Synergistic effects of lipopolysaccharide and rote-none on dopamine neuronal damage in rats / 中国药理学与毒理学杂志

OBJECTIVE The etiology of Parkinson disease(PD)is still unknown.Until now,oxidative stress and neuroinflammation play a crucial role in the patho-genesis of PD.However,the specific synergistic role of oxidative stress and neuroinflammation in the occurrence and development of PD remains unclear.METHODS The changes in motor behavior,dopamine(DA)neurons quantification and their mitochondrial respiratory chain,glial cells activation and secreted cytokines,Nrf2 signal-ing pathway,and redox balance in the brain of rats were evaluated.RESULTS Lipopolysaccharide(LPS)-induced neuroinflammation and rotenone(ROT)-induced oxidative stress synergistically aggravated motor dysfunc-tion,DA neuron damage,activation of glial cells,and release of related mediators,activation of Nrf2 signaling and destruction of oxidative balance.In addition,further studies indicated that after ROT-induced oxidative stress caused direct damage to DA neurons,LPS-induced inflammatory effects had stronger promoting neurotoxic effects on the above aspects.CONCLUSION Neuroinflammation and oxidative stress synergistically aggravated DA neuronal loss.Furtherly,oxidative stress followed by neuroinflammation caused more DA neuro-nal loss than neuroinflammation followed by oxidative stress.

Effects and mechanisms of SNP-9 on Parkinson''s disease cell model induced by rotenone / 中国药科大学学报

@#In this article, the effects and mechanisms of SNP-9 on Parkinson''s disease (PD) cell model were investigated.SH-SY5Y cells were treated with rotenone to establish PD cell model; the effects of rotenone and SNP-9 on cell viability were detected by MTT assay; Hoechst/PI double staining assay was used to detect the effects of rotenone and SNP-9 on cell apoptosis; DCFH-DA probe was used to detect the effects of rotenone and SNP-9 on cellular reactive oxygen species (ROS) levels; and Western blot was used to detect the effects of rotenone and SNP-9 on protein levels of tyrosine hydroxylase (TH), α-synuclein (α-syn), Bcl-2 and Bax.The results showed that SNP-9 could alleviate abnormalities in cell viability, levels of TH and α-syn, apoptosis, ROS and apoptotic relative protein Bax/Bcl-2 induced by rotenone.Our findings suggest that SNP-9 may alleviate rotenone-induced injury in neuronal cells by regulating cell apoptosis related pathway.

Effects of rotenone exposure during pregnancy on GSK-3β and β-catenin protein expression in rat placenta / 环境与职业医学

Background Research on non-target organ damage of biological pesticides has attracted much attention. Rotenone exposure may be far beyond the occupational environment, and the exposureduring pregnancy may be increased through bioaccumulation, fruit or vegetable residues, and other forms of oral intake. At present, the effects of rotenone on placental development and its mechanism are still unknown. Objective To investigate the developmental damage of rat placenta and evaluate the expression levels of glycogen synthase kinase (GSK-3β) and beta catenin (β-catenin) followed by rotenone exposure through the placental barrier during pregnancy, as well as to propose possible associated mechanisms. Methods Eighteen sexually mature SD female infertile rats without specific pathogens were selected and divided into three groups: blank control group (0.9% saline), corn oil group, and rotenone group (corn oil + 2 mg·kg−1 rotenone) by random number method, six female animals in each group. Another six male rats were selected and mated to the female rats at night with a female to male ratio of 3:1 per cage. Pregnant rats were given 0.9% saline, corn oil, and 2 mg·kg−1 rotenone preparation by isovolumetric gavage once daily for the entire gestation period (19 d), and their conditions were observed after the last dose. The pregnant rats were anesthetized, and the size of the placenta and blood perfusion were detected by ultrasound the next day of the last dose of rotenone. Then, 3 pregnant rats in each group were sacrificed immediately and the placenta and umbilical cord tissues were dissected. The remaining 9 pregnant rats gave birth naturally, and the fetuses were observed for developmental evaluation and weighed. The histopathological changes of umbilical cord and placenta were observed by hematoxylin-eosin staining. The reactive oxygen species levels of placenta tissues were detected by flow cytometry. The Ca2+-ATPase activity of placenta tissues was detected by colorimetric method. The localization and levels of GSK-3β and β-catenin expression of placenta were detected by immunohistochemistry. The p-GSK-3β/GSK-3β and p-β-catenin/β-catenin protein expression in placental tissues were measured by Western blotting. Results No injury or death was recorded during the pregnant rats receiving rotennon administration. Adverse pregnancy outcomes such as fetal absorption and postpartum stillbirth were found in the rotenone group, and the weight of the fetal mice decreased (P<0.05). The B-ultrasound showed disc-shaped placenta with a thick middle and thin edge, smooth fetal surface, rough maternal surface, visible placental lobules, granular echotexture of the placenta with comma-like echogenic densities, and chorionic plate showing deep indentations, no calcification, degeneration, or necrosis in each group. Compared with the corn oil group, the fetal surface diameter of the placenta was reduced in the rotenone group (P<0.05). The Doppler color ultrasound showed that interplacental blood flow was reduced in the rotenone group, while interplacental blood flow was abundant in the blank control and the corn oil groups. The hematoxylin-eosin staining results showed that smooth muscle cells in the umbilical cord tissues of rats were loosely arranged, with fuzzy nuclei and inflammatory infiltration in the rotenone group. The placental trophoblast cells were small in size, disorderly arranged with nuclear fragmentation and cytoplasm turbidity. The tissue reactive oxygen species level in the rotenone group was higher than those in the other two groups (P<0.05). The Ca2+-ATPase activity of placental tissues was reduced in the rotenone group (P<0.05). The immunofluorescence low-magnification observation showed that GSK-3β and β-catenin were expressed in placental tissue, weak fluorescence expression in the decidua basalis, strong fluorescence expression in the labyrinthine layer structure. The labyrinthine layer under high magnification showed that compared with the blank control group and the corn oil group, the brightness of β-catenin fluorescence expression in the rotenone group decreased (P<0.05), and the brightness of GSK-3β expression increased (P<0.05). The Western blotting results showed that the expression of β-catenin and p-GSK-3β proteins decreased (P<0.01), and the expression of GSK-3β protein increased (P<0.01) in the rotenone group. No significant expression of p-β-catenin protein was detected in the placenta tissue of each group. Conclusion Rotenone exposure during pregnancy induces placental hypoperfusion, growth retardation, and oxidative stress in rats, as well as down-regulation of β-catenin and p-GSK-3β protein expression, and up-regulation of GSK-3β protein expression, which may further lead to abnormal pregnancy and fetal restricted growth.

