Aqueous extract of Whitmania pigra Whitman ameliorates ferric chloride-induced venous thrombosis in rats via antioxidation.

Whitmania pigra Whitman (W. pigra) has been widely employed in decoction for the treatment of blood stasis syndrome for many years in China. The aim of the present study was to explore the anti-venous thrombosis (VT) mechanism of the aqueous extract of W. pigra (AEW) in rats. Rats were orally administered with AEW. A inferior vena cava (IVC) thrombosis model was established. Thrombosed IVC was weighed and histopathological analyses were performed. Blood coagulation, blood fibrinolysis, blood cell count, blood viscosity and platelet activity were evaluated. Reactive oxygen species (ROS) accumulation was analyzed. Malondialdehyde (MDA) content in thrombosed IVC and antioxidants in serum were detected. Protein expression of nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) in thrombosed IVC was determined. AEW significantly reduced thrombus weight. It did not affect blood coagulation, blood fibrinolysis, blood cell count, platelet activity, or whole blood viscosity. However, AEW remarkably alleviated vascular injury, reduced ROS accumulation and MDA content, enhanced the total antioxidant capacity and the activities of superoxide dismutase, glutathione peroxidase and glutathione reductase. It increased the glutathione/oxidized glutathione ratio and the protein expression levels of Nrf2 and HO-1. In summary, W. pigra may prevent VT via Nrf2-mediated antioxidation.

Protective Role of Endothelial SIRT1 in Deep Vein Thrombosis and Hypoxia-induced Endothelial Dysfunction Mediated by NF-κB Deacetylation.

Venous hypoxia is considered as the major pathogenetic mechanism linking blood flow stagnancy with deep vein thrombosis (DVT). Our previous study showed that activating SIRT1 may attenuate inferior vena cava (IVC) stenosis-induced DVT in rats. This study was aimed to investigate the role of endothelial SIRT1 in DVT and hypoxia-induced endothelial dysfunction as well as the underlying mechanism. Protein profiling of IVCs and blood plasma of DVT rats induced by IVC stenosis was analysed by 4D Label free proteomics analysis. To verify the independent role of SIRT1 in DVT and oxygen-glucose deprivation (OGD)-induced endothelial dysfunction, SIRT1 specific activator SRT1720 and SIRT1 knockdown in both local IVCs and endothelial cells were employed. Moreover, the role of the NF-κB were investigated using NF-κB inhibitor caffeic acid phenethyl ester (CAPE). SRT1720 significantly inhibited thrombus burden, leukocytes infiltration, protein expressions of cell adhesion molecules and chemokines, as well as acetylation level of NF-κB/p65 in wild DVT rats, while these protective effects of SRT1720 were abolished in rats with SIRT1 knockdown in local IVCs. In vitro, SRT1720 protected endothelial cells against OGD-induced dysfunction characterized with enhanced adhesion of monocytes as well as the protein expressions of cell adhesion molecules and chemokines, whereas these protective effects of SRT1720 were vanished by SIRT1 stable knockdown. Furthermore, CAPE attenuated endothelial cell dysfunction and abolished these effects of SIRT1 knockdown. Collectively, these data suggested that endothelial SIRT1 plays an independent role in ameliorating hypoxia-induced endothelial dysfunction and thrombotic inflammation in DVT, and this effect is mediated by NF-κB deacetylation.

Temporary Upregulation of Nrf2 by Naringenin Alleviates Oxidative Damage in the Retina and ARPE-19 Cells.

Dry age-related macular degeneration (dAMD) is a chronic degenerative ophthalmopathy that leads to serious burden of visual impairment. Antioxidation in retinal pigment epithelium (RPE) cells is considered as a potential treatment for dAMD. Our previous studies have showed that naringenin (NAR) protects RPE cells from oxidative damage partly through SIRT1-mediated antioxidation. In this study, we tested the hypothesis that the Nrf2 signaling is another protective mechanism of NAR on dAMD. NaIO3-induced mouse retinopathy and ARPE-19 cell injury models were established. Immunochemical staining, immunofluorescence, and western blotting were performed to detect the protein expressions of Nrf2 and HO-1. In addition, ML385 (activity inhibitor of Nrf2) and zinc protoporphyrin (ZnPP, activity inhibitor of HO-1) were applied to explore the effect of NaIO3 or NAR. The results showed that NAR increased the protein expressions of Nrf2 and HO-1 in the retinas in mice exposed to NaIO3 at the early stage. NAR treatment also resulted in a stronger activation of Nrf2 at the early stage in NaIO3-treated ARPE-19 cells. Moreover, inhibition of HO-1 by ZnPP weakened the cytoprotective effect of NAR. The constitutive accumulation and activation of Nrf2 induced by NaIO3 led to the death of RPE cells. However, NAR decreased the protein expressions of Nrf2 and HO-1 towards normal level in the mouse retinas and ARPE-19 cells exposed to NaIO3 at the late stage. Our findings indicate that NAR protects RPE cells from oxidative damage via activating the Nrf2 signaling pathway.

Aqueous extract of Paeoniae Radix Rubra prevents deep vein thrombosis by ameliorating inflammation through inhibiting GSK3ß activity.

BACKGROUND: Deep vein thrombosis (DVT) is a kind of blood stasis syndrome. Paeoniae Radix Rubra (PRR) has long been widely used for eliminating blood stasis in China, but its effect on DVT has not yet been reported. PURPOSE: The present study aimed to assess the potential inhibitory effect of the aqueous extract of PRR (i.e.,PRR dispensing granule, PRRDG) on DVT and explore the underlying mechanism. STUDY DESIGN/METHODS: The chemical profile of PRRDG was analyzed by high-performance liquid chromatography. Sprague-Dawley rats were intragastrically treated with PRRDG (0.625, 1.25 and 1.875 g crude drug/kg/d) once daily for 7 consecutive days. On the sixth day, a model of inferior vena cava (IVC) stenosis-induced DVT was established. All rats were sacrificed on the seventh day. Serum was collected for enzyme-linked immunosorbent assay. Thrombus-containing IVC was weighed and further processed for histopathologic examination, immunohistochemical analysis and western blotting. LiCl and LY294002 were adopted to block and increase the activity of glycogen synthase kinase 3ß (GSK3ß), respectively. RESULTS: The chemical profile analysis showed that paeoniflorin, benzoylpaeoniflorin, albiflorin, gallic acid and catechin were the main constituents of PRRDG. LiCl decreased thrombus weight, reduced the number of inflammatory cells in thrombus and vein wall, down-regulated phosphorylated NF-κB p65 (p-p65) protein expression. Similarly, PRRDG decreased thrombus weight and tissue factor (TF) protein expression. PRRDG reduced the protein expression levels of P-selectin, monocyte chemoattractant protein-1 (MCP-1), intercellular cell adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in venous endothelium, serum levels of tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß), and the number of inflammatory cells in thrombus and vein wall. Moreover, PRRDG down-regulated p-p65 protein expression and up-regulated phosphorylated GSK3ß (p-GSK3ß) protein expression. LY294002 abrogated the inhibitory effects of PRRDG on thrombus weight, TF protein expression, TNF-α and IL-1ß serum levels, inflammatory cells influxes, and p-p65 protein expression. CONCLUSION: PRRDG prevents DVT by ameliorating inflammation through inhibiting GSK3ß activity.