Chetna Singh

The involvement of nitric oxide in maneb-and paraquat-induced oxidative stress in rat polymorphonuclear leukocytes

Oxidative stress plays a crucial role in the manifestations of maneb (MB) and paraquat (PQ)-induced toxicity including MB+PQ-induced Parkinsons disease (PD). Polymorphonuclear leukocytes (PMNs) actively participate in the oxidative stress-mediated inflammation and organ toxicity. The present study was undertaken to investigate the MB- and/or PQ-induced alterations in the indices of oxidative stress in rat PMNs. Animals were treated with or without MB and/or PQ in an exposure time dependent manner. In some sets of experiments, the animals were pre-treated with NOS inhibitors NG-nitro-L-arginine methyl ester (L-NAME) and aminoguanidine (AG) along with respective controls. A significant increase in myeloperoxidase (MPO), superoxide dismutase (SOD), nitric oxide, iNOS expression and lipid peroxidation (LPO) was observed in PMNs of MB- and/or PQ-treated animals, while catalase and glutathione S-transferase (GST) activities were attenuated. L-NAME and AG significantly reduced the augmented nitrite content, iNOS expression and MPO activity to control level in MB and PQ exposed animals. Although the augmented LPO was also reduced significantly in L-NAME and AG treated rat PMNs, the level was still higher as compared with controls. Alterations induced in SOD and GST activities were not affected by NOS inhibitors. The results thus suggest that MB and/or PQ induce iNOS-mediated nitric oxide production, which in turn increases MPO activity and lipid peroxidation, thereby oxidative stress.

Biochemical and molecular mechanisms of N-acetyl cysteine and silymarin-mediated protection against maneb- and paraquat-induced hepatotoxicity in rats

Oxidative stress is one of the major players in the pathogenesis of maneb (MB) and paraquat (PQ)-induced disorders. N-acetyl cysteine (NAC), a glutathione (GSH) precursor and silymarin (SIL), a naturally occurring antioxidant, encounter oxidative stress-mediated cellular damage. The present study was aimed to investigate the effects of NAC and SIL against MB and/or PQ-induced hepatotoxicity in rats. The levels of hepatotoxicity markers - alanine aminotransaminase (ALT), aspartate aminotransaminase (AST) and total bilirubin, histological changes, oxidative stress indices, phase I and phase II xenobiotic metabolizing enzymes - cytochrome P450 (CYP) and glutathione S-transferase (GST) and pro-inflammatory molecules - inducible nitric oxide synthase (iNOS), tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) were measured in animals treated with MB and/or PQ in the presence or absence of NAC and SIL. MB and/or PQ augmented ALT, AST, total bilirubin, lipid peroxidation and nitrite contents and catalytic activities of superoxide dismutase and glutathione peroxidase however, the GSH content was attenuated. NAC and SIL restored the above-mentioned alterations towards basal levels but the restorations were more pronounced in SIL treated groups. Similarly, MB and/or PQ-mediated histopathological symptoms and changes in the catalytic activities/expressions of CYP1A2, CYP2E1, iNOS, TNF-α, and IL-1β were alleviated by NAC and SIL. Conversely, MB and/or PQ-induced GSTA4-4 expression/activity was further increased by NAC/SIL and glutathione reductase activity was also increased. The results obtained thus suggest that NAC and SIL protect MB and/or PQ-induced hepatotoxicity by reducing oxidative stress, inflammation and by modulating xenobitic metabolizing machinery and SIL seems to be more effective.

Involvement of NADPH oxidase and glutathione in zinc-induced dopaminergic neurodegeneration in rats: similarity with paraquat neurotoxicity.

