Resveratrol and pinostilbene confer neuroprotection against aging-related deficits through an ERK1/2-dependent mechanism

Age-related declines in motor function may be due, in part, to an increase in oxidative stress in the aging brain leading to dopamine (DA) neuronal cell death. In this study, we examined the neuroprotective effects of natural antioxidants resveratrol and pinostilbene against age-related DAergic cell death and motor dysfunction using SH-SY5Y neuroblastoma cells and young, middle-aged, and old male C57BL/6 mice. Resveratrol and pinostilbene protected SH-SY5Y cells from a DA-induced decrease in cell viability. Dietary supplementation with resveratrol and pinostilbene inhibited the decline of motor function observed with age. While DA and its metabolites (DOPAC and HVA), dopamine transporter, and tyrosine hydroxylase levels remain unchanged during aging or treatment, resveratrol and pinostilbene increased ERK1/2 activation in vitro and in vivo in an age-dependent manner. Inhibition of ERK1/2 in SH-SY5Y cells decreased the protective effects of both compounds. These data suggest that resveratrol and pinostilbene alleviate age-related motor decline via the promotion of DA neuronal survival and activation of the ERK1/2 pathways.     

Dietary oxyresveratrol prevents parkinsonian mimetic 6-hydroxydopamine neurotoxicity

Oxyresveratrol (OXY) is a polyhydroxylated stilbene existing in mulberry. Increasing lines of evidence have shown its neuroprotective effects against Alzheimer disease and stroke. However, little is known about its neuroprotective effect in Parkinson disease (PD). Owing to its antioxidant activity, blood-brain barrier permeativity, and water solubility, we hypothesized that OXY may exert neuroprotective effects against parkinsonian mimetic 6-hydroxydopamine (6-OHDA) neurotoxicity. Neuroblastoma SH-SY5Y cells have long been used as dopaminergic neurons in PD research. We found that both pretreatment and posttreatment with OXY on SH-SY5Y cells significantly reduced the release of lactate dehydrogenase, the activity of caspase-3, and the generation of intracellular reactive oxygen species triggered by 6-OHDA. Compared to resveratrol, OXY exhibited a wider effective dosage range. We proved that OXY could penetrate the cell membrane by HPLC analysis of cell extracts. These results suggest that OXY may act as an intracellular antioxidant to reduce oxidative stress induced by 6-OHDA. Western blot analysis demonstrated that OXY markedly attenuated 6-OHDA-induced phosphorylation of JNK and c-Jun. Furthermore, we proved that OXY increased the basal levels of SIRT1, which may disclose new pathways accounting for the neuroprotective effects of OXY. Taken together, our results suggest OXY, a dietary phenolic compound, as a potential nutritional candidate for protection against neurodegeneration in PD.     

Gallic Acid Protects 6-OHDA Induced Neurotoxicity by Attenuating Oxidative Stress in Human Dopaminergic Cell Line

Gallic acid is one of the most important polyphenolic compounds, which is considered an excellent free radical scavenger. 6-Hydroxydopamine (6-OHDA) is a neurotoxin, which has been implicated in mainly Parkinson’s disease (PD). In this study, we investigated the molecular mechanism of the neuroprotective effects of gallic acid on 6-OHDA induced apoptosis in human dopaminergic cells, SH-SY5Y. Our results showed that 6-OHDA induced cytotoxicity in SH-SY5Y cells was suppressed by pre-treatment with gallic acid. The percentage of live cells (90%) was high in the pre-treatment of gallic acid when compared with 6-OHDA alone treated cell line. Moreover, gallic acid was very effective in attenuating the disruption of mitochondrial membrane potential, elevated levels of intracellular ROS and apoptotic cell death induced by 6-OHDA. Gallic acid also lowered the ratio of the pro-apoptotic Bax protein and the anti-apoptotic Bcl-2 protein in SH-SY5Y cells. 6-OHDA exposure was up-regulated caspase-3 and Keap-1 and, down-regulated Nrf2, BDNF and p-CREB, which were sufficiently reverted by gallic acid pre-treatment. These findings indicate that gallic acid is able to protect the neuronal cells against 6-OHDA induced injury and proved that gallic acid might potentially serve as an agent for prevention of several human neurodegenerative diseases caused by oxidative stress and apoptosis.     

