nervous system is closely related to the pathogenesis of NAFLD. In this study, we investigated the effects of pyridostigmine (PYR), a cholinesterase (AChE) inhibitor, on HFD-induced liver injury and explored the potential mechanisms involving mitochondrial damage and oxidative stress. A murine model of HFD-induced obesity was established using the C57BL/6 mice, and PYR (3 mg/kg/d) or placebo was administered for 20 weeks. PYR reduced the body weight and liver weight of the HFD-fed mice. Additionally, the serum levels of IL-6, TNF-α, cholesterol, and triglyceride were significantly lower in the PYR-treated versus the untreated mice, corresponding to a decrease in hepatic fibrosis, lipid accumulation, and apoptosis in the former. Furthermore, the mitochondrial morphology improved significantly in the PYR-treated group. Consistently, PYR upregulated ATP production and the mRNA level of the mitochondrial dynamic factors OPA1, Drp1 and Fis1, and the mitochondrial unfolded protein response (UPRmt) factors LONP1 and HSP60. Moreover, PYR treatment activated the Keap1/Nrf2 pathway and upregulated HO-1 and NQO-1, which mitigated oxidative injury as indicated by decreased 8-OHDG, MDA and H2 O2 levels, and increased SOD activity. Finally, PYR elevated acetylcholine (ACh) levels by inhibiting AChE, and upregulated the α7nAChR and M3AChR proteins in the HFD-fed mice. PYR alleviated obesity-induced hepatic injury in mice by mitigating mitochondrial damage and oxidative stress via α7nAChR and M3AChR.

Rhaponticum carthamoides improved energy metabolism and oxidative stress through the SIRT6/Nrf2 pathway to ameliorate myocardial injury.

BACKGROUND: Rhaponticum carthamoides (Willd.) Iljin (Rha) is a member of the family Compositae that is widely used in folk medicine as a dietary supplement to treat cardiovascular diseases (CVDs), such as senile cardiac insufficiency, and to restore myocardial function after surgery. Sirtuin 6 (SIRT6), an NAD+-dependent class III histone deacetylase, plays a considerable role in the administration of CVDs. However, the specific effects and mechanism of Rha on myocardial injury remain unknown. PURPOSE: This study aimed to explore the therapeutic potential of Rha against myocardial injury as well as its underlying mechanisms in vivo and in vitro. METHODS: A myocardial ischaemia model was established in male SD rats by subcutaneously injecting ISO. The rats were gavaged with Rha (40, 80, 160 mg/kg) or Rho (6 ml/kg) for 14 successive days and then injected subcutaneously with ISO or saline solution on the 13th and 14th days. The positive effects of Rha against myocardial injury in rats were evaluated by ECG assessment, BP measurements, H&E staining, and myocardial enzyme detection. Biochemical indicators of energy metabolism and oxidative stress, such as NAD+/NADH, ATP, and MDA, were analysed by assay kits to assess the effects of Rha. The protein and mRNA expression levels of SIRT6 and Nrf2 in the myocardium were determined by western blotting and real-time PCR. RESULTS: Our results showed that Rha ameliorated myocardial ischaemia and inhibited energy metabolism disorders (NAD+/NADH ratio, ATP, and LD) and oxidative stress (SOD, ROS, etc.) in rat myocardial tissue and H9c2 cells. In addition, Rha upregulated SIRT6 and Nrf2 expression in myocardial injury. Mechanistic studies then found that SIRT6 knockdown reduced the expression of Nrf2 as well as the effects of Rha on the levels of ATP, LD, and ROS, whereas activation of Nrf2 improved the effects of Rha in cells. In summary, Rha might exert its cardioprotective effects via the SIRT6-mediated Nrf2 signaling pathway. CONCLUSION: The results suggest that Rha regulates energy metabolism and oxidative stress through the SIRT6/Nrf2 signaling pathway to play a protective role in myocardial injury.

MiR-145 alleviates Hcy-induced VSMC proliferation, migration, and phenotypic switch through repression of the PI3K/Akt/mTOR pathway.

