RESUMO
Iron is widely linked with the onset and development of Parkinson's disease (PD). Accumulation of iron induces free radical generation and promotes α-synuclein aggregation, oxidative stress, and autophagy impairment. Deferoxamine, an iron chelator, is shown to ameliorate iron dyshomeostasis in rodents and humans. However, the role of deferoxamine in cypermethrin-induced iron accumulation is not yet known. Although an iron accumulation and impaired chaperone-mediated autophagy (CMA) contribute to PD, a link between the two is not yet widely understood. Current study is undertaken to explore the possible association between an iron accumulation and CMA in cypermethrin model of PD in the presence of deferoxamine. Level of iron, iron transporter proteins, oxidative stress, and CMA proteins along with indicators of Parkinsonism were measured. Deferoxamine attenuated cypermethrin-induced iron accumulation and number of iron-positive cells and ameliorated the demise of dopaminergic cells and dopamine content. Deferoxamine significantly normalizes cypermethrin-induced changes in iron transporter proteins, α-synuclein, lysosome-associated membrane protein-2A, and oxidative stress. The results demonstrate that deferoxamine ameliorates cypermethrin-induced iron dyshomeostasis and impairment in CMA.
RESUMO
Cadmium (Cd) exposure to the animals including humans is reported as nephrotoxic compounds i.e., disturbing redox status (increase oxidative stress), mitochondrial dysfunction, renal cell death and altered transporters in the renal system. Hsp27 (a small heat shock protein) has been shown as one of the modulators in the renal dysfunction and increased against the Cd induced toxicity. However, no studies are reported on the genetic modulation of stress protein against the Cd-induced nephrotoxicity. The current study aimed to examine the protective role of hsp27 overexpression against the Cd-induced nephrotoxicity using Drosophila melanogaster as an animal model. D. melanogaster renal system includes nephrocytes and Malpighian tubules (MTs) that show the functional similarity with mammalian kidney nephron. Overexpression of the hsp27 was found to reduce the Cd induced oxidative stress, rescue cell death in MTs of Cd exposed D. melanogaster larvae. The rescued GSH level, NADPH level and glucose 6 phosphate dehydrogenase (G6PD) activity were also observed in the MTs of the Cd exposed organism. Function (efflux activity and fluid secretion rate) of the MTs was restored in Cd exposed hsp27 overexpressed larvae. Further, results were confirmed by restored brush border microvilli density and reduced uric acid level. Tissue specific knockdown of hsp27 developed Cd like phenotypes in MTs and the phenotypes enhanced in Cd exposed condition. The present study clearly shows the role of hsp27 overexpression in restoration of the MTs function and protection against the Cd induced renal toxicity.
Assuntos
Cádmio , Drosophila melanogaster , Humanos , Animais , Drosophila melanogaster/genética , Drosophila melanogaster/metabolismo , Cádmio/toxicidade , Cádmio/metabolismo , Rim/metabolismo , Estresse Oxidativo , Proteínas de Choque Térmico/genética , Proteínas de Choque Térmico/metabolismo , Mamíferos/metabolismoRESUMO
The etiology of Parkinson's disease (PD) is highly complex and is still indefinable. However, a number of studies have indicated the involvement of pesticides and transition metals. Copper, magnesium, iron, and zinc have emerged as important metal contributors. Exposure to pesticides causes an accumulation of transition metals in the substantia nigra (SN) region of the brain. The cypermethrin model of PD is characterized by mitochondrial dysfunction, autophagy impairment, oxidative stress, etc. However, the effect of cypermethrin on metal homeostasis is not yet explored. The study was designed to delineate the role of metals and their transporter proteins in cypermethrin-induced animal and cellular models of PD. The level of copper, magnesium, iron, and zinc was checked in the nigrostriatal tissue and serum by atomic absorption spectroscopy. Since cypermethrin consistently increased iron content in the nigrostriatal tissue and serum after 12 weeks of exposure, the level of iron transporter proteins, such as divalent metal transporter-1 (DMT-1), ceruloplasmin, transferrin, ferroportin, and hepcidin, and their in silico interaction with cypermethrin were checked. 3,3'-Diaminobenzidine-enhanced Perl's staining showed an elevated number of iron-positive cells in the SN of cypermethrin-treated rats. Molecular docking studies revealed a strong binding affinity between cypermethrin and iron transporter protein receptors of humans and rats. Furthermore, cypermethrin increased the expression of DMT-1 and hepcidin while reducing the expression of transferrin, ceruloplasmin, and ferroportin in the nigrostriatal tissue and human neuroblastoma cells. These observations suggest that cypermethrin alters the expression of iron transporter proteins leading to iron dyshomeostasis, which could contribute to dopaminergic neurotoxicity.
