Neuroprotective effects of the stable nitroxide compound Tempol on 1-methyl-4-phenylpyridinium ion-induced neurotoxicity in the Nerve Growth Factor-differentiated model of pheochromocytoma PC12 cells

Nerve growth factor (NGF) differentiated pheochromocytoma PC12 cells exposed to 1-methyl-4-phenylpyridinium (MPP+) toxin were used as an in vitro pharmacological model of Parkinson's disease to examine the neuroprotective effects of 4-hydroxy-2,2,6,6-tetramethyl piperidine-n-oxyl (Tempol), a free radical scavenger and a superoxide dismutase-mimetic compound. MPP+-induced PC12 cell death was measured 72 h after exposure to 1.5 mM MPP+ by the release of lactate dehydrogenease, caspase-3 activation and stimulation of survival and stress mitogen-activated protein kinases. Exposure of PC12 cells to MPP+ activated ERK1 and ERK2 (forty-fold over control after 72 h), JNK1 and JNK2 (fourfold after 48 h) and p-38alpha (tenfold after 24 h). Pretreatment of PC12 cells with 500 microM Tempol, 1 h before induction of the MPP+ insult, reduced by 70% the release of LDH into the medium, inhibited caspase-3 activity by 30% and improved by 33% mitochondrial function, effects correlated with a 70% reduction in ERK1 and ERK2 phosphorylation activity. These findings support the neuroprotective effect of Tempol in the MPP+-induced PC12 cell death model and its use as a potential drug for treatment of Parkinson's disease.     

脂质立方液晶设计制备与表征评价研究进展

目的：研究伊拉地平（ ISR ）对1-甲基-4-苯基吡啶离子（ MPP＋）损伤的PC12细胞的保护作用及可能机制。方法MPP ＋处理PC12细胞建立帕金森病细胞模型;4-甲基偶氮唑蓝（ MTT）比色法检测细胞存活率;双氯荧光黄乙酸乙酯（ DCFH-DA）染色流式细胞术检测细胞内活性氧（ ROS）的生成;JC-1染色流式细胞术检测细胞线粒体膜电位（ MMP）。结果1 mmol · L－1MPP＋处理PC12细胞24 h后能明显抑制细胞生长（P＜0．01）;降低线粒体膜电位;ROS含量增加。2μmol· L－1伊拉地平预处理后, PC12细胞存活率显著增加（ P＜0．01）;线粒体膜电位升高;ROS生成减少。结论伊拉地平对MPP＋损伤的PC12细胞具有保护作用,其作用机制可能与维持线粒体正常膜电位,稳定线粒体功能,阻止线粒体氧化应激发生有关。     

毛蕊花苷对MPP~+诱导的SHSY5Y细胞凋亡的保护作用

目的观察肉苁蓉提取物毛蕊花苷对MPP+诱导的SHSY5Y细胞损伤的影响。方法用MTT法检测细胞存活率,以流式细胞仪检测细胞内活性氧的产生和线粒体膜电位的变化,以及细胞凋亡的发生,并用荧光酶标仪测定caspase-3的活性,蛋白印迹测定Bcl-2的表达水平。结果200μmol·L-1MPP+处理细胞24h降低细胞的存活率;诱导细胞发生凋亡,凋亡率达38.9%;细胞内活性氧水平及caspase-3的活性升高;而线粒体膜电位却明显降低。而预先给予0.1、1或者10mg·L-1浓度的毛蕊花苷处理细胞12h,可提高细胞存活率;流式细胞仪检测凋亡率分别降低到29.5%,15.3%和8.6%,而且细胞内活性氧的水平明显降低,并可逐渐恢复线粒体的高能量状态;caspase-3的活性不断降低,Bcl-2的表达水平增高,并呈现一定的剂量依赖性。结论毛蕊花苷能抑制MPP+诱导的SHSY5Y细胞凋亡,其神经细胞保护作用可能与其降低细胞内活性氧水平,维持线粒体膜电位的高能状态和抑制caspase-3的活性有关。     

