Xiaodi Zhang

Antioxidant activities of oleanolic acid in vitro: possible role of Nrf2 and MAP kinases.

Oleanolic acid (OA) is a natural triterpenoid, which has been used in Chinese medicine for the treatment of liver disorders for many years. Its pharmacological activities have been the focus of intense research in recent years. However, there is little research on the antioxidant activities of OA. In the present study, we aim to investigate whether OA produces its protective effects mainly through antioxidant mechanisms and whether OA plays as an antioxidant through quenching reactive oxygen species (ROS), inhibiting lipid peroxidation or stimulating cellular antioxidant defenses. In the in vitro antioxidant activity-assessing models, OA acted as not only a free radical-scavenger through direct chemical reactions but also a biological molecule, which may enhance the antioxidant defenses. tert-Butyl hydroperoxide (tBHP) induced ROS generation, damaged plasma membrane and decreased cell viability and the expression of key antioxidant enzymes and MAP kinases in QZG cells. OA ameliorated the oxidative injury induced by tBHP through increasing the generation of antioxidant (glutathione) and the expression of key antioxidant enzymes mediated by nuclear factorerythroid 2 p45-related factor 2 (Nrf2), in which process, activation of JNK and ERK, but not p38, was involved. The present study, for the first time, investigated the antioxidant activities of OA systematically. OA probably functions mainly through indirect biological effect and protects QZG cells against cytotoxicity induced by tBHP through increasing the generation of antioxidant and the expression of oxidative stress sensitive transcription factor-Nrf2, and MAP kinases, mainly JNK and ERK. These findings may significantly better the understanding of OA and advance therapeutic approaches to the diseases which are associated with oxidative stress.

Oleanolic acid improves hepatic insulin resistance via antioxidant, hypolipidemic and anti-inflammatory effects

Insulin resistance is the hallmark of type 2 diabetes mellitus (T2DM), which is closely related to disorder of lipid metabolism. The study was designed to evaluate the effects of oleanolic acid (OA) on hepatic insulin resistance and underlying mechanisms in Lep(db)(/)(db) obese diabetic mice. db/db Mice were administered with OA (20mg/kg/day, i.p.) for two weeks. OA reduced body weight, liver weight, and fat weight, and protected liver morphology and function. OA decreased fasting blood glucose, improved glucose and insulin tolerance, enhanced insulin signaling and inhibited gluconeogenesis. In livers, mitochondrial biogenesis, ultrastructure and function were influenced, accompanied by increased cellular and mitochondrial ROS production. OA inhibited all these changes, in which process Nrf2-GCLc mediated stabilization of mitochondrial glutathione pool may be involved. Moreover, OA decreased serum triglyceride, total cholesterol, LDL, HDL, and free fatty acids, increased serum HDL, and reduced hepatic lipid accumulation. Furthermore, inflammatory condition in db/db mice was improved by OA, as evidenced by decreased level of IL-1 β, IL-6, and TNFα in circulation and in liver. The evidence suggests that OA improves hepatic insulin resistance through inhibition of mitochondrial ROS, hypolipidemic and anti-inflammatory effects. The effectiveness of OA leads to interesting therapeutic perspectives.

Glucose oxidase induces insulin resistance via influencing multiple targets in vitro and in vivo: The central role of oxidative stress