Neuroprotective effects of sinapic acid involve the iron regulatory role on the rotenone-induced Parkinson's disease model

Abstract In the last decades, ferroptosis and its relationship with Parkinson's disease have gained significant attention. Compounds that affect ferroptosis and iron-dependent pathways in particular, have possible candidates for study in this context.Sinapic acid is an iron-chelator and high antioxidant bioactive phenolic acid. Its neuroprotective action, due to the antioxidant capacity, has been shown in several experimental models.However, the relationship between iron and antioxidant actions is still misunderstood and therefore, in the current study, we tried to investigate the effects of sinapic acid in rotenone-induced Parkinson's disease with the aspect of ferroptosis and iron-dependent alterations.The Parkinson's disease model was induced by a single dose intrastriatal and intrategmental rotenone (5µg/µl) injection.Sinapic acid (30mg/ kg) was orally administered during a 28-day period after the Parkinson's disease model was validated.Our results demonstrated that sinapic acid treatment attenuated rotenone-induced increase of serum transferrin and iron levels.Furthermore, sinapic acid inhibited rotenone-induced heme oxygenase-1(HO-1) increase and decrease of glutathione peroxidase-4 (GPx-4) levels in brain tissue. Also, sinapic acid treatment decreased motor impairment, likely as a result of the ameliorative effects on the tyrosine hydroxylase immunoreactivity loss after the rotenone insult.Our study suggests that the iron regulatory role of sinapic acid possibly plays a role in the protective effect on rotenone-induced neuronal damage.

Effects of VHL inhibitor on rotenone-induced Caenorhabditis elegans model of Parkinson′s disease / 中国药科大学学报

@#To investigate the effects of VHL (von Hippel-Lindau) inhibitor on Caenorhabditis elegans (C.elegans) model of Parkinson''s disease (PD),C.elegans were exposed to rotenone and treated with VHL inhibitor VH298.The death,dopaminergic neurodegeneration and mitochondrial unfolded protein response (mito-UPR) of transgenic strains with the markers zcIs9 and otIs181 exposed to different concentrations of rotenone were investigated. The death,dopaminergic neurodegeneration,and changes of behaviors including head thrashes,body bends and foraging behavior of C.elegans model of PD treated with different concentrations of VH298 were explored.The results showed that different concentrations of rotenone can lead to the death,dopaminergic neurodegeneration and abnormal mito-UPR of transgenic nematodes with zcIs9; otIs181,while the VHL inhibitor can decrease the death rate and alleviate dopaminergic neurodegeneration of rotenone-induced C.elegans model of PD.The VHL inhibitor can also attenuate the behavioral abnormalities of head thrashes,body bends and foraging behavior of C.elegans model.These results suggest that rotenone may cause mitochondrial damage in the transgenic nematodes with zcIs9; otIs181, and then destroy mitochondrial homeostasis,thereby resulting in dopaminergic neurodegeneration and death of the nematodes. The VHL inhibitor VH298 may promote the survival of rotenone-induced C.elegans model of PD,and alleviate dopaminergic neurodegeneration,thereby improving the behavioral abnormalities of C.elegans model of PD.