Sulforaphane as an inducer of glutathione prevents oxidative stress-induced cell death in a dopaminergic-like neuroblastoma cell line

The total GSH depletion observed in the substantia nigra (SN) appears to be responsible for subsequent oxidative stress (OS), mitochondrial dysfunction, and dopaminergic cell loss in patients with Parkinson's disease. A strategy to prevent the OS of dopaminergic cells in the SN may be the use of chemopreventive agents as inducers of endogenous GSH, antioxidant and phase 2 enzymes. In this study, we demonstrated that treatment of the dopaminergic-like neuroblastoma SH-SY5Y cell line with sulforaphane (SF), a cruciferous vegetables inducer, resulted in significant increases of total GSH level, NAD(P)H : quinone oxidoreductase-1, GSH-transferase and -reductase, but not GSH-peroxidase, catalase and superoxide dismutase activities. Further, the elevation of GSH levels, GSH-transferase and NAD(P)H:quinone oxidoreductase-1 activities was correlated to an increase of the resistance of SH-SY5Y cells to toxicity induced by H₂O₂ or 6-hydroxydopamine (6-OHDA). The pre-treatment of SH-SY5Y cells with SF was also shown to prevent various apoptotic events (mitochondrial depolarization, caspase 9 and 3 activation and DNA fragmentation) and necrosis elicited by 6-OHDA. Further, the impairment of antioxidant capacity and reactive oxygen species formation at intracellular level after exposure to 6-OHDA was effectively counteracted by pre-treatment with SF. Last, both the cytoprotective and antioxidant effects of SF were abolished by the addition of buthionine sulfoximine supporting the main role of GSH in the neuroprotective effects displayed by SF. These findings show that SF may play a role in preventing Parkinson's disease.     

Neuroblastoma cells expressing the noradrenaline transporter are destroyed more selectively by 6-fluorodopamine than by 6-hydroxydopamine

6-Hydroxydopamine (6-OHDA) has been used for lesioning catecholaminergic neurons and attempted purging of neuroblastoma cells from hematopoietic stem cells in autologous bone marrow transplantation (ABMT). Neurotoxicity is mediated primarily by reactive oxygen species. In ABMT, 6-OHDA, as a purging agent, has been unsuccessful. At physiological pH it autooxidizes before targeted uptake, resulting in nonspecific cytotoxicity of nontarget cells. A catecholamine analogue, similar to 6-OHDA but with a lower rate of autooxidation enabling uptake by target cells, is thus required. Electron paramagnetic resonance spectra in this study show that 6-fluorodopamine (6-FDA) hydrolyzes slowly to 6-OHDA at physiological pH. Oxygen consumption, H(2)O(2), and quinone production are found to be intermediate between those of 6-OHDA and dopamine (DA). Relative neurotoxicity of these compounds was assessed by cell viability and DNA damage in the human neuroblastoma lines SH-SY5Y and SK-N-LO, which express and lack the noradrenaline transporter, respectively. Specific uptake of DA and 6-FDA by SH-SY5Y cells was demonstrated by competitive m-[(131)I]iodobenzylguanidine uptake inhibition. The competition by 6-OHDA was low owing to rapid autooxidation during incubation with equal toxicity toward both cell types. 6-FDA toxicity was preferential for SH-SY5Y cells and reduced in the presence of desipramine, a catecholamine uptake inhibitor. We demonstrate that 6-FDA cytotoxicity is more specific for cells expressing catecholamine reuptake systems than is 6-OHDA cytotoxicity.     