An association between excessive zinc (Zn) accumulation in brain and incidences of Parkinsons disease (PD) has been shown in several epidemiological and experimental investigations. The involvement of nicotinamide adenine dinucleotide phosphate (NADPH) oxidase and glutathione (GSH) in the pathogenesis of PD has also been proposed in a few studies. Despite the implicated role of oxidative stress in PD, the entire mechanism of Zn-induced dopaminergic neurodegeneration has not yet been clearly understood. The present study aimed to investigate the involvement of NADPH oxidase and GSH in Zn-induced dopaminergic neurodegeneration and also to assess its similarity with paraquat (PQ)-induced rat model of PD. Male Wistar rats were treated either with Zn (20 mg/kg; i.p.) or PQ (5 mg/kg; i.p.) in the presence and absence of NADPH oxidase inhibitor, apocynin (10 mg/kg; i.p.) and a GSH precursor, N-acetyl cysteine (NAC; 200 mg/kg; i.p.) either alone or in combination along with the respective controls. Apocynin and/or NAC pre-treatment significantly alleviated Zn- and PQ-induced changes in neurobehavioral deficits, number of dopaminergic neurons and contents of the striatal dopamine and its metabolites. Apocynin and/or NAC also mitigated Zn- and PQ-induced alterations in oxidative stress, NADPH oxidase activation and cytochrome c release, caspases-9 and -3 activation and CD11b expression. The results obtained thus suggest that Zn induces oxidative stress via the activation of NADPH oxidase and depletion of GSH, which in turn activate the apoptotic machinery leading to dopaminergic neurodegeneration similar to PQ.

Effect of zinc and paraquat co-exposure on neurodegeneration: Modulation of oxidative stress and expression of metallothioneins, toxicant responsive and transporter genes in rats

Abstract Oxidative stress is implicated in Parkinsons disease (PD). Metallothioneins (MT), cytochrome P450 IIE1 (CYP2E1) and glutathione S-transferases alpha4-4 (GSTA4-4) are involved in oxidative stress-mediated damage. Altered dopamine transporter (DAT) and vesicular monoamine transporter-2 (VMAT-2) are also documented in PD. The present study was undertaken to investigate the effect of Zn and PQ co-exposure on neurodegeneration in rats. A significant reduction was observed in spontaneous locomotor activity (SLA), striatal dopamine (DA) levels, tyrosine hydroxylase (TH) immunoreactivity, glutathione reductase (GR) and catalase activity along with increased lipid peroxidation (LPO) and glutathione peroxidase (GPx) activity after Zn and/or PQ exposure. Zn and/or PQ exposure increased gene expression of DAT, CYP2E1, GSTA4-4, MT-I and MT-II, but reduced the expression of VMAT-2. Protein expression analysis of TH, VMAT-2 and DAT showed results similar to those obtained with gene expression study. Zn and PQ co-exposure caused a more pronounced effect than that of individual exposure. The results obtained in this study suggest that, similar to PQ, Zn induced neurodegeneration via alterations in oxidative stress and expression of the above-mentioned genes. However, the effect of Zn+PQ was only slightly higher than that of alone, indicating that probably Zn and PQ follow some different molecular events leading to neurodegeneration.

The expression of CYP2D22, an ortholog of human CYP2D6, in mouse striatum and its modulation in 1-methyl 4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinson's disease phenotype and nicotine-mediated neuroprotection.

The main contributory factors of Parkinsons disease (PD) are aging, genetic factors, and environmental exposure to pesticides and heavy metals. CYP2D22, a mouse ortholog of human CYP2D6, is expected to participate in a chemically induced PD phenotype due to its structural resemblance with CYP2D6. Despite its expected participation in PD, its expression in the nigrostriatal tissues and modulation by the chemicals that induce PD or offer neuroprotection have not yet been investigated. The present study was undertaken to investigate CYP2D22 expression in mouse striatum and to assess its involvement in the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD phenotype and nicotine-mediated neuroprotection. The animals were treated intraperitoneally daily with nicotine (1 mg/kg) for 8 weeks, followed by MPTP (20 mg/kg) + nicotine (1 mg/kg) for 2-4 weeks and vice versa, along with respective controls. In some sets of experiments, the animals were treated with ketoconazole (300 mg/kg), a CYP3AH/CYP2D22 inhibitor, along with nicotine and/or MPTP. Tyrosine hydroxylase (TH)-immunoreactivity in the substantia nigra, the expression of nicotinic acetylcholine receptors (nAChRs) alpha6 and alpha4, dopamine content, and 1-methyl-4-phenylpyridinium ion (MPP(+)) level in the striatum were measured to confirm the MPTP-induced PD phenotype and nicotine-mediated neuroprotection. CYP2D22 and nAChRs expressions were measured in the striatum by RT-PCR/western blotting and dopamine level; CYP2D22 catalytic activity and MPP(+) content were determined by high-performance liquid chromatography (HPLC). MPTP increased dopaminergic neuronal degeneration and the striatal MPP(+) level and reduced striatal dopamine content; it attenuated expression/activity of CYP2D22 and nAChRs that were significantly restored in nicotine-treated animals. Ketoconazole reduced the nicotine-mediated increase in CYP2D22 expression and activity, dopamine content, and TH-immunoreactivity. The results indicate the expression of CYP2D22 in mouse striatum and its possible role in the MPTP-induced PD phenotype and nicotine-mediated neuroprotection.

