Kolaviron protects against nigrostriatal degeneration and gut oxidative damage in a stereotaxic rotenone model of Parkinson's disease.

The asymptomatic and clinical stages of Parkinson's disease (PD) are associated with comorbid non-motor symptoms including gastrointestinal (GI) dysfunction. Although the neuroprotective and gastroprotective roles of kolaviron (KV) have been reported independently, whether KV-mediated GI-protective capacity could be beneficial in PD is unknown. We therefore investigated the modulatory effects of KV on the loss of dopaminergic neurons, locomotor abnormalities, and ileal oxidative damage when rats are lesioned in the nigrostriatal pathway. KV treatment markedly suppressed the behavioral deficit and apomorphine-induced rotations associated with rotenone lesioning. KV attenuated the loss of nigrostriatal dopaminergic neurons and perturbations in the striatal glucose-regulated protein (GRP78) and X-box binding protein 1 (XBP1) levels. Ileal epithelial injury following stereotaxic rotenone infusion was associated with oxidative stress and marked inhibition of acetylcholine esterase activity and reduced expression of occludin in the crypt and villi. While KV treatment attenuated the redox imbalance in the gut and enhanced occludin immunoreactivity, acetylcholinesterase activity was not affected. Our data demonstrate ileal oxidative damage as a characteristic non-motor gut dysfunction in PD while showing the potential dual efficacy of KV in the attenuation of both neural defects and gut abnormalities associated with PD.

Potential therapeutic effects of antagonizing adenosine A2A receptor, curcumin and niacin in rotenone-induced Parkinson's disease mice model.

Parkinson's disease (PD) is the second common age-related neurodegenerative disease. It is characterized by control loss of voluntary movements control, resting tremor, postural instability, bradykinesia, and rigidity. The aim of the present work is to evaluate curcumin, niacin, dopaminergic and non-dopaminergic drugs in mice model of Parkinson's disease through behavioral, biochemical, genetic and histopathological observations. Mice treated with rotenone rerecorded significant increase in adenosine A2A receptor (A2AR) gene expression, α synuclein, acetylcholinesterase (AchE), malondialdehyde (MDA), angiotensin-II (Ang-II), c-reactive protein (CRP), interleukin-6 (IL-6), caspase-3 (Cas-3) and DNA fragmentation levels as compared with the control group. While, significant decrease in dopamine (DA), norepinephrine (NE), serotonin (5-HT), superoxide dismutase (SOD), reduced glutathione (GSH), ATP, succinate and lactate dehydrogenases (SDH &LDH) levels were detected. Treatment with curcumin, niacin, adenosine A2AR antagonist; ZM241385 and their combination enhanced the animals' behavior and restored all the selected parameters with variable degrees of improvement. The brain histopathological features of hippocampal and substantia nigra regions confirmed our results. In conclusion, the combination of curcumin, niacin and ZM241385 recorded the most potent treatment effect in Parkinsonism mice followed by ZM241385, as a single treatment. ZM241385 succeeded to antagonize adenosine A2A receptor by diminishing its gene expression and ameliorating all biochemical parameters under investigation. The newly investigated agent; ZM241385 has almost the same pattern of improvement as the classical drug; Sinemet®. This could shed the light to the need of detailed studies on ZM241385 for its possible role as a promising treatment against PD. Additionally, food supplements such as curcumin and niacin were effective in Parkinson's disease eradication.

Tephrosin attenuates sepsis induced acute lung injury in rats by impeding expression of ICAM-1 and MIP-2.

Acute lung injury (ALI), a devastating form of respiratory infections, is characterized by increased edema, release of cytokines, weakened arterial oxygenation and infiltration of neutrophils and lymphocytes. The objective of the research envisaged was to reveal protective effects of tephrosin (TP) in ALI. In the present investigation, sepsis was triggered in rats by cecal ligation and puncture (CLP) method, and TP was administered intraperitonially. Five groups - Group A (control), Group B (Sham group) Group C (infected and untreated), and Group D and E (infected and treated with 25 and 50 mg/kg TP respectively) - of ten rats each, were used for the investigation. Evaluation parameters included measurement of arterial oxygenation, lung water content, protein determination, cytokine determination, neutrophil and lymphocyte count in the bronchoalveolar lavage fluid (BALF). As indicated by histopathological examination, the lung injury score was maximum in group C, but indicated reduction in group D and E. Intracellular adhesion molecule (ICAM)-1 and macrophage inflammatory protein-2 (MIP-2) are known to be important mediators responsible for ALI. Reduction in the ICAM-1 and MIP-2 expression was found to reduce after treatment with TP. In comparison to group D, group E reflected higher magnitude of ICAM-1 and MIP-2 suppression due to administration of higher TP dose. Compared to Group A and B, Group E indicated slightly higher expression of ICAM-1 and MIP-2. The research envisaged thus supports that TP attenuates ICAM-1 and MIP-2 expression in sepsis induced ALI rat model.