Assuntos
Doença de Parkinson , Praguicidas , Ratos , Humanos , Animais , Ferro/metabolismo , Doença de Parkinson/metabolismo , Hepcidinas/metabolismo , Cobre/metabolismo , Ceruloplasmina , Magnésio/farmacologia , Simulação de Acoplamento Molecular , Substância Negra/metabolismo , Transferrina/metabolismo , Zinco/metabolismoRESUMO
Benzodiazepines can make victims more docile, they are frequently used in drug-facilitated crimes, such as robberies and sexual assaults. Therefore, it is essential to develop techniques for determining whether these chemicals are present in relation with illegal activity is crucial. Therefore, to determine the presence of five benzodiazepines (alprazolam, clonazepam, diazepam, lorazepam, and oxazepam) in water, alcoholic beverages, and non-alcoholic beverages, a simple and direct, miniaturized, and effective vortex assisted ultrasound based microextraction using solidification of floating organic droplets (VAUS-ME-SFO) in combination with LC-MS/MS was developed. 1-Undecanol and acetonitrile, respectively, served as the extractant and disperser solvents. Many other parameters affect the efficiency of the developed analytical procedure VAUS-ME-SFO/LC-MS/MS. These parameters were optimized using Plackett Burman Design and Central Composite Design to obtain reliable results. The optimum conditions for the extraction were: 10.0 mL of sample; 180 µL acetonitrile, as a dispersive solvent; 200 µL of 1-undecanol, as an extraction solvent; pH 7; 105 s of vortex agitation; 120 s of ultrasonication application and 3 min of centrifugation at 7000 rpm. The benzodiazepines were separated by a chromatographic separation technique carried out by a UPLC system consisting of a binary mobile phase. The solvent system comprises of 0.1% Formic acid in Milli-Q (Solvent A) and 0.1% Formic acid in ACN (Solvent B) with a gradient flow of 3.5 min total analysis time. Under the optimized conditions, the calibration curve was studied in the range of 0.124-7.810 ng mL-1. The regression correlation coefficient (R2) value of all targeted analytes ranges from 0.993 to 0.999. The LOD and LOQ of VAUS-ME-SFO methods using LC-MS/MS analysis range from 0.316 to 0.968 ng mL-1 and 1.055-3.277 ng mL-1 respectively. The repeatability within a day varied from 0.6 to 3.5%, and the reproducibility across days varied from 2.2 to 6.3%. The recoveries ranges for water, alcoholic and non-alcoholic beverages from 70.77 to 114.53%, 63.20-102.21% and 66.23-113.28% respectively. Further, the degradation kinetics was studied to establish the half-life of each targeted analyte in the matrix undertaken in the study. The water samples were classified based on their BDZs residues. This implies that the more health care and anthropogenic activity, the more the BDZs residue will be in water samples.
Assuntos
Benzodiazepinas , Microextração em Fase Líquida , Cromatografia Líquida/métodos , Água , Reprodutibilidade dos Testes , Espectrometria de Massas em Tandem/métodos , Solventes/química , Bebidas , Microextração em Fase Líquida/métodosRESUMO
In recent years, special attention has been given to emission research that led to the deposition of toxicants from road traffic. Thus, it is imperative to focus on heavy metal (HM) stressors in food items, their source contribution, and health risk assessment providing insight into their spatial role at the population level. In this study, heavy metal in the street vended noodles was studied while correlating the quality of noodle with different environmental origins. The samples were prepared using acid digestion and analysed by flame atomic absorption spectrophotometer, except Hg which was analysed by direct mercury analyser. The results showed that some heavy metals like Cr, Pb, Mn, Cd, and Hg exceed their permissible limits established by the international legislation for food products. In the noodle samples, the concentration of heavy metal ranged from < 0.1 to 0.904 mg/kg for Pb, < 0.09 to 0.843 mg/kg for Ni, < 0.004 to 0.201 mg/kg for Cd, < 0.0001 to 0.004 mg/kg for Hg, < 0.01 to 1.388 mg/kg for Cu, < 0.015 to 8.049 mg/kg for Mn, and < 0.02 to 16.514 mg/kg for Cr. Noodle samples vended on high traffic density streets are directly associated with increased HM content due to atmospheric deposition from the surrounding. Source apportionment study determines that HM contamination belongs to the same source of origin, except Cr. Based on the cluster analysis, these samples fall into three major groups that were further validated by the canonical discriminant function. Health risk prediction by Monte Carlo simulation revealed an elevated non-carcinogenic health hazard risk to consumers with a hazard index (HI) shift from 71 to 75%. Health hazard analysis showed that consumers of high traffic density street vended food are at higher risk of developing health-related issues. This study is important to evaluate the health risk of the population exposed to heavy metals due to ingestion of street vended food.
Assuntos
Mercúrio , Metais Pesados , Poluentes do Solo , Cádmio/análise , Chumbo/análise , Metais Pesados/análise , Medição de Risco , Mercúrio/análise , Monitoramento Ambiental , China , Poluentes do Solo/análiseRESUMO
BACKGROUND: Combined maneb (MB) and paraquat (PQ), two widely used pesticides, increases oxidative stress leading to Parkinsonism. Xenobiotic metabolizing enzymes, cytochrome P450 (CYP) 2D6 and its mouse ortholog Cyp2d22 protect against Parkinsonism. Resveratrol, an antioxidant, restores antioxidant defense system through the activation of nuclear factor erythroid 2- related factor 2 (Nrf2). However, a crosstalk between Cyp2d22/CYP2D6-mediated protection and resveratrol-induced Nrf2 activation leading to neuroprotection is not yet elucidated. OBJECTIVE: The study aimed to decipher the effect of resveratrol on Nrf2 activation and expression of its downstream mediators, nicotinamide adenine dinucleotide phosphate quinone oxidoreductase 1 (NQO1) and thioredoxin 1 (Trx1) along with Cyp2d22/CYP2D6 activity in combined MB and PQ mouse model of Parkinsonism and differentiated neuroblastoma cells. RESULTS: MB and PQ reduced the dopamine content (mouse) and Cyp2d22/CYP2D6 activity (mouse/neuroblastoma cells) and increased the nuclear translocation of Nrf2 and expression of NQO1 and Trx1 (both). Resveratrol ameliorated pesticides-induced changes in dopamine content and Cyp2d22/CYP2D6 activity. It was found to promote nuclear translocation of Nrf2 and expression of NQO1 and Trx1 proteins. Since Cyp2d22/CYP2D6 inhibitor (ketoconazole/quinidine) per se reduced Cyp2d22/CYP2D6 activity and dopamine content, it was found to substantially increase the pesticides-induced reduction in Cyp2d22/CYP2D6 activity and dopamine content. Inhibitors normalized the pesticides induced changes in Nrf2 translocation and NQO1 and Trx1 levels in pesticides treated groups. CONCLUSION: The results suggest that resveratrol promotes the catalytic activity of xenobiotic metabolizing enzyme, Cyp2d22/CYP2D6, which partially contributes to Nrf2 activation in pesticides- induced Parkinsonism.