Autophagy inhibitor 3-methyladenine could not modulate rotenone neurotoxicity in primary mesencephalic cell culture / El inhibidor de la autofagia 3-metiladenina no logra modular la neurotoxicidad de la rotenona en el cultivo primario de células mesencefálicas

Dysregulated autophagy, whether excessive or downregulated, has been thought to be associated with neurodegenerative disorders including Parkinson's disease. Accordingly, the present study was carried out to investigate whether 3-methyladenine, an autophagy inhibitor, can modulate the effects of rotenone on dopaminergic neurons in primary mesencephalic cell culture. Cultures were prepared from embryonic mouse mesencephala at gestation day 14. Four groups of cultures were treated on the 10th DIV for 48 h as follows: the first was kept as an untreated control, the second was treated with 3-methyladenine alone (1, 10, 100, 200 mM), the third was treated with 20 nM rotenone and the fourth was co-treated with 20 nM rotenone and 3-methyladenine (1, 10, 100, 200 mM). On the 12th DIV, cultured cells were stained immunohistochemically against tyrosine hydroxylase and culture media were used to measure the levels of lactate dehydrogenase. 3methyladenine had no effects on both the survival of dopaminergic neurons and the release of lactate dehydrogenase. Rotenone significantly decreased the number of dopaminergic neurons and increased the levels of lactate dehydrogenase in the culture media. When cultures concomitantly treated with 3-methyladenine and rotenone, 3-methyladenine had no effect against rotenone-induced dopaminergic cell damage and lactate dehydrogenase release into the culture medium. In conclusion, the autophagy inhibitor 3-methyladenine could not modulate rotenone-induced dopaminergic cell damage in primary mesencephalic cell culture.

Se estima que la autofagia desregulada, ya sea excesiva o con baja regulación, está asociada con trastornos neurodegenerativos, incluyendo la enfermedad de Parkinson. En consecuencia, el se realizó este estudio para investigar si la 3metiladenina, un inhibidor de la autofagia,puede modular los efectos de la rotenona en las neuronas dopaminérgicas en el cultivo primario de células mesencefálicas. Los cultivos se prepararon a partir de mesencéfalo de ratón embrionario el día 14 de gestación. Cuatro grupos de cultivos se trataron en el 10º DIV durante 48 h de la siguiente manera: el primer grupo se mantuvo como un control no tratado, el segundo se trató con 3-metiladenina sola (1, 10, 100, 200 mM), el tercer grupo se trató con rotenona 20 nM y el cuarto se trató conjuntamente con rotenona 20 nM y 3-metiladenina (1, 10, 100, 200 mM). En el 12º DIV; las células cultivadas fueron tratadas mediante tinción inmunohistoquímica en tirosina hidroxilasa y se usaron medios de cultivo para medir los niveles de lactato deshidrogenasa. La 3-metiladenina no tuvo efectos tanto en la supervivencia de las neuronas dopaminérgicas como en la liberación de lactato deshidrogenasa. La rotenona disminuyó significativamente el número de neuronas dopaminérgicas y se observó un aumento de los niveles de lactato deshidrogenasa en los medios de cultivo. Cuando los cultivos tratados concomitantemente con 3-metiladenina y rotenona, la 3metiladenina no tuvo efecto contra el daño celular dopaminérgico inducido por la rotenona y la liberación de lactato deshidrogenasa en el medio de cultivo. En conclusión, el inhibidor de la autofagia 3-metiladenina no moduló el daño celular dopaminérgico inducido por la rotenona en el cultivo celular mesencefálico primario.

Anti-proliferative potential of sodium thiosulfate against HT 29 human colon cancer cells with augmented effect in the presence of mitochondrial electron transport chain inhibitors

Objective: To compare the anti-proliferative effect of sodium thiosulfate on human colorectal cancer cells (HT-29) and normal small intestine cells (IEC6). Methods: Cells (HT-29 and IEC6) were treated with different concentrations of sodium thiosulfate ranging from 0.5 mM to 80 mM for 24 h. Cell viability was measured via crystal violet and MTT assays. HT-29 cells were further treated in the presence and absence of mitochondrial electron transport chain (ETC) inhibitors, KATP channel opener and closer and H2S inhibitors for 24 h followed by sodium thiosulfate in order to study their respective roles in the anti-proliferative activity of sodium thiosulfate. Results: The IC50 values of sodium thiosulfate on HT-29 cells were 40.93 mM and 42.45 mM by crystal violet and MTT assay whereas, in the case of IEC6 cells, the values were 45.17 mM and 47.22 mM. The inhibition of endogenous H2S enzymes and KATP channel induced no change in the anti-proliferative capacity of sodium thiosulfate. However, the anti-proliferative activity of sodium thiosulfate was enhanced in the presence of mitochondrial ETC inhibitors. Conclusions: HT-29 cell growth is effectively attenuated by sodium thiosulfate and the anti-proliferative activity of sodium thiosulfate is enhanced in the presence of mitochondrial ETC inhibitors.

