Salidroside simultaneously reduces de novo lipogenesis and cholesterol biosynthesis to attenuate atherosclerosis in mice.

Salidroside is a kind of phenylethanoid glycoside and widespread in the plants from Rhodiola and Ligustrum species. Our previous study has reported that salidroside can prevent atherosclerosis progression by ameliorating glyerolipid and glycerophospholipid metabolism in apoE-deficient (apoE-/-) mice. However, its effect on neutral lipids and underlying mechanism remains largely unclear. Here we investigated the molecular mechanism of salidroside action from the perspective of metabolic regulation by integrating metabonomics and transcriptomics pattern. The results showed that salidroside significantly reduced cholesterols, esterified cholesterols, fatty acids, unsaturated fatty acids and triacylclycerols biosynthesis in liver through down-regulating the genes expressions of sterol regulatory element-binding proteins (Srebf1 and Srebf2). The expressions of SREBPs targeted and downstream genes, such as the encoding genes of fatty acid synthase (Fasn), glycerol-3-phosphate acyltransferase (Gpam), stearoyl-CoA desaturase (Scd), 3-hydroxy-3-methylglutaryl-CoA reductase (Hmgcr), and proprotein convertase subtilisin/kexin type 9 (Pcsk9), were also inhibited after salidroside administration. ATP citrate lyase gene (Acly) that encodes an important enzyme producing acetyl-CoA for cholesterol and fatty acid biosynthesis significantly decreased after treatment as well. Moreover, one of ketone body products, 3-hydroxybutyrate, was significantly up-regulated in drug-treated group, indicating that fatty acid degradation was accelerated by salidroside at the same time. Our findings identify salidroside as a regulator of lipid homeostasis in atherosclerotic mice, suggesting its potential to be an alternative medicine for lowering the risks of atherosclerosis-related diseases.

Astragaloside IV Attenuates Myocardial Ischemia-Reperfusion Injury from Oxidative Stress by Regulating Succinate, Lysophospholipid Metabolism, and ROS Scavenging System.

Astragaloside IV is one of the main active ingredients isolated from Astragalus membranaceus. Here we confirmed its protective effect against cardiac ischemia-reperfusion (I/R) injury and aimed to investigate the potential molecular mechanisms involved. Pretreatment of ex vivo and in vivo I/R-induced rat models by astragaloside IV significantly prevented the ratio of myocardium infarct size, systolic and diastolic dysfunction, and the production of creatine kinase and lactate dehydrogenase. Metabolic analyses showed that I/R injury caused a notable reduction of succinate and elevation of lysophospholipids, indicating excessive reactive oxygen species (ROS) generation driven by succinate's rapid reoxidization and glycerophospholipid degradation. Molecular validation mechanistically revealed that astragaloside IV stimulated nuclear factor (erythroid-derived 2)-like 2 (Nrf2) released from Kelch-like ECH-associated protein 1 (Keap1) and translocated to the nucleus to combine with musculoaponeurotic fibrosarcoma (Maf) to initiate the transcription of antioxidative gene heme oxygenase-1 (HO-1), which performed a wide range of ROS scavenging processes against pathological oxidative stress in the hearts. As expected, increasing succinate and decreasing lysophospholipid levels were observed in the astragaloside IV-pretreated group compared with the I/R model group. These results suggested that astragaloside IV ameliorated myocardial I/R injury by modulating succinate and lysophospholipid metabolism and scavenging ROS via the Nrf2 signal pathway.

Cardenolides from the leaves of Nerium oleander.

Six new cardenolides (1-6), including three 14-hydroxylated cardenolides and three 14-carbonylated cardenolides were isolated from the dried aerial parts of Nerium oleander Linn in addition to twenty-seven known compounds (7-33). Their structures were elucidated on the basis of extensive spectroscopic evidences and single-crystal X-ray diffraction analysis. Compounds 1, 4, 7-10 and 13 exhibited significant cytotoxicity against four colon cancer cell lines (HCT116, HT29, SW620, RKO), one gastric cancer cell line (GT) and one cervical cancer cell line (HeLa) in vitro.