Therapeutic effects of baicalein on rotenone-induced Parkinson's disease through protecting mitochondrial function and biogenesis.

Mitochondrial dysfunction has been implicated in the pathogenesis of Parkinson's disease (PD) for several decades, and disturbed mitochondrial biogenesis (mitobiogenesis) was recently found to be a common phenomenon in PD. Baicalein, a major bioactive flavone of Scutellaria baicalensis Georgi, exerted neuroprotective effects in several experimental PD models. However, the effects of baicalein in rotenone-induced PD rats and the possible mechanisms remain poorly understood. In this study, we evaluated the therapeutic effects of baicalein and explored its mechanism of action in rotenone-induced PD models. The results indicated that behavioural impairments and the depletion of dopaminergic neurons induced by rotenone were attenuated by baicalein. Furthermore, in rotenone-induced parkinsonian rats, baicalein treatment effectively restored mitochondrial function and improved mitobiogenesis, as determined by measuring the mitochondrial density and key regulators involved in mitobiogenesis. Additionally, we confirmed that baicalein enhanced mitobiogenesis through the cAMP-responsive element binding protein (CREB) and glycogen synthase kinase-3ß (GSK-3ß) pathways in rotenone-treated SH-SY5Y cells. Moreover, we demonstrated that the cytoprotective effects of baicalein could be attenuated by the mitobiogenesis inhibitor chloramphenicol as well as CREB siRNA transfection. Overall, our results suggested that baicalein partially enhanced mitobiogenesis to restore mitochondrial function, thus exerting therapeutic effects in rotenone-induced PD models.

Ammonia promotes endothelial cell survival via the heme oxygenase-1-mediated release of carbon monoxide.

Although endothelial cells produce substantial quantities of ammonia during cell metabolism, the physiologic role of this gas in these cells is not known. In this study, we investigated if ammonia regulates the expression of heme oxygenase-1 (HO-1), and if this enzyme influences the biological actions of ammonia on endothelial cells. Exogenously administered ammonia, given as ammonium chloride or ammonium hydroxide, or endogenously generated ammonia stimulated HO-1 protein expression in cultured human and murine endothelial cells. Dietary supplementation of ammonia also induced HO-1 protein expression in murine arteries. The increase in HO-1 protein by ammonia in endothelial cells was first detected 4h after ammonia exposure and was associated with the induction of HO-1 mRNA, enhanced production of reactive oxygen species (ROS), and increased expression and activity of NF-E2-related factor-2 (Nrf2). Ammonia also activated the HO-1 promoter and this was blocked by mutating the antioxidant responsive element or by overexpressing dominant-negative Nrf2. The induction of HO-1 expression by ammonia was dependent on ROS formation and prevented by N-acetylcysteine or rotenone. Finally, prior treatment of endothelial cells with ammonia inhibited tumor necrosis factor-α-stimulated cell death. However, silencing HO-1 expression abrogated the protective action of ammonia and this was reversed by the administration of carbon monoxide but not bilirubin or iron. In conclusion, this study demonstrates that ammonia stimulates the expression of HO-1 in endothelial cells via the ROS-Nrf2 pathway, and that the induction of HO-1 contributes to the cytoprotective action of ammonia by generating carbon monoxide. Moreover, it identifies ammonia as a potentially important signaling gas in the vasculature that promotes endothelial cell survival.

Pyrroloquinoline quinone (PQQ) producing Escherichia coli Nissle 1917 (EcN) alleviates age associated oxidative stress and hyperlipidemia, and improves mitochondrial function in ageing rats.