Neuroprotective effects of kukoamine A against rotenone-induced neurotoxicity in PC12 cells / 中草药

Objective: To investigate the neuroprotective effects of kukoamine A (KuA) on rotenone-induced PC12 cells damage and to preliminary verify its potential action mechanisms. The present study may lay the foundation for finding leading compounds with anti-Parkinson's disease (PD) effects. Methods: A PD model induced by rotenone was established in vitro, and MTT, LDH, and Hoechst33342 staining were used for preliminary confirmation of KuA resistance to rotenone-induced PC12 cell injury in vitro. The effects of KuA on superoxide dismutase (SOD) activity, malondialdehyde (MDA) and reactive oxygen species (ROS) content, and mitochondrial membrane potential (MMP) were investigated by colorimetric method and fluorescence staining, respectively. Western blotting was applied to explore the underlying mechanisms of protective effects of KuA against rotenone-induced PC12 cells damage. Results: The PC12 cell viability was significantly decreased after exposure to 0.5 μmol/L rotenone, whereas pretreatment with different concentrations of KuA could attenuate the cell injury induced by rotenone. Compared with the rotenone-treated group, KuA could decrease the ROS production and MDA level, while increase the SOD activity. In addition, KuA could effectively increase the MMP, decrease the cytochrome c release and the Bax/Bcl-2 ratio as well as inhibit caspase-3, caspase-9, and α-synuclein protein expressions. Conclusion: KuA showed neuroprotective ability on rotenone-induced PC12 cells PD model and the potential protective mechanisms of KuA can be related with inhibition of ROS generation, protection of MMP, regulation of protein expressions involved in the mitochondrial apoptosis pathway and reduction of α-synuclein expression.

Protective Effect and Mechanism of Velvet Antler Peptide on Rotenone-induced SH-SY5Y Cell Damage / 中国实验方剂学杂志

Objective:To investigate the intervening effect of velvet antler peptide （VAP） on rotenone-induced neuroblastoma （SH-SY5Y） cell damage and explore its related mechanism. Method:0.5 μmol·L-1 rotenone was used to SH-SY5Y cells to establish an in vitro model of Parkinson's disease （PD）. A blank control group， a model group， high， medium and low dose VAP groups （150，100，50 mg·L-1， respectively） and a rapamycin group were established. The number of lewy bodies， changes in mitochondrial membrane potential， content of reactive oxygen species （ROS） and α-synuclein （α-syn）， protein kinase B （Akt）， and mammalian target of rapamycin （mTOR） were observed by hematoxylin-eosin（HE） staining， rhodamine 123 staining， DCFH-DA staining and immunohistochemical staining expression respectively. Result:The results of HE staining showed that as compared with the blank group， the number of cells in model group was reduced， the tentacle structure became dull， the shape became round， and eosinophilic Lewy bodies were visible in cytoplasm. As compared with model group， there was no significant difference in cell morphology from rapamycin group and VAP high， medium and low dose groups， but there were fewer Lewy bodies in cytoplasm in these four groups. Rhodamine 123 staining showed that as compared with blank group， the mitochondrial membrane potential was increased significantly in model group （P<0.05）. As compared with the model group， the mitochondrial membrane potential was decreased in rapamycin group and VAP high， medium and low dose groups （P<0.05）. DCFH-DA staining results showed that as compared with blank group， the content of ROS was increased significantly in cells of model group （P<0.05）. As compared with model group， the content of ROS was decreased in rapamycin group and VAP high， medium and low dose groups （P<0.05）. Immunohistochemical staining showed that as compared with blank group， the protein expression levels of α-syn，Akt，and mTOR were increased significantly in model group （P<0.05）. As compared with model group， the protein expression levels of α-syn and mTOR were significantly reduced in rapamycin group and VAP high and medium dose groups （P<0.05）， and the expression levels of Akt were significantly reduced in rapamycin group and VAP high-dose group （P<0.05）. Conclusion:Velvet antler peptides may play a neuroprotective role by regulating the Akt/mTOR signaling pathway and promoting the degradation of α-syn in SH-SY5Y cells.

Isorhamnetin prevents pc12 cell from rotenone-induced iiyury via pdk/akt/gsk-3p/creb signaling pathway / 中国药理学通报

Aim To investigate whether isorhamnetin could protect against rotenone-induced PC 12 cells injury via regulating PI3K/Akt/GSK-3p/CREB pathway. Methods PC 12 cell viability was determined by MTT and LDH assays. The expressions of Akt,p-Akt,GSK-3p,p-GSK-3p,CREB and p-CREB were measured by Western blot. Results The cell viability and level of phospho-CREB significantly decreased in PC 12 cells exposed to rotenone when compared to control group. Both the cell viability and the expression of phospho-CREB in cells pretreated with isorhamnetin were higher than those of cells exposed to rotenone alone. Moreover, pretreatment of PC 12 cells with isorhamnetin enhanced phosphorylation of Akt and GSK-3fi. The addition of LY294002 could suppress the phosphorylation of Akt, GSK-3p and CREB, resulting in abolishment of neuroprotective effects of isorhamnetin on cells exposed to rotenone. Conclusion Isorhamnetin might alleviate rotenone-induced PC 12 cells injury partially via the PI3K/Akt/GSK-3 (3/CREB signaling pathway.