利福平抑制MPP^＋所致PC12细胞凋亡的研究

目的探讨利福平对MPP＋（1-甲基4-苯基吡啶离子）诱导的分化大鼠嗜铬细胞瘤细胞株（ratphaeochmmocytoma,PC12）细胞活性、细胞形态、调亡率的影响及其影响的机制。方法（1）利用MPP^＋诱导分化PCI2细胞建立帕金森病细胞模型;（2）MTT法检测细胞活性,（3）Tunel原位末端标记法检测细胞形态及半定量细胞凋亡率,（4）流式细胞术检测caspase－3激活率。结果（1）MPP＋作用后,细胞生长活性明显受到抑制,凋亡细胞数量增多,调亡率增加,caspase－3激活率明显增高;（2）而经100、200和300μmol／L各浓度利福平预处理后,利福平各浓度组内细胞活性增高,细胞凋亡程度、凋亡率及caspase－3激活率降低,且与利福平作用浓度存在剂量一效应关系。结论利福平可抑制MPP＋所致PCI2细胞的凋亡,这一作用是通过caspase-3途径起作用的,且存在剂量一效应关系。     

Rosmarinic acid antagonized 1-methyl-4-phenylpyridinium (MPP+)-induced neurotoxicity in MES23.5 dopaminergic cells

Rosmarinic acid (RA) is a naturally occurring polyphenolic compound found in various plant families. We previously reported that RA exerted protective effects against 6-hydroxydopamine (6-OHDA)-induced neurotoxicity through antioxidative properties. In this study, we investigated whether RA could prevent effects of 1-methyl-4-phenylpyridinium (MPP(+))-induced insult in MES23.5 dopamineric cells. 1-Methyl-4-phenylpyridinium treatment decreased cell viability and dopamine content, as well as caused apoptotic morphological changes. 1-Methyl-4-phenylpyridinium-induced mitochondrial dysfunction, indicated by inhibition of activity associated with mitochondrial respiratory chain complex I, suggested mitochondrial transmembrane potential collapse and generation of reactive oxygen species. Decreased Bcl-2/Bax ratio and caspase 3 activation were also observed. Rosmarinic acid pretreatment restored the complex I activity of the mitochondrial respiratory chain and partially reversed the other damaging effects of MPP(+). Our results indicate that RA plays a neuroprotective role by ameliorating mitochondrial dysfunction against MPP(+)-induced cell apoptosis and suggest that RA has the potential to be considered an aid for prevention of Parkinson's disease.     

瓜子金皂苷己对MPP~+诱导PC12细胞凋亡的保护作用

目的观察瓜子金皂苷己(polygalasaponin F,PS-F)对1-甲基-4-苯基-吡啶离子(1-methyl-4-phenylpyridinium,MPP+)诱导的PC12细胞损伤的影响,并且探讨其作用机制。方法 MTT法检测细胞存活率,Annexin V/PI染色流式细胞术检测PC12细胞凋亡,JC-1染色倒置显微镜检测细胞线粒体膜电位(mitochondrial membrane potential,MMP),Western blot检测Caspase-3蛋白的水平。结果 500μmol.L-1MPP+作用PC12细胞48 h,能明显抑制细胞生长(P<0.01),诱导细胞发生凋亡,同时降低MMP,增加活性Caspase-3的蛋白水平。同时给予不同浓度PS-F处理,PC12细胞存活率增加(P<0.01);凋亡细胞量减少;MMP增高;活性Caspase-3蛋白水平降低(P<0.01)。结论 PS-F能抑制MPP+诱导的PC12细胞的凋亡,其作用机制可能与维持线粒体正常膜电位,稳定线粒体功能,降低活性Caspase-3蛋白水平有关。     