The proliferation, migration, and cellular morphology of vascular smooth muscle cells (VSMCs) play important roles in the pathogenesis of atherosclerosis (AS). Homocysteine (Hcy) is a sulfur-containing amino acid, which is an intermediate product of methionine metabolism. Hcy can induce proliferation, migration, and phenotypic switch of VSMCs, but details of these mechanisms are still unclear. The phosphatidylinositol 3-kinase (PI3K/Akt/mTOR) signaling pathway is involved in a host of cellular functions. In this study, we sought to determine if this multifunctional pathway played a role in Hcy-induced proliferation, migration, and phenotypic transformation of VSMCs, which has not been previously reported. miR-145 has been previously reported to suppress the effects of Hcy in VSMCs. In our study, using qRT-PCR, we found that Hcy itself reduced the expression of miR-145 in VSMCs, while overexpression of miR-145 reduced the proliferation, migration, and phenotypic transformation of VSMCs caused by Hcy. Using Western blot analysis, we found that VSMCs exposed to Hcy exhibited significant increases in the levels of PI3K, Akt, and mTOR proteins. Additionally, overexpression of miR-145 dramatically decreased PI3K, Akt, and mTOR expression. Using qRT-PCR we found that miR-145 expression increased after blocking PI3K using an inhibitor. Inhibition of the PI3K signaling pathway also prevented Hcy-induced VSMC proliferation, migration, and phenotypic switch. Taken together, our results suggest that miR-145 could inhibit VSMC proliferation, migration, and phenotype switching by preventing activation of the PI3K/Akt/mTOR signaling pathway.

Sitagliptin inhibits vascular inflammation via the SIRT6-dependent signaling pathway.

Sitagliptin has recently been shown to inhibit inflammatory response in cardiovascular disease. Sirtuin6 (SIRT6), a NAD+-dependent class III histone deacetylase, participates in the regulation of cellular inflammation. We hypothesized that sitagliptin could attenuate vascular inflammation via modulation of SIRT6 pathway. It was found that sitagliptin decreased the expression of monocyte chemotactic protein-1 (MCP-1), interleukin (IL)-6 and IL-1ß, but up-regulated SIRT6 expression, both in mice and in TNF-α-stimulated endothelial cells. Moreover, knockdown of SIRT6 reversed the inhibitory effect of sitagliptin on MCP-1, IL-6 and IL-1ß expression. Further study revealed that sitagliptin also decreased the expression of MCP-1, IL-6 and IL-1ß partly through suppression of reactive oxygen species (ROS). In vivo, hypercholesterolemia in mice was induced by intraperitoneal administration of poloxamer 407 for 1 month. Hyperlipidemia-induced production of MCP-1, IL-6 and IL-1ß was significantly suppressed in the sitagliptin-supplemented animals, but the effect of was abolished by SIRT6 knockout in endothelium. These results indicate that sitagliptin protects against vascular inflammation via the SIRT6/ROS-dependent signaling pathway.

The protective effect of hydroxytyrosol acetate against inflammation of vascular endothelial cells partly through the SIRT6-mediated PKM2 signaling pathway.

Hydroxytyrosol acetate (HT-AC), a polyphenolic compound in olive oil, exerts an anti-inflammatory effect on murine collagen-induced arthritis. However, the effect of HT-AC on inflammatory response in cardiovascular disease remains unclear. Thus, in this study, we aimed to investigate the effect of HT-AC on the inflammation response of vascular endothelial cells and the related molecular mechanism. Our results showed that HT-AC inhibited the inflammatory response in hypercholesterolemic mice and tumor necrosis factor (TNF)-stimulated HUVECs. Meanwhile, HT-AC also up-regulated SIRT6 expression in hypercholesterolemic mice and HUVECs. To further investigate whether SIRT6 is involved in the regulation of endothelial inflammatory response by HT-AC, endothelium-specific Sirt6 knockout (Sirt6endo-/-) mice were used. Our study found that Sirt6endo-/- abolished the inhibition of inflammatory response by HT-AC in the thoracic aorta of hypercholesterolemic mice. In vitro study also showed that knockdown of SIRT6 reduced the inhibition of inflammatory response by HT-AC, whereas overexpression of SIRT6 augmented the inhibition of inflammatory response by HT-AC in HUVECs. Further study demonstrated that HT-AC exerts its anti-inflammatory effect partly via the SIRT6-mediated PKM2 signaling pathway. In addition, HT-AC inhibited TNF-induced inflammatory response through the TNF receptor superfamily member 1A (TNFRSF1A) signaling pathway. These findings indicate that HT-AC regulates the vascular endothelial inflammatory response partly through the TNFRSF1A/SIRT6/PKM2-mediated signaling pathway.