Anti-proliferative potential of sodium thiosulfate against HT 29 human colon cancer cells with augmented effect in the presence of mitochondrial electron transport chain inhibitors

Objective: To compare the anti-proliferative effect of sodium thiosulfate on human colorectal cancer cells (HT-29) and normal small intestine cells (IEC6). Methods: Cells (HT-29 and IEC6) were treated with different concentrations of sodium thiosulfate ranging from 0.5 mM to 80 mM for 24 h. Cell viability was measured via crystal violet and MTT assays. HT-29 cells were further treated in the presence and absence of mitochondrial electron transport chain (ETC) inhibitors, K

Evaluation of Haematological Alterations in Intraperitoneal and Oral Rotenone Induced Parkinson’s Disease Wistar Rats

This study was aimed at developing a single RP-HPLC method for determination of Natamycin in 20 different cheesesamples purchased from local Turkish supermarkets. Chromatographic separation was achieved on a X-Terra RP18column (250 mm x 4.6 mm x 5 µm) with a mobile phase of acetonitrile: water (30:70 v/v/v), at 0.8 mL/min flow rate withDAD detection at 305 nm. In twenty cheese samples, Natamycin was analyzed by using sample preparation method of ISO9233-2, 2007 (IDF 140-2, 2007). The results of analysis have been fully validated statistically and recovery studiesconfirmed the accuracy of the proposed method. The precision (intra-day & inter-day) of method was found within limits(RSD < 2%). The sensitivity of the method was assessed by determining limit of detection and limit of quantification.Findings dealing with the presence of Natamycin in cheese samples are presented.

Treadmill Exercise Improves Motor Function by Suppressing Purkinje Cell Loss in Parkinson Disease Rats / 대한배뇨장애요실금학회지

PURPOSE: Rotenone is the most widely used neurotoxin for the making Parkinson disease (PD) animal model. The neurodegenerative disorder PD shows symptoms, such as slowness of movements, tremor at resting, rigidity, disturbance of gait, and instability of posture. We investigated whether treadmill running improves motor ability using rotenone-caused PD rats. The effect of treadmill running on PD was also assessed in relation with apoptosis of cerebellar Purkinje cells. METHODS: Treadmill running was applied to the rats in the exercise groups for 30 minutes once a day for 4 weeks, starting 4 weeks after birth. We used rota-rod test for the determination of motor coordination and balance. In this experiment, terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) staining, immunohistochemistry for calbindin, glial fibrillary acidic protein (GFAP), Iba-1, and western blot analysis for Bax and Bcl-2 were performed. RESULTS: Treadmill running enhanced motor balance and coordination by preventing the loss of Purkinje cells in the cerebellar vermis. Treadmill running suppressed PD-induced expression of GFAP-positive reactive astrocytes and Iba-1-positive microglia, showing that treadmill running suppressed reactive astrogliosis and microglia activation. Treadmill running suppressed TUNEL-positive cell number and Bax expression and enhanced Bcl-2 expression, demonstrating that treadmill running inhibited the progress of apoptosis in the cerebellum of rotenone-induced PD rats. CONCLUSIONS: Treadmill running improved motor ability of the rotenone-induced PD rats by inhibiting apoptosis in the cerebellum. Apoptosis suppressing effect of treadmill running on rotenone-induced PD was achieved via suppression of reactive astrocyte and inhibition of microglial activation.

Neuroprotection by misoprostol against rotenone-induced neurotoxicity in rat brain

Objective:To investigate the effect of the prostaglandin E1 analogue misoprostol on oxidative stress and neurodegeration caused by subcutaneous rotenone administration in rats.Methods:Rotenone was administered in a dose of 1.5 mg/kg every other day for 2 weeks. Starting from the 1st day of rotenone injection, rats were subcutaneously treated with misoprostol at doses of 10, 100 or 1 000 μ g/kg. Rats were evaluated for brain lipid peroxidation (malondialdehyde: MDA), reduced glutathione (GSH), nitric oxide (NO) levels, and paraoxonase-1 (PON-1) activity. The concentrations of the anti-apoptotic protein B cell/lymphoma-2 (Bcl-2) were determined in the striatum. Histopathologic examination and the expression of inducible nitric oxide synthase (iNOS) in the cerebral cortex and striatum were also performed.Results:Compared with the vehicle-treated group, rotenone caused a significant increase in brain lipid proxidation (MDA) by 61% (P<0.05) accompanied by an increase in NO by 73.1% (P<0.05) and a decrease in GSH concentration by 29.4% (P<0.05). In addition, brain PON-1 activity significantly decreased by 63.0% (P<0.05) and striatal Bcl-2 significantly decreased by 27.9% (P<0.05) with respect to the corresponding control value. Brain sections from rotenone treated rats showed extensive dark pyknotic and apoptotic nuclei in neurons, shrunken cytoplasm and perineuronal vacuolation. Rotenone also caused pronounced expression of iNOS in the cerebral cortex and striatum. Treatment with misoprostol at doses of 100 and 1 000 μ g/kg resulted in decreased brain MDA (by 16.5%-23.0%) (P<0.05) and NO levels (by 37.1%-40.7%) (P<0.05) and increased GSH concentrations (by 18.8%-30.1%) (P<0.05). PON-1 activity was significantly increased by 80.0%-114.8% (P<0.05) by misoprostol at 100 and 1 000 μ g/kg, respectively. In addition, misoprostol treatment restored striatal Bcl-2 concentrations to its normal value. Misoprostol treatment resulted in markedly reduced brain injury and decreased iNOS expression in the cerebral cortex and striatum of rotenone intoxicated rats.Conclusions:These data suggest that misoprostol prevents the rotenone-induced neurodegeneration in rat brain by reducing brain oxidative stress.