Assuntos
Antioxidantes/metabolismo , Família 2 do Citocromo P450/biossíntese , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Fator 2 Relacionado a NF-E2/metabolismo , Neuroproteção/efeitos dos fármacos , Doença de Parkinson Secundária , Praguicidas/toxicidade , Resveratrol/farmacologia , Animais , Linhagem Celular Tumoral , Masculino , Camundongos , NAD(P)H Desidrogenase (Quinona) , Doença de Parkinson Secundária/induzido quimicamente , Doença de Parkinson Secundária/metabolismo , Doença de Parkinson Secundária/prevenção & controle , Tiorredoxinas/biossínteseRESUMO
Epidemiological evidences have shown an association of exposure to pesticides or heavy metals with increased incidences of Parkinson's disease (PD) in humans. Exposure to pesticides or metals during the decisive period of the brain development increases the susceptibility of dopaminergic neurons upon re-exposure in adult rodents. However, the effect of early life exposure to pesticide on the heavy metal-induced neurodegeneration or heavy metal on pesticide-induced neurodegeneration is not yet explored. The current study explored the effect of developmental exposure to zinc (Zn), a metal or paraquat (PQ), a pesticide on the nigrostriatal dopaminergic neurons of rats challenged to Zn or PQ during adulthood. Exposure of Zn or PQ during adulthood alone exhibited marked reduction in motor activities, striatal dopamine and metabolites, glutathione content and number of dopaminergic neurons. However, the levels of lipid peroxidation, protein carbonyls, superoxide dismutase activity, pro-inflammatory cytokines and 4-hydroxynonenal-protein adducts were increased. While the expression of vesicular monoamine transporter-2 and tyrosine hydroxylase were attenuated, dopamine transporter and microglial marker Iba-1 expression, activated microglia, nuclear factor-kappa B activation, mitochondrial cytochrome c release and caspase-3/9 activation were augmented following Zn or PQ exposure. Albeit postnatal alone exposure did not alter any of the studied parameters, the developmental administration of Zn/PQ in re-challenged adult rats produced more pronounced changes in the aforementioned variables as compared with adulthood Zn or PQ alone intoxicated animals. The results demonstrate that postnatal Zn/PQ intoxication dents the oxidative stress, inflammation, cell death and dopamine metabolism and storage regulating machineries, which speed up the toxicant-induced degeneration during adulthood.
Assuntos
Neurônios Dopaminérgicos/citologia , Doenças Neurodegenerativas/metabolismo , Paraquat/efeitos adversos , Zinco/efeitos adversos , Aldeídos/metabolismo , Animais , Animais Recém-Nascidos , Sobrevivência Celular/efeitos dos fármacos , Citocinas/metabolismo , Modelos Animais de Doenças , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Glutationa/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Doenças Neurodegenerativas/induzido quimicamente , Estresse Oxidativo/efeitos dos fármacos , Ratos , Superóxido Dismutase/metabolismoRESUMO
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) exacerbates mitochondrial impairment and α-synuclein expression leading to Parkinsonism. Impaired mitochondria and over-expressed α-synuclein are degraded and eliminated via macroautophagy and chaperone-mediated autophagy. Owing to multiple properties, silymarin protects from oxidative stress-mediated cellular injury. However, its effect on MPTP-induced changes in autophagy is not yet known. The study aimed to decipher the effect of silymarin on MPTP-induced changes in autophagy. Male mice (20-25 g) were treated with silymarin (intraperitoneally, daily, 40 mg/kg) for 2 weeks. On day 7, a few animals were also administered with MPTP (intraperitoneally, 20 mg/kg, 4 injections at 2-h interval) along with vehicles. Striatal dopamine content was determined. Western blot analysis was done to assess α-synuclein, beclin-1, sequestosome, phosphorylated 5' adenosine monophosphate-activated protein kinase (p-AMPK), lysosome-associated membrane protein-2 (LAMP-2), heat shock cognate-70 (Hsc-70), LAMP-2A, phosphorylated unc-51-like autophagy activating kinase (p-Ulk1), and phosphorylated mechanistic target of rapamycin (p-mTOR) levels in the nigrostriatal tissue. Silymarin rescued from MPTP-induced increase in beclin-1, sequestosome, p-AMPK, and p-Ulk1 and decrease in LAMP-2, p-mTOR, and LAMP-2A levels. Silymarin defended against MPTP-induced increase in α-synuclein and reduction in dopamine content. The results demonstrate that silymarin protects against MPTP-induced changes in autophagy leading to Parkinsonism.