It is well known that reactive oxygen species (ROS) plays a role in the pathogenesis of insulin resistance which is the hallmark of type 2 diabetes. However, it is still needed to clarify the mechanism underlying insulin resistance. Glucose oxidase (GOD) is an oxi-reductase catalyzing the conversion of glucose to glucolactone, which is further converted to glucuronic acid and H(2)O(2). The present study was designed to establish a rat model of insulin resistance using GOD and to investigate possible mechanisms. The results showed that three days administration of GOD could significantly increase fasting blood glucose, resulting in impaired glucose and insulin tolerance. Moreover, GOD disrupted insulin signaling both in rats and in hepatocytes, as evidenced by decreased phosphorylation of insulin-stimulated Akt, GSK3 and FOXO1α. Furthermore, GOD administration decreased the expression of PPARγ, alterated the phosphorylation of MAPKs, including p38, ERK and JNK, increased the expression of GRP78 and reduced the expression of PGC-1α and decreased the activities of ATPase and respiratory complexes, all of which have been reported to contribute to insulin resistance. Redox balance was evaluated by detecting the expression of antioxidant defenses and ROS generation. After the treatment with GOD, nuclear factorerythroid 2 p45-related factor 2 (Nrf2)-regulated antioxidant enzymes were damaged and ROS production increased significantly. N-acetyl-L-cysteine (NAC), a potent antioxidant, could notably inhibit these effects of GOD. Although further studies are needed to investigate the clear mechanism, these data also support the conclusion that, if not the most early event, ROS generation is the most important event that plays a central role in the pathogenesis of insulin resistance. Overall, our study established an insulin resistant animal model induced by GOD, elucidated the importance of ROS in pathogenesis of insulin resistance and provided the clue for further studies on the underlying mechanisms.

Enhanced antioxidant effect of caffeic acid phenethyl ester and Trolox in combination against radiation induced-oxidative stress

Combinations of antioxidants are believed to be more effective than single antioxidant because when antioxidants are combined they support each other synergistically to create a magnified effect. Discovering the enhancer effects or synergies between bioactive components is valuable for resisting oxidative stress and improving health benefits. The aim of this study was to investigate a possible cooperation of natural antioxidant caffeic acid phenethyl ester (CAPE) with synthetic antioxidant Trolox in the model systems of chemical generation of free radicals, lipid peroxidation of microsomes and radiation-induced oxidative injury in L929 cells. Based on the intermolecular interaction between CAPE and Trolox, the present study shows a synergistic effect of CAPE and Trolox in combination on elimination of three different free radicals and inhibition of lipid peroxidation initiated by three different systems. CAPE and Trolox added simultaneously to the L929 cells exerted an enhanced preventive effect on the oxidative injury induced by radiation through decreasing ROS generation, protecting plasma membrane and increasing the ratios of reduced glutathione/oxidized glutathione and the expression of key antioxidant enzymes mediated by nuclear factor erythroid 2 p45-related factor 2 (Nrf2). Our results showed for the first time that administration of CAPE and Trolox in combination may exert synergistic antioxidant effects, and further indicate that CAPE and Trolox combination functions mainly through scavenging ROS directly, inhibiting lipid peroxidation and promoting redox cycle of GSH mediated by Nrf2-regulated glutathione peroxidase and glutathione reductase expression.

Mechanism of acute lung injury due to phosgene exposition and its protection by cafeic acid phenethyl ester in the rat.

The mechanism of phosgene-induced acute lung injury (ALI) remains unclear and it is still lack of effective treatments. Previous study indicated that oxidative stress was involved in phosgene-induced ALI. Caffeic acid phenethyl ester (CAPE) has been proved to be an anti-inflammatory agent and a potent free radical scavenger. The purpose of this study was to investigate the protective effects of CAPE on phosgene-induced ALI and identify the mechanism, in which oxidative stress and inflammation were involved. The phosgene was used to induce ALI in rats. The results showed that after phosgene exposure, total protein content in BALF was not significantly changed. The increase of MDA level and SOD activity induced by phosgene was significantly reduced by CAPE administration, and the decrease of GSH level in BALF and lung were significantly reversed by CAPE. CAPE also partially blocked the translocation of NF-κB p65 to the nucleus, but it had little effect on the phosphorylation of p38 MAPK. In conclusion, CAPE showed protective effects on lung against phosgene-induced ALI, which may be related with a combination of the antioxidant and anti-inflammatory functions of CAPE.

Correlation between sPLA2-IIA and phosgene-induced rat acute lung injury.