Influence of Rotenone on Activity of Protein Phosphates 2A in MN9D Cells / 中国康复理论与实践

Objective To study whether and how rotenone reduces the activity of protein phophatase 2A (PP2A). Methods MN9D cells (mouse midbrain dopaminergic cell line) were divided into normal group (normal cultured), con-trol group (dimethyl sulfoxide of same volume with rotenone group was added in medium), rotenone group (50 nmol/L rotenone was added to the culture medium for 24 hours) and rotenone+C2 group (pretreatment of 5 mol/L C2-ceramide for eight hours, and then exposed to 50 nmol/L rotenone for 24 hours). MTT was used to detect cell viability. Total PP2A levels and tyrosine phosphorylation levels were measured with Western blotting. PP2A activity was detected with colorimetric assay.Results Compared with the control group, the cell viability reduced (P<0.01), phosphorylation of tyrosine 307 of PP2A inceased (P<0.01), and activity of PP2A decreased in the rotenone group (P<0.05). And compared with the rote-none group, the cell viability improved (P<0.01), phosphorylation of tyrosine 307 of PP2A deceased (P<0.01), and activity of PP2A increased (P<0.05) in the rotenone+C2 group. Conclusion Rotenone can inhibit activity of PP2A through increasing phosphorylation of tyrosine 307 at the catalytic subunit of PP2A, which might be involved in reducing cell viability, and implicate a new treatment strategy for Parkinson's disease.

Protective effect of water soluble CoQ10 on rotenone-induced apoptosis in PC12 cells / 西安交通大学学报(医学版)

Objective To investigate the protective effect and the underlying mechanism of water soluble coenzyme Q10 (CoQ10)against rotenone induced injury on PC12 cells model.Methods PC12 cells were cultured with rotenone,water-soluble CoQ1 0 was added to the culture media 3 hours prior to the rotenone incubation.We determined cell viability by CCK8;reactive oxygen species (ROS)was detected by spectrophotometer;and Bcl-2, Bax,active Caspase-3,Caspase-9 and apoptosis-inducing factor (AIF)were measured by Western blotting after 24-hour rotenone incubation.Results After the treatment by rotenone,cell viability decreased significantly (P＜0.01)and ROS level increased (P＜0.01).CoQ10 could improve PC12 cell viability (P＜0.01)and reduce the level of ROS (P＜0.01).Western blotting experiments showed that CoQ10 could reduce rotenone-induced Caspase-9 (P＜0.05),active Caspase-3 (P＜0.05)and Bax (P＜0.01)expressions,increase the expression of Bcl-2 (P＜0.01),and prevent nuclear translocation of AIF (P＜0.05).Conclusion CoQ10 has a protective effect on rotenone-induced apoptosis in PC12 cells,the mechanism of which may be through scavenging ROS in cells;decreasing caspase-9 ,active caspase-3 and Bax expressions;and increasing the expression of Bcl-2 ;and preventing AIF nuclear translocation.

Preliminary optimization of a Chinese herbal medicine formula based on the neuroprotective effects in a rat model of rotenone-induced Parkinson's disease / 中西医结合学报

<p><b>OBJECTIVE</b>The main objective of this study was to preliminarily determine the optimum formulation of a Chinese herbal formula that may have neuroprotective effects against rotenone-induced Parkinson's disease (PD).</p><p><b>METHODS</b>Seven recipes were made from Dihuang (DH, Rehmannia glutinosa Libosch), Roucongrong (RCR, Cistanche deserticola Y.C.Ma), Niuxi (NX, Achyranthes bidentata Bl.) and Shanzhuyu (SZY, Cornus officinalis Sieb. et Zucc) in different proportions, according to the principles of uniform design (4 factors 7 levels). Tyrosine hydroxylase (TH)-positive neurons in substantia nigra pars compacta (SNpc) were detected by immunohistochemistry and rotenone-exposure days necessary to induce PD symptoms were recorded. To probe one likely mechanism of the formulas, echinacoside (ECH) concentrations of all seven recipes were determined by high-performance liquid chromatography and related to number of TH-positive neurons.</p><p><b>RESULTS</b>The data showed that recipe 4 (DH:RCR:SZY:NX = 1:1:1:1) and recipe 7 (DH:RCR:SZY:NX = 7:5:3:1) partially reversed rotenone-induced death of TH-positive neurons in the SNpc and significantly increased rotenone-exposed days compared with model group. Pharmacologically, there was not a strong correlation between ECH concentration and TH-positive neurons.</p><p><b>CONCLUSION</b>The investigated formulations of Chinese herbs had neuroprotective effects against PD models, and the neuroprotective effects were weakly related to the proportion of key herbs. However the neuroprotective effects of the formula may not result from a single active constituent.</p>