Effects of cypermethrin on monoamine transporters, xenobiotic metabolizing enzymes and lipid peroxidation in the rat nigrostriatal system

Abstract Long-term exposure to cypermethrin induces the nigrostriatal dopaminergic neurodegeneration in adult rats and its pre-exposure in the critical periods of brain development enhances the susceptibility during adulthood. Monoamine transporters, xenobiotic metabolizing enzymes and oxidative stress play critical roles in the nigrostriatal dopaminergic neurodegeneration. The study was undertaken to investigate the effects of cypermethrin on DAT, VMAT 2, CYP2E1, GST Ya, GST Yc and GSTA4-4 expressions, CYP2E1 and GST activities and lipid peroxidation in the nigrostriatal system of adult rats with/without post-natal exposure to cypermethrin. Cypermethrin reduced VMAT 2 and increased CYP2E1 expressions without causing significant change in DAT. Although GSTA4-4 mRNA expression and lipid peroxidation were increased, no significant changes were observed in GST Ya and GST Yc expressions and total GST activity. The results obtained demonstrate that long-term exposure to cypermethrin modulates VMAT 2, CYP2E1, GSTA4-4 expressions and lipid peroxidation, which could contribute to the nigrostriatal dopaminergic neurodegeneration.

Nicotine and caffeine-mediated modulation in the expression of toxicant responsive genes and vesicular monoamine transporter-2 in 1-methyl 4-phenyl-1,2,3,6-tetrahydropyridine-induced Parkinson's disease phenotype in mouse

Epidemiological evidence revealed that cigarette smokers and coffee drinkers have lower risk of Parkinsons disease (PD). Nicotine inhibits monoamine oxidase activity, and induces expression of neurotrophic factors and nicotinic acetylcholinergic receptors. However, caffeine is capable of antagonizing adenosine A(2A) receptor. Toxicant responsive enzymes and vesicular monoamine transporter-2 (VMAT-2) play critical roles in chemically induced PD. Despite some known functions, the effects of nicotine and caffeine on the expression and activity of toxicant responsive genes and on VMAT-2 are still not known. The study was therefore undertaken to investigate the effect of nicotine and caffeine on the expression and activity of toxicant responsive genes, i.e., CYP1A1, CYP2E1, GST-ya, GST-yc, GSTA4-4 and VMAT-2 in the striatum of control and MPTP-induced PD phenotype in mouse. The animals were treated intraperitoneally daily with nicotine (1 mg/kg) or caffeine (20 mg/kg) for 8 weeks, followed by 1-methyl 4-phenyl-1,2,3,6-tetrahydropyridine (MPTP; 20 mg/kg)+nicotine or caffeine for 4 weeks. MPTP significantly attenuated CYP1A1 and VMAT-2, and augmented CYP2E1, GST-ya, GST-yc and GSTA4-4 expression/activity. Nicotine or caffeine-treated animals showed significant restoration against most of the MPTP-induced alterations. The results obtained thus suggest that nicotine and caffeine modulate MPTP-induced alterations in CYP1A1, CYP2E1, GST-ya, GST-yc, GSTA4-4 and VMAT-2 expression/activity.