没食子儿茶素没食子酸酯对MPP~+诱导大鼠PC12细胞凋亡的拮抗作用

目的探讨没食子儿茶素没食子酸酯(EGCG)对MPP+诱导大鼠PC12细胞凋亡的拮抗作用。方法培养PC12细胞给予MPP+(900μmol.L-1)诱导细胞凋亡,实验分为6组:空白对照组、MPP+组、维生素E(10μmol.L-1)组和EGCG高(100μmol.L-1)、中(50μmol.L-1)、低(10μmol.L-1)剂量组。药物处理0.5h后,加入MPP+损伤细胞,4h后取细胞测定乳酸脱氢酶(LDH)漏出量,MTT法检测细胞活力,Hoechst33342荧光染色法和流式细胞术检测细胞凋亡,透射电镜观察凋亡细胞线粒体形态结构改变。结果MPP+处理后,细胞活力下降,LDH漏出量增加,细胞凋亡率增加,线粒体肿胀,出现空泡和嵴断裂等细胞凋亡征象。维生素E和不同剂量EGCG处理后,明显提高细胞活力,降低LDH漏出和细胞凋亡率,并减少MPP+引起的线粒体结构损伤。结论EGCG具有抑制MPP+诱导的大鼠PC12细胞凋亡作用,其作用与保护细胞线粒体结构的完整性有关。     

Protective effect of verbascoside on 1-methyl-4-phenylpyridinium ion-induced neurotoxicity in PC12 cells

The neuroprotective effects of verbascoside, one of phenylpropanoid glucoside isolated from the Chinese herbal medicine Buddleja officinalis Maxim, on 1-methyl-4-phenylpyridinium ion (MPP(+)) induced apoptosis and oxidative stress in PC12 neuronal cells were investigated. Treatment of PC12 cells with MPP(+) for 48 h induced apoptotic death as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and flow cytometry, the activation of caspase-3 measured by the caspase-3 activity assay kit, the reduction in mitochondrial membrane potential with laser scanning confocal microscopy and the increase in the extracellular hydrogen peroxide level. Simultaneous treatment with verbascoside markedly attenuated MPP(+)-induced apoptotic death, increased extracellular hydrogen peroxide level, the activation of caspase-3 and the collapse of mitochondrial membrane potential. These results strongly indicate that verbascoside may provide a useful therapeutic strategy for the treatment of oxidative stress-induced neurodegenerative disease such as Parkinson's disease.     

Possible role of interleukin-6 in PC12 cell death induced by MPP+ and tetrahydroisoquinoline

Interleukin (IL)-6 has been shown to protect neuronal cells from cell death induced by various stimulants. Although neuronal cells including PC12 cells were shown to produce IL-6, little is known about the effects of dopaminergic neurotoxins, 1,2,3,4-tetrahydroisoquinoline (TIQ) and 1-methyl-4-phenylpyridinium ion (MPP(+)), on IL-6 expression in PC12 cells. In the present study, we investigated the role of IL-6 in the TIQ- and MPP(+)-induced cell death in PC12 cells. Treatment with 3.2 mM TIQ for 24 h caused a delayed cell death (lactate dehydrogenase (LDH) leakage and nuclear DNA fragmentation) markedly 72 h after the addition. Addition of 0.4 mM MPP(+) caused LDH leakage and nuclear DNA fragmentation 24 h after the addition. The cell death induced by MPP(+) was inhibited by an inhibitor of caspases, z-Val-Ala-Asp(OMe)-fluoromethylketone. The cell death induced by TIQ or MPP(+) was inhibited by nerve growth factor and 10% serum and significantly enhanced by the treatment with anti-IL-6 antibody. Both neurotoxins decreased the IL-6 mRNA level in PC12 cells without changing the other tested mRNA levels (IL-1 alpha, beta-actin, etc.). These findings suggest that dopaminergic neurotoxins cause cell death in PC12 cells at least partially by changing IL-6 expression.     