Neuroprotection by misoprostol against rotenone-induced neurotoxicity in rat brain

Objective: To investigate the effect of the prostaglandin E1 analogue misoprostol on oxidative stress and neurodegeration caused by subcutaneous rotenone administration in rats. Methods: Rotenone was administered in a dose of 1.5 mg/kg every other day for 2 weeks. Starting from the 1st day of rotenone injection, rats were subcutaneously treated with misoprostol at doses of 10, 100 or 1 000

Glucose promotes resistance in lymphocytes against oxidative stress-induced apoptosis through signaling and metabolic pathways. Implications for Parkinson's disease / La glucosa promueve la resistencia en linfocitos contra el estrés oxidativo que induce apoptosis a través de rutas de señalización y metabólica. Impacto en la enfermedad de Parkinson / A glucose promove a resistência em linfócitos contra o estresse oxidativo que induz apoptose a través de rotas de sinalização e metabólica. Impacto na doença de Parkinson

SUMMARY Introduction: Parkinson's disease (PD) is a neurological disorder associated with the selective loss of dopaminergic (DAergic) neurons. Clinical data suggest that oxidative stress (OS) and dysregulation of glucose (G) metabolism are early events in PD. However, no data are available to explain the molecular connection between glucose metabolism, OS, and neuronal demise in PD. Human lymphocytes share a similar dopaminergic signaling mechanism with DAergic neurons. Further, rotenone (ROT) is a mitochondrial complex I inhibitor that selectively induces apoptosis through OS in dopaminergic neurons and lymphocytes. Thus, to test the hypothesis that G metabolism and OS are linked in dopaminergic system toxicity and PD, human lymphocytes were cultured with ROT in the presence or absence of various concentrations of glucose. Objective: This study examines the response of human lymphocytes to glucose (11, 55, 166, 277, 555 mM G) in the absence or presence of ROT (250 microM). Methods: Light and fluorescence microscopy and immunocytochemistry techniques were used to evaluate morphological and biochemical changes in human lymphocytes. Results: 55 mM G was effective in suppressing ROT-induced apoptosis in lymphocytes via 5 pathways: (i) pentose phosphate pathway (PPP), (ii) glutathione (GSH) pathway, (iii) superoxide dismutase (SOD) and catalase (CAT) antioxidant systems, and (iv) Phosphoinositide 3-kinase (PI3-K) signaling. Additionally, we report for the first time that G rescued lymphocytes from ROT-induced apoptosis by (v) activating NF-kB and down-regulating p53 and caspase-3. Signaling (e.g., LY294002) and metabolic inhibitors of these pathways (e.g., dehydroepiandrosterone (DHEA), L-buthioninesulfoximine (BSO), 1,3-Bis (2-chloroethyl)-1-nitrosourea (BCNU), mercaptosuccinic acid (MS), 3-amino-1,2,4- triazole (AT), sodium diethyldithiocarbamate (DDC)) only partially reversed the protective effect of 55 mM G on lymphocytes exposed to ROT. Conclusion: These data suggest that high G simultaneously triggers cellular signaling and antioxidant systems to ensure global cell protection against stressful conditions in DAergic.

RESUMEN Introducción: la enfermedad de Parkinson (EP) es un trastorno neurológico asociado con la pérdida selectiva de neuronas dopaminérgicas (DAérgicas). Datos clínicos sugieren que el estrés oxidativo (EO) y la desregulación del metabolismo de la glucosa (G) son eventos tempranos en la EP. Sin embargo, no existe información que explique la posible asociación molecular entre el metabolismo de la glucosa, el EO y la muerte neuronal. Los linfocitos humanos comparten mecanismos de señalización DAérgicos comunes. Más aún, la rotenona (ROT) es un inhibidor que selectivamente induce apoptosis vía EO en neuronas DAérgicas y linfocitos. Para evaluar la hipótesis que el metabolismo de la G y el EO están asociados con la toxicidad del sistema DAérgicas y EP, se cultivaron linfocitos humanos con ROT en presencia o ausencia de varias concentraciones de G. Objetivo: este estudio examina la respuesta de los linfocitos a G (11, 55, 166, 277, 555 mM) en ausencia o presencia de ROT (250 microM). Métodos: se utilizaron técnicas de microscopía de luz y fluorescencia e inmunocitoquímica para evaluar los cambios morfológicos y bioquímicos de los linfocitos. Resultados: la G 55 mM fue eficaz en suprimir la apoptosis en linfocitos inducida por ROT vía activación de 5 rutas metabólicas: (i) la vía pentosa fosfato, (ii) la vía glutatión; (iii) los sistemas antioxidantes superóxido dismutasa (SOD) y catalasa (CAT); (iv) fosfoinositol 3 cinasa (PI3-K). Además, se observó por primera vez que la G rescata linfocitos de la apoptosis inducida por ROT vía (v) activación del factor nuclear kappa-B (NFkB) y por regulación a la baja de p53 y de la caspasa-3. Se demostró que los inhibidores de señalización (v.gr. LY294002) e inhibidores metabólicos (v.gr. DHEA, BSO, BCNU, MS, DCC) revierten parcialmente los efectos citoprotectores de la G 55 mM en linfocitos expuestos a ROT. Conclusión: estos hallazgos sugieren que la alta concentración de G induce simultáneamente sistemas de señalización y antioxidantes para asegurar la protección global de la célula contra condiciones estresantes en células DAérgicas.