Secreted phospholipase A2 of group IIA (sPLA2-IIA) has been involved in a variety of inflammatory diseases, including acute lung injury. However, the specific role of sPLA2-IIA in phosgene-induced acute lung injury remains unidentified. The aim of the present study was to investigate the correlation between sPLA2-IIA activity and the severity of phosgene-induced acute lung injury. Adult male rats were randomly exposed to either normal room air (control group) or a concentration of 400 ppm phosgene (phosgene-exposed group) for there are 5 phosgene-exposed groups altogether. For the time points of 1, 3, 6, 12 and 24 h post-exposure, one phosgene-exposed group was sacrificed at each time point. The severity of acute lung injury was assessed by PaO2/FIO2 ratio, wet-to-dry lung-weight ratio, and bronchoalveolar lavage (BAL) fluid protein concentration. sPLA2-IIA activity in BAL fluid markedly increased between 1 h and 12 h after phosgene exposure, and reached its highest level at 6 h. Moreover, the trend of this elevation correlated well with the severity of lung injury. These results indicate that sPLA2-IIA probably participates in phosgene-induced acute lung injury.

Apoptosis of ATII cells in mice induced by phosgene

Phosgene inhalation can induced pulmonary edema formation. The purpose of this study was to investigate cell of apoptosis in pulmonary edema mice induced by phosgene. Fifty-two BALB/c mice were random divided into a negative group and a positive group with 26 mice in each. Mice were exposed for 5 min to air and phosgene in the negative group and in the positive one, respectively. The dose of phosgene was 539 ppm. After 4 h of exposure, all mice were anesthetized. Lungs were analyzed for lung wet/dry weight ratio and pathological alternation. The method of isolation and culture of alveolar type II cells (ATII cells) was established to observe their apoptosis by electron microscope and flow cytometry. Apoptosis of lung cells was observed by DNA gel electrophoresis and TUNEL. The lung wet/dry weight ratio was significantly higher in the positive group (6.42 ± 1.00) than in the negative group (4.25 ± 0.47, p < 0.05). A large amount of fluid effusion was observed in the alveolus of mice induced by phosgene. Alveolar type II cells were identified by tannic acid stainning and electron microscope. The apoptotic signs in alveolar type II cells, alveolar type I cells, eosinophils, macrophages, symphocytes, and ciliated cells were viewed under electron microscope in positive group. The ratio of apoptosis cells (40.26 ± 7.74) in positive was higher than that (1.58 ± 1.01, p < 0.001) in the negative group by flow cytometry. DNA ladder alternation was seen through DNA gel electrophoresis. Apoptosis of epithelia and vascular endothelia in lung were found by TUNEL. These results indicate that there is success in establishing a model of pulmonary edema and method of isolation and culture of AT II cells in BALB/c mice. Phosgene can induce apoptosis of cells in the lungs of BALB/c mice, and this indicates that pulmonary edema is related to apoptosis of lung cells in mice, induced by phosgene.

The effects of Insulin Pre-Administration in Mice Exposed to Ethanol: Alleviating Hepatic Oxidative Injury through Anti-Oxidative, Anti-Apoptotic Activities and Deteriorating Hepatic Steatosis through SRBEP-1c Activation

Alcoholic liver disease (ALD) has become an important liver disease hazard to public and personal health. Oxidative stress is believed to be responsible for the pathological changes in ALD. Previous studies have showed that insulin, a classic regulator of glucose metabolism, has significant anti-oxidative function and plays an important role in maintaining the redox balance. For addressing the effects and mechanisms of insulin pre-administration on ethanol-induced liver oxidative injury, we investigated histopathology, inflammatory factors, apoptosis, mitochondrial dysfunction, oxidative stress, antioxidant defense system, ethanol metabolic enzymes and lipid disorder in liver of ethanol-exposed mice pretreatment with insulin or not. There are several novel findings in our study. First, we found insulin pre-administration alleviated acute ethanol exposure-induced liver injury and inflammation reflected by the decrease of serum AST and ALT activities, the improvement of pathological alteration and the inhibition of TNF-α and IL-6 expressions. Second, insulin pre-administration could significantly reduce apoptosis and ameliorate mitochondrial dysfunction in liver of mice exposed to ethanol, supporting by decreasing caspases-3 activities and the ratio of Bax/Bcl-2, increasing mitochondrial viability and mitochondrial oxygen consumption, inhibition of the decline of ATP levels and mitochondrial ROS accumulation. Third, insulin pre-administration prevented ethanol-mediated oxidative stress and enhance antioxidant defense system, which is evaluated by the decline of MDA levels and the rise of GSH/GSSG, the up-regulations of antioxidant enzymes CAT, SOD, GR through Nrf-2 dependent pathway. Forth, the modification of ethanol metabolism pathway such as the inhibition of CYP2E1, the activation of ALDH might be involved in the anti-oxidative and protective effects exerted by insulin pre-administration against acute ethanol exposure in mice. Finally, insulin pre-administration deteriorated hepatic steatosis in mice exposed to ethanol might be through SRBEP-1c activation. In summary, these results indicated that insulin pre-administration effectively alleviated liver oxidative injury through anti-inflammatory, anti-oxidative and anti-apoptotic activities but also deteriorated hepatic steatosis through SRBEP-1c activation in mice exposed to ethanol. Our study provided novel insight about the effects and mechanisms of insulin on ethanol-induced liver injury.