Oxidative stress in zinc-induced dopaminergic neurodegeneration: Implications of superoxide dismutase and heme oxygenase-1

Abstract The study was undertaken to investigate the effect of zinc (Zn) on glutathione S-transferase (GST) and superoxide dismutases (SOD) activities and on the expressions of cytosolic Cu, Zn-SOD (SOD1), mitochondrial Mn-SOD (SOD2), γ-glutamyl cysteine synthetase (γ-GCS) and heme oxygenase-1 (HO-1) in the nigrostriatal tissue of rats. Additionally, Zn-induced alterations in the neurobehavioral parameters, lipid peroxidation (LPO), striatal dopamine and its metabolites and tyrosine hydroxylase (TH) protein expression were measured to assess their correlations with the oxidative stress. Zn exposure reduced the locomotor activity, rotarod performance, striatal dopamine and its metabolites and TH protein expression. LPO, total SOD, SOD1 and SOD2 activities were increased while GST and catalase were reduced in a dose and time dependent manner. Expressions of SOD1 and HO-1 were increased while no change was observed in SOD2 and γ-GCS expressions. The results obtained suggest that Zn-induced augmentation of total SOD, SOD1, SOD2 and HO-1 was associated with increased oxidative stress and neurodegenerative indexes indicating the involvement of both cytosolic and mitochondrial machinery in Zn-induced oxidative stress leading to dopaminergic neurodegeneration.

Maneb and paraquat-induced modulation of toxicant responsive genes in the rat liver: Comparison with polymorphonuclear leukocytes

Experimental studies have shown that toxicant responsive genes, cytochrome P450s (CYPs) and glutathione S-transferases (GSTs) play a critical role in pesticide-induced toxicity. CYPs play pro-oxidant role and GSTs offer protection in maneb (MB) and paraquat (PQ)-induced brain and lung toxicities. The present study aimed to investigate the effect of repeated exposures of MB and/or PQ on lipid peroxidation (LPO), glutathione content (GSH) and toxicant responsive genes, i.e., CYP1A1, 1A2, 2E1, GSTA4-4, GSTA1-1 and GSTA3-3 in the liver and to correlate the same with polymorphonuclear leukocytes (PMNs). A significant augmentation in LPO and reduction in GSH content was observed in a time of exposure dependent manner in the liver and PMNs of MB and/or PQ treated animals. The expression and catalytic activity of CYP2E1 and GSTA4-4 were significantly increased following MB and/or PQ exposure both in the liver and PMNs. Although the expression of GSTA3-3 was increased, the expression of GSTA1-1 was unaltered after MB and/or PQ treatment in both the liver and PMNs. MB augmented the expression and catalytic activity of CYP1A1 in the liver, however, CYP1A2 was unaffected. PQ, on the other hand, significantly increased hepatic CYP1A2 expression and catalytic activity. MB and/or PQ did not produce any significant changes in CYP1A1 and CYP1A2 in PMNs. The results of the study thus demonstrate that MB and PQ differentially regulate hepatic CYP1A1 and CYP1A2 while LPO, GSH, CYP2E1, GSTA4-4 and GSTA3-3 are modulated in the similar fashions both in the liver and PMNs.

Solubilisation of low grade rock phosphate by composting with a farm waste, pearl-millet Boobla

Abstract Indigenous low grade rock phosphate (RP) was solubilised by composting with pearl-millet Boobla (pmb) in the presence of nitrogen, molasses and pyrites. The solubilisation of RP was rapid up to 15 days in pmb + N + RP and reached a maximum of 60% in 120 days of composting. Molasses had no beneficial effect on solubilisation while pyrites decreased it drastically in spite of decreasing the pH and increasing the citric-soluble P of the composting material. The concentration of total P increased in all treatments as organic matter decreased during composting. However, the release of Olsens and water-soluble P increased up to 60 days and decreased thereafter except with pmb + N + RP, where Olsens P increased up to 90 days. Loss of organic matter and organic P increased regularly with composting period and was at a maximum in the pmb + N + RP treatment.