RESUMO Introdução: a doença de Parkinson (PD) é uma desordem neurológica associada a perda seletiva de neurônios dopaminérgicos (Daergicas). Dados clínicos sugerem que o estresse oxidativo (EO) e a desregulação do metabolismo da glicose (G) são eventos iniciais no PD. No entanto, não há nenhuma informação que explica a possível associação molecular entre o metabolismo da glicose, EO e morte neuronal. Linfócitos humanos compartilham comum Daergicos mecanismos de sinalização. Além disso, a rotenona (ROT) é um inibidor que seletivamente induz apoptose através de EO nos neurônios Daergicas e linfócitos. Para avaliar a hipótese de que o metabolismo do G e EO estão associadas com a toxicidade do Daergicas e sistema de EP, linfócitos humanos com ROT foram cultivados na presença ou ausência de várias concentrações de G. Objetivo: este estudo analisa a resposta dos linfócitos a G (55 11, 166, 277, 555 mM) na ausência ou presença de podridão (250 microM). Métodos: luz e imunocitoquímica, microscopia de fluorescência foram utilizadas técnicas para avaliar as alterações morfológicas e bioquímicas de linfócitos. Resultados: a 55mM G foi eficaz em suprimir a apoptose em linfócitos induzidos por (ROT) a través da ativação de 5 vias metabólicas: (i) a través do fosfato de pentose, (ii) a través de glutationa; (iii) a superóxido dismutase de antioxidante (SOD) e catalase (CAT) sistemas; (iv) Phosphoinositide 3 cinase (PI3-K). Além disso, foi observada pela primeira vez que o G resgata a apoptose de linfócitos induzida por (ROT) via ativação (v) por fator nuclear kappa-B, (NF-kB) e por regulação por baixo de p53 e Caspase-3. Mostrou que inibidores (por exemplo, LY294002), inibidores de sinalização e metabólicos (por exemplo, DHEA, BSO, BCNU, MS, DCC) reverteu parcialmente os efeitos dos tilacoides do G 55 mM de linfócitos expostos à (ROT). Conclusão: estes resultados sugerem que a alta concentração de G induz simultaneamente antioxidantes e sistemas de sinalização para garantir a proteção global da célula de condições estressantes nas células Daergicas.

Effect of siRNA-interfered endogenous Kit6.2 gene in PC12 cells on rotenone-induced cell damage and signal pathway / 中华老年心脑血管病杂志

Objective To study the effect of siRNA-interfered endogenous Kir6.2 gene deletion on rotenone-induced cell damage and signal pathway according to the expression of siRNA-interfered endogenous Kir6.2 gene.Methods The expression of endogenous Kir6.2 gene in PC12 cells was detected by RT-PCR and Western bolt.The viability of PC12 cells was tested using WST-1.The expression of PKC and phosphorylated PKC in PC12 cells was detected by Western blot in negative control group,siRNA/Kir6.2 group,siRNA/Kir6.2 + rotenone group,siRNA/Kir6.2+ rotenone+PKC inhibitor group before and afer treatment with rotenone.Results RT-PCR showed that the interference was successful in siRNA/Kir6.2/group 1 and siRNA/Kir6.2/ group 2.Western blot displayed that the expression level of endogenous Kir 6.2 gene was significantly lower in siRNA/Kir6.2/ group 1 than in negative control group (0.55±0.07 vs 0.89±0.09,P＜ 0.05).WST-1 revealed that the viability of PC12 cells was significantly lower in siRNA/Kir6.2 group than in negative control group (P＜0.05).Western blot demonstrated that the expression of PKC and phosphorylated PKC was significantly lower in siRNA/Kir6.2+-rotenone+PKC inhibitor group than in the other three group (P＜0.05).Conclusion Endogenous Kir6.2 can protect PC12 cells against the toxicity of rotenone and plays an important role in regulating the rotenone-induced viability of PC12 cells by activating the phosphorylated PKC.