Assuntos
Autofagia , Intoxicação por MPTP/metabolismo , Fármacos Neuroprotetores/farmacologia , Silimarina/farmacologia , Quinases Proteína-Quinases Ativadas por AMP , Animais , Proteína Homóloga à Proteína-1 Relacionada à Autofagia/metabolismo , Proteína Beclina-1/metabolismo , Corpo Estriado/efeitos dos fármacos , Corpo Estriado/metabolismo , Dopamina/metabolismo , Proteína 2 de Membrana Associada ao Lisossomo/metabolismo , Intoxicação por MPTP/tratamento farmacológico , Masculino , Camundongos , Fármacos Neuroprotetores/uso terapêutico , Proteínas Quinases/metabolismo , Silimarina/uso terapêutico , Serina-Treonina Quinases TOR/metabolismo , alfa-Sinucleína/metabolismoRESUMO
Microgliosis and inflammation are major wrongdoers in cypermethrin-induced Parkinsonism along with oxidative stress, mitochondrial dysfunction and α-synuclein aggregation. Dopamine depletion could alter dendritic morphology, length and spine number in the striatum. Present study investigated the effect of ibuprofen on the dendritic morphology, length and spine density in cypermethrin PD model. Male pups were treated intraperitoneally with cypermethrin during postnatal days followed by adulthood to induce Parkinsonism using standard procedure along with controls. Subsets of animals were pre-treated with ibuprofen 2 h prior to cypermethrin treatment during adulthood. Standard methods were used to confirm Parkinsonism/neuroprotection. Striatal dendritic morphology, length, spine number and expression of synaptophysin and postsynaptic density protein-95 (PSD-95) along with the nigrostriatal pro-inflammatory and apoptotic proteins were measured. Cypermethrin induced Parkinsonian traits and attenuated the dendritic length, spine number and expression of synaptophysin and PSD-95. While cypermethrin increased the expression of interleukin-1ß, interleukin-4, interferon-γ, inducible nitric oxide synthase, caspase-3, caspase-9 and B-cell lymphoma (Bcl)-xl proteins, it attenuated Bcl-2 expression. Ibuprofen normalized the changes in dendritic morphology, length, spine number and expression of synaptophysin, PSD-95, and pro-inflammatory and apoptotic proteins. Results demonstrate that cypermethrin induces inflammation and alters dendritic morphology, length and spine number, which are encountered by ibuprofen.
Assuntos
Corpo Estriado/patologia , Dendritos/patologia , Espinhas Dendríticas/patologia , Ibuprofeno/uso terapêutico , Fármacos Neuroprotetores/uso terapêutico , Piretrinas/toxicidade , Animais , Animais Recém-Nascidos , Anti-Inflamatórios não Esteroides/farmacologia , Anti-Inflamatórios não Esteroides/uso terapêutico , Corpo Estriado/efeitos dos fármacos , Dendritos/efeitos dos fármacos , Espinhas Dendríticas/efeitos dos fármacos , Ibuprofeno/farmacologia , Masculino , Fármacos Neuroprotetores/farmacologia , Transtornos Parkinsonianos/induzido quimicamente , Transtornos Parkinsonianos/patologia , Transtornos Parkinsonianos/prevenção & controle , Ratos , Ratos WistarRESUMO
Inflammation is decisive in zinc (Zn)-induced nigrostriatal dopaminergic neurodegeneration; however, the contribution of cyclooxygenase-2 (COX-2) is not yet known. The present study aimed to explore the role of COX-2 in Zn-induced Parkinsonism and its association with the microglial activation. Male Wistar rats were treated intraperitoneally (i.p.) with Zn as zinc sulphate (20 mg/kg) along with respective controls for 2-12 weeks. In a few sets, animals were also treated with/without celecoxcib (CXB, 20 mg/kg, i.p.), a selective COX-2 inhibitor. Indexes of the nigrostriatal neurodegeneration, oxidative stress, inflammation and apoptosis were measured in the animals/nigrostriatal tissue. Zn induced time-dependent increase in the expression of COX-2 while COX-1 expression was unaltered. Zn reduced the neurobehavioral activities, striatal dopamine content, tyrosine hydroxylase (TH) expression and number of dopaminergic neurons. While oxidative stress; microglial activation; expression of microglial cell surface marker-CD11b; cytochrome c release; caspase-9/3 activation; level of pro-inflammatory cytokines, such as TNF-α, IL-1ß and IL-6 and Bcl-2-associated protein x (Bax) translocation from the cytosol to mitochondria were induced in the Zn-treated group, expression of B-cell lymphoma-2 (Bcl-2) was found to be reduced. CXB significantly attenuated Zn-induced increase in COX-2 expression and restored TH-expression, dopamine content, level of inflammatory cytokines and neurobehavioral indexes towards normalcy. Moreover, CXB also attenuated Zn-induced increase in microglial activation, oxidative stress and apoptotic markers towards normal levels. Results of the study thus demonstrate that COX-2 induces microglial activation that provokes the release of inflammatory mediators, which in turn augments oxidative stress and intrinsic apoptosis leading to dopaminergic neurodegeneration in Zn-induced Parkinsonism.