BACKGROUND: Ageing involves oxidative stress mediated by Reactive Oxygen Species (ROS) and mitochondrial dysfunction. The present work demonstrates the protective effect of PQQ producing EcN against rotenone induced mitochondrial oxidative stress and consequence of mitochondrial and cellular dysfunction in naturally ageing rat model. PQQ is a potent antioxidant molecule also known to stimulate mitochondrial biogenesis and function in mammals. METHODS: Firstly, adult rats (16-18 weeks old) were treated with rotenone (2.5 mg/kg body weight; i.p.) daily for 28 days along with PQQ (10 mg/kg diet, daily) and modified probiotic EcN strains (10(8) CFU twice weekly). Secondly, ageing rats (48-50 weeks old) were gavaged with probiotic EcN strains (10(8)CFU twice weekly) and PQQ (10 mg/kg diet, daily) for 8 months. RESULTS: PQQ producing EcN-5 treatment prevented rotenone induced hepatic oxidative stress and mitochondrial damage in rats as assessed by reduced lipid peroxidation (29%), elevated glutathione (GSH) content (43%), increased catalase (52%) and superoxide dismutase (52%) activities when compared to only rotenone treatment. Moreover, increased hepatic mitochondrial content (41%), peroxisome proliferator-activated receptor-gamma coactivator 1-alpha (PGC-1α) mRNA (25%) and mitochondrial Superoxide Dismutase (Mit-SOD) activity (94%) were also observed in EcN-5 treated rats. Rotenone treated rats did not exhibit gain in body weight, whereas rats co-treated with EcN-5 showed significant restoration in body weight gain. Furthermore, weekly administration of EcN-5 to naturally ageing rats for eight months resulted in significant reduction of oxidative stress in hepatic and colonic tissues (assessed by lipid peroxidation, GSH content and catalase and SOD enzyme activities) along with increase in hepatic mitochondrial enzyme activities (Mit-SOD and succinate dehydrogenase) and biogenesis, when compared to untreated rats. Additionally, these rats also exhibited reduced expression of fatty acid synthase (50%) and increased expression of acyl coenzyme oxidase (225%) genes in liver in contrast to untreated rats resulting in lowered triglyceride (13% & 13.5%) and cholesterol (21% & 27%) levels in plasma and liver, respectively. Increased levels of butyrate (93%), propionate (45%) and acetate (18%) were also found in colonic content of these rats. PQQ administered daily (supplemented in diet) exhibited more or less similar effect as weekly gavaged EcN-5 in both the experiments, which substantiate that these effects are mediated by PQQ. CONCLUSION: These results suggest that genetically modified EcN-5 can be used as a nutritional supplement which can reduce age related oxidative stress and hyperlipidemia. Furthermore, it also rejuvenates healthy mitochondria by stimulating mitochondrial biogenesis and metabolism.

Mirabijalone E: a novel rotenoid from Mirabilis himalaica inhibited A549 cell growth in vitro and in vivo.

ETHNOPHARMACOLOGICAL RELEVANCE: The roots of Mirabilis himalaica have been used in Tibetan folk medicine for treatment of uterine cancer, nephritis edematous, renal calculus and arthrodynia. In our previous work, the ethanol extract of roots had shown potent cytotoxicity against human cancer cells. However, no information is available on the antitumor effect of Mirabilis himalaica. The aim of the present study was to investigate the active constituents guided by bioassay and evaluate the related antitumor efficacy in vitro and in vivo. MATERIALS AND METHODS: The active subextract (ethyl acetate) was subjected to successive chemical separation using a combination of silica gel, LH-20 chromatography and semi-preparative HPLC. The structures were determined by spectroscopic analysis techniques such as nuclear magnetic resonance (NMR) and mass spectrometry. Three human cancer cell lines, A549, HepG2 and HeLa were used for in vitro cytotoxicity evaluation of all isolated compounds by MTT-assay. Then, the potent and novel compound mirabijalone E was employed to the mechanism study againstA549 cells. BrdU immunofluorescence, soft agar assay and cell cycle analysis were employed to detect the cell proliferation effects. Annexin V-FITC/PI staining assay was used for examining apoptotic effects. Expression levels of apoptosis-related proteins were determined by western blot assay. in vivo tumorigenic assay was used to evaluate the xenograft tumor growth treated with mirabijalone E. RESULTS: One new rotenoid compound, mirabijalone E, together with eight known rotenoids was isolated from Mirabilis himalaica. Mirabijalone E, 9-O-methyl-inone B, boeravinone C and boeravinone H exhibited cytotoxicity against A 549 and HeLa cells. Further study on mirabijalone E was carried out in vitro and in vivo. Mirabijalone E inhibited A549 cells growth in a time and dose-dependent manner, which arrested cell cycle in S phase. Mechanistically, mirabijalone E treatment resulted in the increase of Bax expression level, the decrease of Bcl-2 level and the activation of caspase-3, which suggested the activation of apoptosis cascades. Consequently, the xenograft treated with mirabijalone E showed markedly suppressed tumor growth. CONCLUSIONS: The result suggested that mirabijalone E, together with active compounds, 9-O-methyl-4-hydroxyboeravinone B, boeravinone C and boeravinone H could be a promising candidate for cancer therapy.