Cell damage following carbon monoxide releasing molecule exposure: implications for therapeutic applications

The cytoprotective properties of carbon monoxide (CO) gas and CO-releasing molecules (CORMs) are well established. Despite promising pre-clinical results, little attention has been paid to the toxicological profile of CORMs. The effects of CORM-2 and its CO-depleted molecule (iCORM-2) (20-400 μM) were compared in primary rat cardiomyocytes and two cell lines [human embryonic kidney (HeK) and Madine-Darby canine kidney Cells (MDCK)]. Cells were assessed for cell viability, apoptosis, necrosis, cytology, mitochondrial energetics, oxidative stress and cell cycle arrest markers. In separate experiments, the anti-apoptotic effects of CORM-2 and i-CORM-2 treatment were compared against CO gas treatment in HeK and MDCK lines. H(2)O(2) -induced cellular damage, measured by lactate dehydrogenase (LDH) release from primary cardiomyocytes, was reduced by 20 μM CORM-2; LDH activity, however, was directly inhibited by 400 μM CORM-2. Both CORM-2/iCORM-2 and CO gas decreased cisplatin-induced caspase-3 activity in MDCK and HeK cells suggesting an anti-apoptotic effect. Conversely, both CORM-2 and iCORM-2 induced significant cellular toxicity in the form of decreased cell viability, abnormal cell cytology, increased apoptosis and necrosis, cell cycle arrest and reduced mitochondrial enzyme activity. Comparison of these markers after CO gas administration to MDCK cells found significantly less cellular toxicity than in 100 μM CORM-2/iCORM-2-treated cells. CO gas did not have an adverse effect on mitochondrial energetics and integrity. Release of CO by low concentrations of intact CORM-2 molecules provides cytoprotective effects. These results show, however, that the ruthenium-based CORM by-product, iCORM-2, is cytotoxic and suggest that the accumulation of iCORM-2 would seriously limit any clinical application of the ruthenium-based CORMs.     

Protocatechuic acid suppresses MPP+ -induced mitochondrial dysfunction and apoptotic cell death in PC12 cells.

Protocatechuic acid (PCA), a phenolic compound isolated from the kernels of Alpinia (A.) oxyphylla, showed antioxidant neuroprotective effect in our previous study. Here, we investigated the effect of PCA on the MPP(+)-induced mitochondrial dysfunction and apoptotic cell death in PC12 cells. The apoptosis in MPP(+)-induced PC12 cells was associated with loss of mitochondrial membrane potential, the formation of reactive oxygen species (ROS), GSH depletion, activation of caspase-3 and down-regulation of Bcl-2. In contrast, treatment of PC12 cells with PCA significantly prevented the above-mentioned mitochondrial dysfunction. Our data pointed to the potential clinical application/use of PCA to overcome neurodegenerative diseases such as Parkinson's disease.     

Acteoside from Cistanche salsa inhibits apoptosis by 1-methyl-4-phenylpyridinium ion in cerebellar granule neurons

In this study we assessed the effect of acteoside that significantly improved cell viability and inhibited lactate dehydrogenase (LDH) release. Furthermore acteoside prevented a neurotoxin 1-methyl-4-phenylpyridinium ion (MPP+)-induced apoptosis in CGNs. Accordingly, our flow cytometric analysis of CGNs after acteoside treatment revealed a decrease in the number of the MPP+-induced apoptotic cells (P < 0.001). Western blot analysis demonstrated that acteoside inhibits the active caspase-3 fragment (17 kDa) (P < 0.001) and the proteolytic poly (ADP-ribose) polymerase (PARP) fragment (85 kDa) expression (P < 0.001) following MPP + treatment in CGNs. We conclude that acteoside prevents the MPP+-induced apoptosis and inhibits the apoptosis-related pathway.     