Assuntos
Apoptose/fisiologia , Ciclo-Oxigenase 2/metabolismo , Mediadores da Inflamação/metabolismo , Microglia/metabolismo , Estresse Oxidativo/fisiologia , Transtornos Parkinsonianos/metabolismo , Zinco/toxicidade , Animais , Apoptose/efeitos dos fármacos , Masculino , Microglia/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Transtornos Parkinsonianos/induzido quimicamente , Ratos , Ratos WistarRESUMO
Parkinson's disease (PD) is the second most familiar, progressive and movement-related neurodegenerative disorder after Alzheimer disease. This study aimed to decipher the role of autophagy in cypermethrin-induced Parkinsonism, an animal model of PD. Indicators of autophagy [expression of beclin 1, autophagy-related protein 12 (Atg 12), unc-51 like autophagy activating kinase 1 (Ulk 1), p62 and lysosome-associated membrane protein 2 (LAMP 2) and conversion of microtubule-associated protein 1A/1B-light chain 3 (LC3) I to II], signalling cascade [phosphorylated (p) 5' adenosine monophosphate-activated protein kinase (p-AMPK), sirtuin 1 (Sirt 1), phosphorylated-mammalian target of rapamycin (p-mTOR), tuberous sclerosis complex 2 (TSC 2), p317Ulk 1 and p757Ulk 1 levels] and lysosome morphology were assessed in control and cypermethrin-treated rat model of PD. Autophagy markers were also measured in cypermethrin-treated neuroblastoma cells in the presence of 3-methyl adenine, a phosphatidylinositol-4,5-bisphosphate 3-kinase (PI3K) class III inhibitor; vinblastine, an autophagosome elongation inhibitor; bafilomycin A1, an autophagolysosome and lysosome fusion/abnormal acidification inhibitor or torin 1, a mechanistic target of rapamycin inhibitor. Cypermethrin reduced LAMP 2 and increased p-AMPK and Sirt 1 without causing any change in other signalling proteins. 3-Methyl adenine did not change LC3 conversion; vinblastine and bafilomycin A1 decreased LAMP 2 expression in controls. While cypermethrin increased LC3 conversion in the presence of 3-methyl adenine, LAMP 2 reduction was more pronounced in vinblastine and bafilomycin A1-treated cells. Torin 1 normalized the expression of LAMP 2 without any change in other autophagy markers. Results demonstrate that albeit cypermethrin activates autophagosome formation, it reduces LAMP 2 expression and lysosome quality leading to autophagy inhibition.
Assuntos
Autofagia/efeitos dos fármacos , Lisossomos/efeitos dos fármacos , Doença de Parkinson/tratamento farmacológico , Piretrinas/farmacologia , Animais , Apoptose/efeitos dos fármacos , Autofagossomos/efeitos dos fármacos , Humanos , Lisossomos/metabolismo , Doenças Neurodegenerativas/tratamento farmacológico , Doenças Neurodegenerativas/metabolismo , Doença de Parkinson/metabolismo , Ratos Wistar , Serina-Treonina Quinases TOR/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismoRESUMO
The presence of pesticides in food items and beverages is a big threat to humankind, and their quantitative estimation with high precision and accuracy is always a challenge for analytical chemists. Hence, a simple and rapid method is proposed for the simultaneous determination of 30 pesticides in beverages (alcoholic and non-alcoholic drinks). The proposed method hyphenated with triple quadrupole liquid chromatography mass spectrometry has only 2 min chromatographic runtime for the analysis of all the pesticides. All the factors affecting the extraction yield have been optimized using an experimental design; and under optimized conditions, the developed method has been validated. The detection limits for all the pesticides were in the range of 0.001-0.348 µg/L with good linearity in the concentration range of 0.01-80.0 µg/L. The coefficient of determination was in the range of (R2 ) ≥ 0.977 to 0.999 for all the pesticides. The method was also checked for the precision of the relative standard deviation, which was below 4.75 (intra-day) and 8.96% (inter-day). The recovery of the method was 92-138%.
Assuntos
Cerveja/análise , Bebidas/análise , Sucos de Frutas e Vegetais/análise , Praguicidas/isolamento & purificação , Vinho/análise , Cromatografia Líquida , Espectrometria de Massas , Praguicidas/químicaRESUMO
The study aimed to investigate the role of NO and neuronal NO synthase (nNOS) in Zn-induced neurodegeneration. Animals were treated with zinc sulfate (20 mg/kg), twice a week, for 2-12 weeks along with control. In a few sets, animals were also treated with/without a NO donor, sodium nitroprusside (SNP), or S-nitroso-N-acetyl penicillamine (SNAP) for 12 weeks. Moreover, human neuroblastoma (SH-SY-5Y) cells were also employed to investigate the role of nNOS in Zn-induced toxicity in in vitro in the presence/absence of nNOS inhibitor, 7-nitroindazole (7-NI). Zn caused time-dependent reduction in nitrite content and total/nNOS activity/expression. SNP/SNAP discernibly alleviated Zn-induced neurobehavioral impairments, dopaminergic neurodegeneration, tyrosine hydroxylase (TH) expression, and striatal dopamine depletion. NO donors also salvage from Zn-induced increase in lipid peroxidation (LPO), mitochondrial cytochrome c release, and caspase-3 activation. While Zn elevated LPO content, it attenuated nitrite content, nNOS activity, and glutathione level along with the expression of TH and nNOS in SH-SY-5Y cells. 7-NI further augmented Zn-induced changes in the cell viability, oxidative stress, and expression of TH and nNOS. The results obtained thus demonstrate that Zn inhibits nNOS that partially contributes to an increase in oxidative stress, which subsequently leads to the nigrostriatal dopaminergic neurodegeneration.