The exploration of rotenone as a toxin for inducing Parkinson's disease in rats, for application in BBB transport and PK-PD experiments.

INTRODUCTION: In search for a suitable rat model to study potentially affected blood-brain barrier (BBB) transport mechanisms in the course of Parkinsons disease (PD) progression, experiments were performed to characterise Parkinsons disease markers following subcutaneous (SC) and intracerebral (IC) infusion of the toxin rotenone in the rat. METHODS: Studies were performed using Male Lewis rats. SC infusion of rotenone (3 mg/kg/day) was performed via an osmotic minipump. IC infusion of rotenone occurred directly into the right medial forebrain bundle at three different dosages. At different times following rotenone infusion, behaviour, histopathology (tyrosine hydroxylase and alpha-synuclein immunocytochemistry), peripheral organ pathology (adrenals, heart, kidney, liver, lung, spleen and stomach) were assessed. In part of the SC and IC rats, BBB transport profiles of the permeability marker sodium fluorescein were determined using microdialysis. RESULTS: SC rotenone failed to produce dopaminergic lesions and led to extensive peripheral organ toxicity. BBB permeability for fluorescein following SC rotenone was changed, however due peripheral toxicity. In contrast, IC rotenone produced a progressive lesion of the nigrostrial dopaminergic pathway over 28 days with no associated peripheral toxicity. IC rotenone also exhibited a large increase in amphetamine induced rotational behaviour. In addition, a few IC rats showed alpha-synuclein immunoreactivity and aggregation. Following IC rotenone, no changes in BBB permeability were detected after 14 days. DISCUSSION: SC rotenone only produced peripheral toxicity. IC rotenone appeared to create a progressive lesion of the rat nigrostrial pathway, and may therefore be a more appropriate model of Parkinson's disease progression, compared with the most commonly used 6-OH-DA rat model.

Deguelin regulates nuclear pore complex proteins Nup98 and Nup88 in U937 cells in vitro.

AIM: To investigate the anticancer effects and the molecular mechanisms of deguelin on human U937 leukemia cells, and to explore the underlying mechanism regulating nucleoporin 98 (Nup98) and nucleoporin 88 (Nup88) in vitro. METHODS: The effects of deguelin on the growth of U937 cells were studied by MTT assay. The effect of deguelin on the cell cycle of U937 cells was studied by using a propidium iodide method. The localization of the nuclear pore complex proteins Nup98 and Nup88 was investigated by using immunofluorescence and immunoelectron microscopy. The expression of Nup98 and Nup88 in U937 cells was investigated by using flow cytometry and Western blot. RESULTS: The proliferation of U937 cells was inhibited in the deguelin-treated group, with a 24-h IC(50) value of 21.61 nmol/L and a 36-h IC(50) value of 17.07 nmol/L. U937 cells treated with deguelin had reduced percentages of cells in the G(0)/G(1) phase, whereas cells accumulated in the S and G(2)/M phases. Nup88 and Nup98 were found on both the nuclear and cytoplasmic sides of the U937 cells by using immunofluorescence and immunoelectron microscopy. The expression of Nup98 was upregulated and that of the Nup88 protein was downregulated in U937 cells treated with deguelin. CONCLUSION: Deguelin is able to inhibit the proliferation of U937 cells by regulating the cell cycle such that cells are arrested at the S and G(2)/M phases, so that the proportion of cells in the G(0)/G(1) phase decreases. The antitumor effects of deguelin are related to upregulating the expression of Nup98 and downregulating the expression of Nup88 protein in U937 cells.

Cancer chemopreventive activity of rotenoids from Derris trifoliata.

A study of the chemical constituents of the stems of Derris trifoliata Lour. (Leguminosae) led to the isolation and identification of one new rotenoid, 6aalpha,12aalpha-12a-hydroxyelliptone ( 3), together with five other known rotenoids. In a search for novel cancer chemopreventive agents (anti-tumor promoters), we carried out a primary screening of five of the rotenoids isolated from the plant for their inhibitory effects on Epstein-Barr virus early antigen (EBV-EA) activation induced by 12- O-tetradecanoylphorbol 13-acetate (TPA) in Raji cells. The inhibitory activity of 3 was found to be equivalent to that of beta-carotene without any cytotoxicity. Deguelin ( 4) and alpha-toxicarol ( 5) exhibited a marked inhibitory effect on mouse skin tumor promotion in an in vivo two-stage carcinogenesis test. This investigation indicated that rotenoids might be valuable anti-tumor promoters.