Overexpression of mitochondrial ferritin sensitizes cells to oxidative stress via an iron-mediated mechanism.

Mitochondrial ferritin (MtFt) is a newly identified H-ferritin-like protein expressed only in mitochondria. Previous studies have shown that its overexpression markedly affects intracellular iron homeostasis and rescues defects caused by frataxin deficiency. To assess how MtFt exerts its function under oxidative stress conditions, MtFt overexpressing cells were treated with tert-butyl-hydroperoxide (tBHP), and the effects of MtFt expression on cell survival and iron homeostasis were examined. We found that MtFt expression was associated with decreased mitochondrial metabolic activity and reduced glutathione levels as well as a concomitant increase in reactive oxygen species levels and apoptosis. Moreover, mechanistic studies demonstrated that tBHP treatment led to a prolonged decrease in cytosolic ferritins levels in MtFt-expressing cells, while ferritin levels recovered to basal levels in control counterparts. tBHP treatment also resulted in elevated transferrin receptors, followed by more iron acquisition in MtFt expressing cells. The high molecular weight desferrioxamine, targeting to lysosomes, as well as the hydrophobic iron chelator salicylaldehyde isonicotinoyl hydrazone significantly attenuated tBHP-induced cell damage. In conclusion, the current study indicates that both the newly acquired iron from the extracellular environment and internal iron redistribution from ferritin degradation may be responsible for the increased sensitivity to oxidative stress in MtFt-expressing cells.

Determination of reactive oxygen species generated in laccase catalyzed oxidation of wood fibers from Chinese fir (Cunninghamia lanceolata) by electron spin resonance spectrometry.

The aim of the present study was to determine whether the radical reaction intermediates--reactive oxygen species (ROS) were formed during the laccase-catalyzed oxidation of wood fibers from Chinese fir (Cunninghamia lanceolata) and to quantify tentatively its production with electron spin resonance (ESR) spectrometry. To investigate the activation pathways triggered by laccase, ESR spin-trapping techniques using N-tert-butyl-alpha-phenylnitrone (PBN) as spin trap followed by ethyl acetate extraction were employed to identify and quantify the free radical intermediates. ROS such as the superoxide and hydroxyl radical was detected and quantified in the laccase catalyzed oxidation of wood fibers, suggesting that ROS is the main free radical intermediates for laccase reaction. Based on the findings of the presence of ROS and previous literature on the free radical reaction of laccase oxidation of wood fibers, a possible reaction mechanism involving ROS-mediated attack on the domains of lignin which is not directly accessible for the enzyme and solubilized low-molecular mass lignins which function as reactive compounds like adhesives and may cling back to the fiber surface, could accordingly describe laccase-catalyzed oxidation of Chinese fir wood fibers.

Simultaneous detection of NO and ROS by ESR in biological systems.

A large body of evidence shows that the generation of nitric oxide (NO) and reactive oxygen species (ROS) and the rate of ROS/NO play an important role in the biological system. We developed a method to simultaneously detect NO free radical and ROS in biological systems using ERS spin trapping technique. The adduct (DETC)2-Fe2+-NO and N-tert- butyl-alpha-phenylnitrone (PBN)-ROS in biological systems can be extracted by organic solvent and then measured on an electron spin resonance (ESR) spectrometer at room temperature because the g = 2.035 of (DETC)2-Fe2+-NO is different from that of PBN-ROS (g = 2.005) and their ESR signals can be separated clearly. Using this method, we measured the production of NO and ROS in plant and animal systems.

The ESR method to determine nitric oxide in plants.

Nitric oxide (NO) plays an important role not only in animal system but also in plant system. We developed a method to detect NO free radical in plant system using electron spin resonance (ERS) spin trapping technique. The adduct (DETC)2-Fe2+-NO in plant can be extracted by organic solvent and then measured on an ESR spectrometer at room temperature (indirect method) or can be measured in live plant on an ESR spectrometer, but the water in the plant has to be partially dehydrated (direct method).