6-Hydroxydopamine (6-OHDA) induces Drp1-dependent mitochondrial fragmentation in SH-SY5Y cells

Mitochondrial alterations have been associated with the cytotoxic effect of 6-hydroxydopamine (6-OHDA), a widely used neurotoxin to study Parkinson's disease. Herein we studied the potential effects of 6-OHDA on mitochondrial morphology in SH-SY5Y neuroblastoma cells. By immunofluorescence and time-lapse fluorescence microscopy we demonstrated that 6-OHDA induced profound mitochondrial fragmentation in SH-SY5Y cells, an event that was similar to mitochondrial fission induced by overexpression of Fis1p, a membrane adaptor for the dynamin-related protein 1 (DLP1/Drp1). 6-OHDA failed to induce any changes in peroxisome morphology. Biochemical experiments revealed that 6-OHDA-induced mitochondrial fragmentation is an early event preceding the collapse of the mitochondrial membrane potential and cytochrome c release in SH-SY5Y cells. Silencing of DLP1/Drp1, which is involved in mitochondrial and peroxisomal fission, prevented 6-OHDA-induced fragmentation of mitochondria. Furthermore, in cells silenced for Drp1, 6-OHDA-induced cell death was reduced, indicating that a block in mitochondrial fission protects SH-SY5Y cells against 6-OHDA toxicity. Experiments in mouse embryonic fibroblasts deficient in Bax or p53 revealed that both proteins are not essential for 6-OHDA-induced mitochondrial fragmentation. Our data demonstrate for the first time an involvement of mitochondrial fragmentation and Drp1 function in 6-OHDA-induced apoptosis.     

Protective effect of (+/-) isoborneol against 6-OHDA-induced apoptosis in SH-SY5Y cells.

Oxidative stress caused by dopamine (DA) may play an important role in the pathogenesis of Parkinson's disease (PD). (+/-) Isoborneol is a monoterpenoid alcohol present in the essential oils of numerous medicinal plants and is a known antioxidant. In this study, we investigated the neuroprotective effect of isoborneol against 6-hydroxydopamine (6-OHDA)-induced cell death in human neuroblastoma SH-SY5Y cells. Pretreatment of SH-SY5Y cells with isoborneol significantly reduced 6-OHDA-induced generation of reactive oxygen species (ROS) and 6-OHDA-induced increases in intracellular calcium. Furthermore, apoptosis induced by 6-OHDA was reversed by isoborneol treatment. Isoborneol protected against 6-OHDA-induced increases in caspase-3 activity and cytochrome C translocation into the cytosol from mitochondria. Isoborneol prevented 6-OHDA from decreasing the Bax/Bcl-2 ratio. We also observed that isoborneol decreased the activation of c-Jun N-terminal kinase and induced activation of protein kinase C (PKC) which had been suppressed by 6-OHDA. Our results indicate that the protective function of isoborneol is dependent upon its antioxidant potential and strongly suggest that isoborneol may be an effective treatment for neurodegenerative diseases associated with oxidative stress.     

Neuroprotective effects of kobophenol A against the withdrawal of tropic support, nitrosative stress, and mitochondrial damage in SH-SY5Y neuroblastoma cells

This study examined the neuroprotective effects of kobophenol A (kob A), oligomeric stillbene, and a resveratrol tetramer. Neuronal death induced by the withdrawal of tropic support was ameliorated by kob A. The protective effect of kob A against nitrosative/oxidative or mitochondrial damages resulted in the inhibition of the ROS, intracellular calcium ion level, and mitochondrial transmembrane potential changes on SH-SY5Y cells.     

Structure-activity relationship analysis of antioxidant ability and neuroprotective effect of gallic acid derivatives

Gallic acid and its derivatives are a group of naturally occurring polyphenol antioxidants which have recently been shown to have potential healthy effects. In order to understand the relationship between the structures of gallic acid derivatives, their antioxidant activities, and neuroprotective effects, we examined their free radical scavenging effects in liposome and anti-apoptotic activities in human SH-SY5Y cell induced by 6-hydrodopamine autooxidation. It was found that these polyphenol antioxidants exhibited different hydrophobicity and could cross through the liposome membrane to react with 1,1-diphenyl-2-picryl-hydrazyl (DPPH) free radical in a time and dose-dependent manner. At the same time, the structure-antioxidant activity relationship of gallic acid derivatives on scavenging DPPH free radical in the liposome was also analyzed based on theoretical investigations. Analysis of cell apoptosis, intracellular GSH levels, production of ROS and the influx of Ca(2+) indicated that the protective effects of gallic acid derivatives in cell systems under oxidative stress depend on both their antioxidant capacities and hydrophobicity. However, the neuroprotective effects of gallic acid derivatives seem to depend more on their molecular polarities rather than antioxidant activities in the human SH-SY5Y cell line. In conclusion, these results reveal that compounds with high antioxidant activity and appropriate hydrophobicity are generally more effective in preventing the injury of oxidative stress in neurodegenerative diseases.     