色霉素对多巴胺能神经元的保护作用

目的研究色霉素对MPP+诱导凋亡的多巴胺能神经元的保护作用。方法体外培养的胎鼠腹侧中脑神经元,以MPP+引起多巴胺能神经元凋亡作为帕金森病的细胞模型,Hoechst 33258荧光核染色法检测神经元的凋亡情况,免疫细胞化学方法检测多巴胺能神经元内磷酸化Tau水平。结果10μmol·L-1 MPP+可以升高Tau磷酸化水平,诱导神经元发生典型的凋亡。而色霉素通过抑制Tau磷酸化,减少神经元凋亡,使TH-阳性细胞数目增加。结论色霉素可以通过抑制Tau磷酸化而对MPP+诱导凋亡的多巴胺能神经元发挥保护作用,色霉素可能用于临床防治帕金森病等神经退行性疾病。     

δ阿片受体激活对过氧化氢损伤的心肌细胞的保护作用

目的研究δ阿片受体激活剂D-丙(2)-D-亮-(5)-脑啡肽(DADLE)对过氧化氢(H2O2)损伤的心肌细胞的保护作用及其机制。方法分离乳大鼠心肌细胞,培养48h后分为正常对照、H2O2(200μmol.L-1)、H2O2+DADLE(1μmol.L-1)、H2O2+DADLE+纳曲吲哚(10μmol.L-1)和H2O2+DADLE+U0126(10nmol.L-1)组,继续培养48h。用[3H]TdR掺入法检测心肌细胞增殖反应,流式细胞仪检测心肌细胞凋亡百分率,乳酸脱氢酶(LDH)活性测定试剂盒测定培养上清LDH活性,硫代巴比妥酸显色法测定细胞内丙二醛(MDA)含量,黄嘌呤氧化酶法测定细胞内超氧化物歧化酶(SOD)活性,Western蛋白印迹法检测细胞外信号调节激酶磷酸化(p-ERK)水平。结果①与正常对照组比较,H2O2组心肌细胞[3H]TdR掺入值明显降低,细胞凋亡率升高;培养上清LDH活性和MDA含量明显增加,SOD活性和Ap-ERK/AERK的比值降低。②与H2O2组比较,DADLE可使心肌细胞[3H]TdR掺入值升高,细胞凋亡率下降;培养上清LDH活性和MDA含量降低,SOD活性和Ap-ERK/AERK的比值升高。③分别加入δ阿片受体拮抗剂纳曲吲哚和ERK拮抗剂U0126,DADLE对上述指标的逆转作用被抑制。结论δ阿片受体激活对H2O2损伤的心肌细胞具有保护作用,其机制可能与其增强心肌细胞的抗氧化功能及促进ERK磷酸化有关。     

Protocatechuic acid from Alpinia oxyphylla against MPP+-induced neurotoxicity in PC12 cells.

An ethyl acetate extract of Alpinia oxyphylla was found to possess neuroprotective activity against 1-methyl-4-phenylpyridinium ion (MPP(+)) induced apotosis and oxidative stress in cultured PC12 cells. From the extract, a phenolic compound was isolated through bioassay-guided fractionation and identified as protocatechuic acid (PCA) by IR, MS, and (1)H and (13)C NMR spectroscopy. It was the first time which was isolated from the kernels of A. oxyphylla. Exposure of PC12 cells to 1mM MPP(+) may cause significant viability loss and apoptotic cell death. PCA stimulated PC12 cellular proliferation and markedly attenuated MPP(+)-induced apoptotic cell death in a dose-dependent manner. By observing the nuclear morphological changes and flow cytometric analysis, PCA showed its significant effect on protecting PC12 cells against MPP(+)-induced apoptosis. Meanwhile, PCA enhanced the activities of superoxide dismutase (SOD) and catalase (CAT) in PC12 cells. In addition, PCA also dose-dependently reduced the hydrogen peroxide (H(2)O(2))- or sodium nitroprusside (SNP)-induced cell death in PC12 cells. The results suggest that PCA may be one of the primary active components in the kernels of A. oxyphylla and provide a useful therapeutic strategy for the treatment of oxidative stress-induced neurodegenerative disease such as Parkinson's disease.     