Pentoxifylline inhibits intercellular adhesion molecule-1 (ICAM-1) and lung injury in experimental phosgene-exposure rats

Phosgene inhalation results in acute lung injury (ALI) mostly, pulmonary edema and even acute respiratory distress syndrome, but there is no specific antidote. Inflammatory cells play an important role in the ALI caused by phosgene. Intercellular adhesion molecule-1 (ICAM-1) is a critical factor for inflammatory organ injury. We hypothesized that pentoxifylline (PTX), an inhibitor of leukocyte activation, would have a protective effect on experimental phosgene-induced lung injury rats by inhibiting ICAM-1. To prove this hypothesis, we used rat models of phosgene (400 ppm × 1 min)-induced injury to investigate: (1) the time course of lung injury (control 1, 3, 6, 12, 24, and 48 h group), including pathological changes in hematoxylin and eosin staining and transmission electron microscope, myeloperoxidase (MPO) activity by colorimetric method and ICAM-1 protein level detected by western blot, (2) At 3 h after phosgene exposure, protective effects of different dosages of PTX (50 mg/kg and 100 mg/kg) administration were evaluated by MPO activity, ICAM-1 differential expression and WBC count in bronchoalveolar lavage fluid. The results showed that inflammatory cells emerged out of lung blood vessels at 3 h after phosgene exposure. The MPO activity of lung tissue increased significantly from 3 to 48 h after phosgene exposure (P < 0.05) and ICAM-1 expression presented a similar change, especially at 3 h and 24 h (P < 0.05). After pretreatment and treatment with PTX (100 mg/kg), significant protective effects were shown (P < 0.05). These data supported our hypothesis that PTX reduced phosgene-induced lung injury, possibly by inhibiting ICAM-1 differential expression.

Suppression of nuclear factor erythroid 2-related factor 2 via extracellular signal-regulated kinase contributes to bleomycin-induced oxidative stress and fibrogenesis

Pulmonary fibrosis is a serious and irreversible lung injury with obscure etiologic mechanisms and no effective treatment to date. This study explored a crucial link between oxidative stress and pulmonary fibrogenesis, focusing on nuclear factor erythroid 2-related factor 2 (Nrf2), a core transcription factor in antioxidative regulation systems. Treatment of C57 BL/6 mice with bleomycin increased fibroblast viability and collagen production and significantly downregulated Nrf2. In addition, prominent oxidative stress was indicated by changes in superoxide dismutase, catalase activity, and glutathione and thiobarbituric acid-reactive substance levels. In a cell-based model, bleomycin suppressed Nrf2 activation via extracellular signal-related kinase phosphorylation, enhancing intracellular reactive oxygen species in lung fibroblasts and stimulating abnormal cell proliferation and collagen secretion. To confirm this novel mechanism of bleomycin-induced fibrogenesis, we attempted to upregulate Nrf2 and related antioxidant proteins in bleomycin-treated fibroblasts using a putative Nrf2 activator, caffeic acid phenethyl ester, and the results showed that bleomycin-induced fibroblast proliferation and collagen content were attenuated through improved redox balance. Collectively, these results disclose a potential regulatory mechanism in pulmonary fibrosis that will aid the development of new therapies.