Assuntos
Neurônios Dopaminérgicos/enzimologia , Neurônios Dopaminérgicos/patologia , Neostriado/patologia , Degeneração Neural/enzimologia , Degeneração Neural/patologia , Óxido Nítrico Sintase Tipo I/metabolismo , Zinco/efeitos adversos , Animais , Comportamento Animal/efeitos dos fármacos , Western Blotting , Caspase 3/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Citocromos c/metabolismo , Ativação Enzimática/efeitos dos fármacos , Glutationa/metabolismo , Humanos , Indazóis/farmacologia , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Metaboloma/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Doadores de Óxido Nítrico/farmacologia , Nitritos/metabolismo , Nitroprussiato/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Ratos Wistar , S-Nitroso-N-Acetilpenicilamina/farmacologia , Tirosina 3-Mono-Oxigenase/metabolismoRESUMO
Clinical evidences showing zinc (Zn) accumulation in the post-mortem brain of Parkinson's disease (PD) patients and experimental studies on rodents chronically exposed to Zn suggested its role in PD. While oxidative stress is implicated in Zn-induced neurodegeneration, roles of inflammation and apoptosis in degeneration of the nigrostriatal dopaminergic neurons have yet been elusive. The present study investigated the contribution of the nuclear factor kappa B (NF-κB), tumor necrosis factor-alpha (TNF-α), interleukin-1ß (IL-1ß), and B-cell lymphoma 2 (Bcl-2) family proteins in Zn-induced Parkinsonism. Male Wistar rats were treated with/without zinc sulfate (Zn; 20 mg/kg, intraperitoneally), twice a week, for 2-12 weeks. In a few sets, animals were treated intraperitoneally with a NF-κB inhibitor, pyrrolidine dithiocarbamate (PDTC; 100 mg/kg), a TNF-α inhibitor, pentoxyfylline (PTX; 50 mg/kg), and an anti-inflammatory agent, dexamethasone (DEX; 5 mg/kg), prior to Zn exposure along with respective controls. Zn caused neurobehavioral impairments and reduction in dopamine and its metabolites, tyrosine hydroxylase (TH)-positive neurons, catalase activity, and expression of TH, Bcl-2, and NOXA. On the contrary, Zn augmented lipid peroxidation, activity of superoxide dismutase, expression of TNF-α, IL-1ß, Bcl-xl, and p53-upregulated modulator of apoptosis (PUMA), and translocation of NF-κB and Bax from the cytosol to the nucleus and mitochondria, respectively, with concomitant increase in the mitochondrial cytochrome c release and activation of procaspase-3 and -9. Pre-treatment with PTX, DEX, or PDTC invariably ameliorated Zn-induced changes in behavioral and neurodegenerative indexes, inflammatory mediators, and apoptosis. Results demonstrate that inflammation regulates Bax expression that subsequently contributes to the nigrostriatal dopaminergic neurodegeneration.
Assuntos
Apoptose/efeitos dos fármacos , Neurônios Dopaminérgicos/patologia , Inflamação/patologia , Neostriado/patologia , Degeneração Neural/patologia , Substância Negra/patologia , Zinco/farmacologia , Proteína X Associada a bcl-2/metabolismo , Animais , Antígenos Nucleares/metabolismo , Apoptose/genética , Proteínas Reguladoras de Apoptose/genética , Proteínas Reguladoras de Apoptose/metabolismo , Comportamento Animal , Citocromos c/metabolismo , Dopamina/metabolismo , Neurônios Dopaminérgicos/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Inflamação/metabolismo , Interleucina-1beta/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Metaboloma , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , NF-kappa B/metabolismo , Degeneração Neural/genética , Proteínas do Tecido Nervoso/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/genética , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Ratos Wistar , Serotonina/metabolismo , Fator de Necrose Tumoral alfa/metabolismo , Tirosina 3-Mono-Oxigenase/metabolismo , Proteína bcl-X/genética , Proteína bcl-X/metabolismoRESUMO
The study aimed to measure the neuroprotective efficacy of caffeine-encapsulated poly(lactic-co-glycolic acid) (PLGA) nanoparticles over bulk and to delineate the mechanism of improvement in efficacy both in vitro and in 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mouse model of Parkinsonism. Caffeine-encapsulated PLGA nanoparticles exhibited more pronounced increase in the endurance of dopaminergic neurons, fibre outgrowth and expression of tyrosine hydroxylase (TH) and growth-associated protein-43 (GAP-43) against 1-methyl-4-phenylpyridinium (MPP+)-induced alterations in vitro. Caffeine-encapsulated PLGA nanoparticles also inhibited MPP(+)-mediated nuclear translocation of nuclear factor-kappa B (NF-κB) and augmented protein kinase B phosphorylation more potentially than bulk counterpart. Conversely, MPTP reduced the striatal dopamine and its metabolites and nigral TH immunoreactivity whereas augmented the nigral microglial activation and nigrostriatal lipid peroxidation and nitrite content, which were shifted towards normalcy by caffeine. The modulations were more evident in caffeine-encapsulated PLGA nanoparticles treated animals as compared with bulk. Moreover, the striatal caffeine and its metabolites were found to be significantly higher in caffeine-encapsulated PLGA nanoparticles-treated mice as compared with bulk. The results thus suggest that nanotization improves the protective efficacy of caffeine against MPTP-induced Parkinsonism owing to enhanced bioavailability, inhibition of the nuclear translocation of NF-κB and activation of protein kinase B phosphorylation.