姜黄素对MPP~+诱导PC12细胞凋亡的影响

目的观察姜黄素对MPP+诱导的PC12细胞凋亡的影响。方法采用透射电镜,hoechst染色和流式细胞仪(FCM)观察PC12细胞凋亡,间接免疫荧光流式细胞术检测PC12细胞bcl2的表达。结果PC12细胞自然凋亡率为(15±01)%,05mmol·L-1、1mmol·L-1和2mmol·L-1MPP+作用24h后,PC12细胞凋亡率分别为(441±38)%、(549±21)%和(822±26)%;20μmol·L-1和40μmol·L-1姜黄素对PC12细胞作用24h后无明显凋亡诱导作用,但可分别使05mmol·L-1MPP+处理组细胞凋亡率由(441±38)%下降到(341±38)%和(179±15)%(P<001);PC12细胞经20μmol·L-1姜黄素处理24h后,其bcl2表达率由正常对照的(3643±790)%增加到(7673±860)%(P<001)。结论姜黄素可以抑制MPP+诱导的PC12细胞凋亡,其作用机制之一可能与促进bcl2的表达有关。     

Melatonin attenuates 1-methyl-4-phenylpyridinium-induced PC12 cell death

Aim: To explore the effect of melatonin on PC 12 cell death induced by 1-methyl-4-phenylpyridinium (MPP^ ). Methods: MTT assay, lactate dehydrogenase (LDH) efflux assay, and immunohistochemistry methods were used to measure neurotoxicity of PC 12 cells treated acutely with MPP^ in low glucose and high glucose conditions, and to assess the neuroprotective effect of melatonin on PC 12 cell death induced by MPP^ .Results: In a low glucose condition, MPP^ significantly induced PC 12 cell death, which showed time and concentration dependence. In a serum-free low glucose condition, the percentages of viability of cells treated with MPP^ for 12, 24, 48, 72, and 96 h were 85.1%, 75.4%, 64.9%, 28.15%, and 9%, respectively. The level of LDH in the culture medium increased and tyrosine hydroxylase positive (TH^ ) cell count decreased. However, in a serum-free high glucose condition, MPP^ did not significantly induce PC12 cell death compared with control at various concentrations and time regimens. When the cells were preincubated with melatonin 250 μmol/L for 48, 72, and 96 h in a serum-free low glucose condition, cell survival rate significantly increased to 78.1%, 58.8%, and 31.6%, respectively. Melatonin abolished the LDH leakage of cells treated with MPP^ and increased TH^ cells count. Conclusion: MPP^ caused concentration-dependent PC12 cell death. The level of glucose was an important factor to MPP^ induced dopaminergic PC 12 cell death. Low glucose level could potentiate MPP^ toxicity, while high glucose level could reduce the toxicity. In addition, melatonin attenuated PC12 cell death induced by MPP^ .     