Protective effects of green tea polyphenols and their major component, (-)-epigallocatechin-3-gallate (EGCG), on 6-hydroxydopamine-induced apoptosis in PC12 cells.

Green tea polyphenols exert a wide range of biochemical and pharmacological effects, and have been shown to possess antimutagenic and anticarcinogenic properties. Oxidative stress is involved in the pathogenesis of Parkinson's disease. However, although green tea polyphenols may be expected to inhibit the progression of Parkinson's disease on the basis of their known antioxidant activity, this has not previously been established. In the present study, we evaluated the neuroprotective effects of green tea polyphenols in the Parkinson's disease pathological cell model. The results show that the natural antioxidants have significant inhibitory effects against apoptosis induced by oxidative stress. 6-Hydroxydopamine (6-OHDA)-induced apoptosis in catecholaminergic PC12 cells was chosen as the in vitro model of Parkinson's disease in our study. Apoptotic characteristics of PC12 cells were assessed by MTT assay, flow cytometry, fluorescence microscopy and DNA fragmentation. Green tea polyphenols and their major component, EGCG at a concentration of 200 microM, exert significant protective effects against 6-OHDA-induced PC12 cell apoptosis. EGCG is more effective than the mixture of green tea polyphenols. The antioxidant function of green tea polyphenols may account for this neuroprotective effect. The present study supports the notion that green tea polyphenols have the potential to be effective as neuropreventive agents for the treatment of neurodegenerative diseases.

Developmental expression and activity variation of nitric oxide synthase in the brain of golden hamster.

Nitric oxide (NO) is the downstream effector after the activation of N-methyl-D-aspartate (NMDA) receptors. It is involved in various physiological processes, such as synapse reconstruction and plasticity, neurotoxity and neuronal death. It also participates in the development and maturation of cortical neurons. The expression of nitric oxide synthase (NOS) during the postnatal development of the visual cortex was investigated by both electron spin resonance (ESR) and Western blot methods. A typical spectrum of (DETC)(2)-Fe(II)-NO complex was found in the visual cortex of different age golden hamsters by ESR method. The signal intensity increased after birth, peaked at postnatal day 14 (PD14) and then gradually decreased. An analysis of variance (ANOVA) implied that the NO synthase expression significantly correlated with the developmental processes (p < 0.05). Results of Western blot further confirmed (one-way ANOVA, p < 0.05) the developmental relating expression pattern of NO synthase shown by ESR technique.

Distinct effects of tea catechins on 6-hydroxydopamine-induced apoptosis in PC12 cells.

Green tea polyphenols have aroused considerable attention in recent years for preventing oxidative stress related diseases including cancer, cardiovascular disease, and degenerative disease. Neurodegenerative diseases are cellular redox status dysfunction related diseases. The present study investigated the different effects of the five main components of green tea polyphenols on 6-hydroxydopamine (6-OHDA)-induced apoptosis in PC12 cells, the in vitro model of Parkinson's disease (PD). When the cells were treated with five catechins respectively for 30 min before exposure to 6-OHDA, (-)-epigallocatechins gallate (EGCG) and (-)-epicatechin gallate (ECG) in 50-200 microM had obvious concentration-dependent protective effects on cell viability, while (-)-epicatechin (EC), (+)-catechin ((+)-C), and (-)-epigallocatechin (EGC) had almost no protective effects. The five catechins also showed the same pattern described above of the different effects against 6-OHDA-induced cell apoptotic characteristics as analyzed by cell viability, fluorescence microscopy, flow cytometry, and DNA fragment electrophoresis methods. The present results indicated that 200 microM EGCG or ECG led to significant inhibition against typical apoptotic characteristics of PC12 cells, while other catechins had little protective effect against 6-OHDA-induced cell death. Therefore, the classified protective effects of the five catechins were in the order ECG> or = EGCG>>EC> or = (+)-C>>EGC. The antiapoptotic activities appear to be structurally related to the 3-gallate group of green tea polyphenols. The present data indicate that EGCG and ECG might be potent neuroprotective agents for PD.