Assuntos
1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/efeitos adversos , Cafeína/química , Cafeína/farmacologia , Portadores de Fármacos/química , Nanopartículas/química , Transtornos Parkinsonianos/induzido quimicamente , Transtornos Parkinsonianos/prevenção & controle , Transporte Ativo do Núcleo Celular/efeitos dos fármacos , Animais , Transporte Biológico , Cafeína/metabolismo , Contagem de Células , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Sobrevivência Celular/efeitos dos fármacos , Dopamina/metabolismo , Neurônios Dopaminérgicos/efeitos dos fármacos , Neurônios Dopaminérgicos/metabolismo , Neurônios Dopaminérgicos/patologia , Liberação Controlada de Fármacos , Fluoresceína-5-Isotiocianato/química , Proteína GAP-43/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Ácido Láctico/química , Peroxidação de Lipídeos/efeitos dos fármacos , Masculino , Camundongos , Microglia/efeitos dos fármacos , Microglia/patologia , NF-kappa B/genética , NF-kappa B/metabolismo , Fármacos Neuroprotetores/química , Fármacos Neuroprotetores/metabolismo , Fármacos Neuroprotetores/farmacologia , Nitritos/metabolismo , Transtornos Parkinsonianos/metabolismo , Transtornos Parkinsonianos/patologia , Fosfoproteínas/metabolismo , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismoRESUMO
Sustained and safe delivery of dopamine across the blood brain barrier (BBB) is a major hurdle for successful therapy in Parkinson's disease (PD), a neurodegenerative disorder. Therefore, in the present study we designed neurotransmitter dopamine-loaded PLGA nanoparticles (DA NPs) to deliver dopamine to the brain. These nanoparticles slowly and constantly released dopamine, showed reduced clearance of dopamine in plasma, reduced quinone adduct formation, and decreased dopamine autoxidation. DA NPs were internalized in dopaminergic SH-SY5Y cells and dopaminergic neurons in the substantia nigra and striatum, regions affected in PD. Treatment with DA NPs did not cause reduction in cell viability and morphological deterioration in SH-SY5Y, as compared to bulk dopamine-treated cells, which showed reduced viability. Herein, we report that these NPs were able to cross the BBB and capillary endothelium in the striatum and substantia nigra in a 6-hydroxydopamine (6-OHDA)-induced rat model of PD. Systemic intravenous administration of DA NPs caused significantly increased levels of dopamine and its metabolites and reduced dopamine-D2 receptor supersensitivity in the striatum of parkinsonian rats. Further, DA NPs significantly recovered neurobehavioral abnormalities in 6-OHDA-induced parkinsonian rats. Dopamine delivered through NPs did not cause additional generation of ROS, dopaminergic neuron degeneration, and ultrastructural changes in the striatum and substantia nigra as compared to 6-OHDA-lesioned rats. Interestingly, dopamine delivery through nanoformulation neither caused alterations in the heart rate and blood pressure nor showed any abrupt pathological change in the brain and other peripheral organs. These results suggest that NPs delivered dopamine into the brain, reduced dopamine autoxidation-mediated toxicity, and ultimately reversed neurochemical and neurobehavioral deficits in parkinsonian rats.
Assuntos
Barreira Hematoencefálica/metabolismo , Dopamina/química , Dopamina/metabolismo , Nanopartículas/química , Doença de Parkinson/tratamento farmacológico , Doença de Parkinson/fisiopatologia , Animais , Linhagem Celular Tumoral , Dopamina/efeitos adversos , Neurônios Dopaminérgicos/metabolismo , Portadores de Fármacos/efeitos adversos , Portadores de Fármacos/química , Portadores de Fármacos/metabolismo , Liberação Controlada de Fármacos , Humanos , Ácido Láctico/química , Neostriado/efeitos dos fármacos , Neostriado/metabolismo , Oxirredução , Oxidopamina/química , Oxidopamina/farmacologia , Oxidopamina/uso terapêutico , Doença de Parkinson/metabolismo , Ácido Poliglicólico/química , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Ratos , Ratos Wistar , Receptores Dopaminérgicos/metabolismo , Segurança , Regulação para Cima/efeitos dos fármacosRESUMO
A 1,1,1-trichloro-2,2-bis(4-chlorophenyl)ethane (DDT) degrading bacterium strain IITR03 producing trehalolipid was isolated and characterized from a pesticides contaminated soil. The strain IITR03 was identified as a member of the genus Rhodococcus based on polyphasic studies. Under aqueous culture conditions, the strain IITR03 degraded 282 µM of DDT and could also utilize 10mM concentration each of 4-chlorobenzoic acid, 3-chlorobenzoic acid and benzoic acid as sole carbon and energy source. The catechol 1,2-dioxygenase enzyme activity resulted in conversion of catechol to form cis,cis-muconic acid. Cloning and sequencing of partial nucleotide sequence of catechol 1,2-dioxygenase gene (cat) from strain IITR03 revealed its similarity to catA gene present in Rhodococcus sp. strain Lin-2 (97% identity) and Rhodococcus strain AN22 (96% identity) degrading benzoate and aniline, respectively. The results suggest that the strain IITR03 could be useful for field bioremediation studies of DDT-residues and chlorinated aromatic compounds present in contaminated sites.