七叶皂苷肾细胞毒性及抗氧化剂的拮抗作用

目的研究七叶皂苷不同组分和七叶皂苷钠注射剂提取物(ESA)的肾毒性及可能的作用机制。方法以成人肾近曲小管上皮细胞(HK-2)为模型,氯化汞(HgCl2)为阳性对照,研究含七叶皂苷Ia(A)与Ib(B)的提取物(A+B)、含异七叶皂苷Ia(C)与Ib(D)提取物(C+D)、含A+B+C+D及ESA的细胞毒性以及抗氧化剂维生素C(Vit C)和还原型谷胱甘肽(GSH)对ESA毒性的拮抗作用。用MTT法检测细胞存活率;乳酸脱氢酶(LDH)释放实验检测细胞膜损伤;倒置显微镜观察细胞形态学改变;流式细胞仪检测细胞周期和细胞凋亡。结果A+B,C+D、A+B+C+D和ESA分别与HK-2细胞作用24h,均显著抑制细胞的存活,并且具有良好的浓度-效应关系,半数抑制浓度(IC50)值分别为:(26.5±1.7),(35.4±2.2),(28.4±1.8)和(23.3±1.7)μmol·L-1。A+B,A+B+C+D(60~100μmol.L-1),C+D和ESA(40~100μmol·L-1)分别与HK-2细胞作用24h,LDH释放率明显升高。80μmo.lL-1A+B,C+D,A+B+C+D和ESA分别与HK-2细胞作用24h,相差显微镜下可观察到细胞收缩、变圆。ESA和A+B+C+D均能导致HK-2细胞出现明显的G2/M期阻滞,同时伴有S期细胞减少,并且随着浓度的增加,G2/M期细胞所占的比例逐渐增加。抗氧化剂Vit C不能拮抗HgCl2和ESA引起的细胞损伤作用;GSH对HgCl2 30 μmol·L-1引起的细胞损伤有一定的拮抗作用,GSH对ESA30及60μmol·L-1引起的细胞损伤均有一定的拮抗作用,但对ESA 100 μmol·L-1引起的细胞损伤无拮抗作用。结论七叶皂苷对肾小管上皮细胞具有明显的毒性,七叶皂苷不同组分之间的毒性存在显著差异。氧化损伤可能是其毒性机制之一。     

Iptakalim protects against MPP+-induced degeneration of dopaminergic neurons in association with astrocyte activation

Astrocyte activation observed in the MPTP mouse model and Parkinson's disease patients participates in the cascade of deleterious events that ultimately leads to death of dopaminergic neurons in the substantia nigra pars compacta (SNpc). The present study aimed to elucidate whether inhibiting astrocyte activation was involved in the protective effects of iptakalim (Ipt), a novel ATP-sensitive potassium channel opener, on MPP+-induced degeneration of dopaminergic neurons. The results showed that Ipt could decrease MPP+-induced TNF-alpha release and p38 MAPK activation in reactive astrocytes. The effects of Ipt were reversed by the mitochondrial KATP blocker, 5-hydroxydecanoate, indicating that mitochondrial KATP channels participate in the regulation of astrocyte activation. Moreover, systematic administration of Ipt could significantly alleviate MPP+-induced behavioural symptoms in motor coordination, the loss of dopaminergic neurons, and the activation of astrocyte and microglia in the SNpc. Together, these findings suggest that Ipt may protect against MPP+-induced degeneration of dopaminergic neurons by inhibiting astrocyte activation and subsequent release of pro-inflammatory factors.     

Dietary phenolic antioxidants, caffeic acid and Trolox, protect rainbow trout gill cells from nitric oxide-induced apoptosis

Caffeic acid (CA) and Trolox are phenolic acids that have beneficial antioxidant effect, but the underlying mechanisms involved are not fully understood. The extent to which CA and Trolox protect against sodium nitroprusside (SNP)-induced oxidative cell injury was investigated in cultured rainbow trout gill cells. The cells exposed to SNP for 24 h displayed a dose-dependent leakage of lactate dehydrogenase (LDH) and decreased cell viability as indicated by the MTT assay (mitochondrial dehydrogenase activity). Both effects were prevented by treatment with 50 μM CA or Trolox. CA or Trolox, protected against SNP-induced caspase-3 activation and DNA fragmentation, indicating a reduction of apoptosis. Thus, the results indicate that SNP induced cell death is caspase-3 related apoptosis and the treatment with CA inhibited the apoptotic pathway. In addition, we studied the effect of CA and Trolox on expression of zinc-responsive antioxidant genes such as metallothioneins (MT), glutathione-S-transferase (GST Class pi) and glucose-6-phosphate dehydrogenase (G6PD) in cultured gill cells. CA, 100 μM, increased accumulation of mRNA for MTA, MTB, GST and G6PD in cells. Thus, in addition to its ability to sequester free radicals, CA may protect against oxidative stress through expression of zinc-induced antioxidant proteins. Because of these properties we suggest that CA could be a beneficial additive to fish feeds in aquaculture.