Assuntos
DDT/metabolismo , Glicolipídeos/biossíntese , Rhodococcus/isolamento & purificação , Rhodococcus/metabolismo , Biodegradação Ambiental/efeitos dos fármacos , Catecol 1,2-Dioxigenase/genética , Catecol 1,2-Dioxigenase/metabolismo , Ácidos Graxos/análise , Cromatografia Gasosa-Espectrometria de Massas , Dados de Sequência Molecular , Fenótipo , Filogenia , Ácido Pirúvico/farmacologia , RNA Ribossômico 16S/genética , Padrões de Referência , Rhodococcus/enzimologia , Rhodococcus/crescimento & desenvolvimento , Espectrofotometria Ultravioleta , Ácido Succínico/farmacologiaRESUMO
For some instances of Parkinson disease (PD), current evidence in the literature is consistent with reactive oxygen species being involved in the etiology of the disease. The management of PD is still challenging owing to its ambiguous etiology and lack of permanent cure. Because nicotine offers neuroprotection against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine-induced parkinsonism, the neuroprotective efficacy of nicotine-encapsulated poly(lactic-co-glycolic) acid (PLGA) nanoparticles and the underlying mechanism of improved efficacy, if any, over bulk nicotine were assessed in this study. The selected indicators of oxidative stress, dopaminergic neurodegeneration and apoptosis, were measured in both in vitro and rodent models of parkinsonism in the presence or absence of "nanotized" or bulk nicotine. The levels of dopamine and its metabolites were measured in the striatum, nicotine and its metabolite in the nigrostriatal tissues while the immunoreactivities of tyrosine hydroxylase (TH), metallothionein-III (MT-III), inducible nitric oxide synthase (iNOS) and microglial activation were checked in the substantia nigra of controls and treated mice. GSTA4-4, heme oxygenase (HO)-1, tumor suppressor protein 53 (p53), caspase-3, lipid peroxidation (LPO), and nitrite levels were measured in the nigrostriatal tissues. Nicotine-encapsulated PLGA nanoparticles improved the endurance of TH-immunoreactive neurons and the number of fiber outgrowths and increased the mRNA expression of TH, neuronal cell adhesion molecule, and growth-associated protein-43 over bulk against 1-methyl-4-phenyl pyridinium ion-induced degeneration in the in vitro model. MPTP reduced TH immunoreactivity and levels of dopamine and its metabolites and increased microglial activation, expression of GSTA4-4, iNOS, MT-III, HO-1, p53, and caspase-3, and levels of nitrite and LPO. Whereas both bulk nicotine and nicotine-encapsulated PLGA nanoparticles modulated the changes toward controls, the modulation was more pronounced in nicotine-encapsulated PLGA nanoparticle-treated parkinsonian mice. The levels of nicotine and cotinine were elevated in nicotine-encapsulated PLGA nanoparticle-treated PD mouse brain compared with bulk. The results obtained from this study demonstrate that nanotization of nicotine improves neuroprotective efficacy by enhancing its bioavailability and subsequent modulation in the indicators of oxidative stress and apoptosis.
Assuntos
Encéfalo/efeitos dos fármacos , Nanoconjugados , Fármacos Neuroprotetores/administração & dosagem , Nicotina/administração & dosagem , Transtornos Parkinsonianos/patologia , Animais , Western Blotting , Encéfalo/patologia , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Cromatografia Líquida de Alta Pressão , Citometria de Fluxo , Imuno-Histoquímica , Ácido Láctico/farmacologia , Camundongos , Neurônios/efeitos dos fármacos , Neurônios/patologia , Estresse Oxidativo/efeitos dos fármacos , Ácido Poliglicólico/farmacologia , Copolímero de Ácido Poliláctico e Ácido Poliglicólico , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Dolomite is a natural mineral of great industrial importance and used worldwide, thus millions of workers are at risk of occupational exposure. Its toxicity is however, meagerly documented. In the present investigation, a dolomite powder obtained from its milling unit was analyzed by some standard methods namely, optical microscopy, transmission electron microscopy and dynamic light scattering. Results showed that dolomite powder contained particles of different shapes and size both microparticles (MPs) and nanoparticles (NPs), suggesting potential occupational exposure of these particles. An attempt was therefore, made to investigate dolomite toxicity in a particle size-dependent manner in human lung epithelial cells A(549). The comparative toxicity evaluation of MPs and NPs was carried out by assessing their effects on cell viability, membrane damage, glutathione, reactive oxygen species (ROS), lipid peroxidation (LPO), micronucleus (MN) and proinflammatory cytokines, namely tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß) and interleukin-6 (IL-6). These markers of cytotoxicity, genotoxicity and inflammation were assayed in cells exposed to MPs and NPs in a dose-and time-dependent manner. Invariably, their toxic effects were dose-and time-dependent while NPs in general were significantly more toxic. Notably, NPs caused oxidative stress, genotoxicity and inflammatory responses, as seen by significant induction of ROS, LPO, MN, TNF-α, IL-1ß and IL-6. Thus, the study tends to suggest that separate health safety standards would be required for micrometer and nanometer scale particles of dolomite.
Assuntos
Carbonato de Cálcio/química , Carbonato de Cálcio/toxicidade , Magnésio/química , Magnésio/toxicidade , Nanopartículas/toxicidade , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Citocinas/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Glutationa/metabolismo , Humanos , L-Lactato Desidrogenase/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Pulmão/citologia , Testes para Micronúcleos , Estresse Oxidativo , Tamanho da Partícula , Pós , Espécies Reativas de Oxigênio/metabolismoRESUMO
An association between excessive zinc (Zn) accumulation in brain and incidences of Parkinson's 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.