Autophagy-dependent and -independent involvement of AMP-activated protein kinase in 6-hydroxydopamine toxicity to SH-SY5Y neuroblastoma cells

The role of the main intracellular energy sensor adenosine monophosphate (AMP)-activated protein kinase (AMPK) in the induction of autophagic response and cell death was investigated in SH-SY5Y human neuroblastoma cells exposed to the dopaminergic neurotoxin 6-hydroxydopamine (6-OHDA). The induction of autophagy in SH-SY5Y cells was demonstrated by acridine orange staining of intracellular acidic vesicles, the presence of autophagosome- and autophagolysosome-like vesicles confirmed by transmission electron microscopy, as well as by microtubule-associated protein 1 light-chain 3 (LC3) conversion and p62 degradation detected by immunoblotting. 6-OHDA induced phosphorylation of AMPK and its target Raptor, followed by the dephosphorylation of the major autophagy inhibitor mammalian target of rapamycin (mTOR) and its substrate p70S6 kinase (S6K). 6-OHDA treatment failed to suppress mTOR/S6K phosphorylation and to increase LC3 conversion, p62 degradation and cytoplasmatic acidification in neuroblastoma cells in which AMPK expression was downregulated by RNA interference. Transfection of SH-SY5Y cells with AMPK or LC3β shRNA, as well as treatment with pharmacological autophagy inhibitors suppressed, while mTOR inhibitor rapamycin potentiated 6-OHDA-induced oxidative stress and apoptotic cell death. 6-OHDA induced phosphorylation of p38 mitogen-activated protein (MAP) kinase in an AMPK-dependent manner, and pharmacological inhibition of p38 MAP kinase reduced neurotoxicity, but not AMPK activation and autophagy triggered by 6-OHDA. Finally, the antioxidant N-acetyl cysteine antagonized 6-OHDA-induced activation of AMPK, p38 and autophagy. These data suggest that oxidative stress-mediated AMPK/mTOR-dependent autophagy and AMPK/p38-dependent apoptosis could be valid therapeutic targets for neuroprotection.     

Identification,characterization and in vitro neuroprotection of N6-(4-hydroxybenzyl) adenine riboside and its metabolites

N⁶-(4-hydroxybenzyl) adenine riboside (NHBA), isolated from Gastrodia elata Blume, has been demonstrated to show great pharmacological effects. The present study aimed to synthesize and identify the metabolites of NHBA, and to determine their neuroprotective potentials in vitro. After incubation with rat liver microsomes in the presence of NADPH, two metabolites were detected, which were further semisynthesized and identified as N⁶-(4-hydroxylbenzyl) purine (NHBP) and N⁶-(3,4-dihydroxylbenzyl) adenine riboside (ONHBA) by UPLC-QTOF-MS, ¹H NMR and ¹³C NMR. Furthermore, the neuroprotective activities of NHBA and two metabolites were evaluated in SH-SY5Y cells. Our results demonstrated that NHBA substantially protected against H₂O₂-induced neuronal death in SH-SY5Y cells. Moreover, both ONHBA and NHBP could significantly prevent Aβ oligomers- and H₂O₂-induced neuronal death in SH-SY5Y cells. These results suggested that NHBA and its metabolites, ONHBA and NHBP, might be suitable for the development of new drugs in the treatment of neurodegenerative diseases, including Alzheimer’s disease in particular.     

Involvement of protein kinase C activation and cell survival/ cell cycle genes in green tea polyphenol (-)-epigallocatechin 3-gallate neuroprotective action

Studies from our laboratory have demonstrated that the major green tea polyphenol, (-)-epigallocatechin 3-gallate (EGCG), exerts potent neuroprotective actions in the mice model of Parkinson's disease. These studies were extended to neuronal cell culture employing the parkinsonism-inducing neurotoxin, 6-hydroxydopamine (6-OHDA). Pretreatment with EGCG (0.1-10 microm) attenuated human neuroblastoma (NB) SH-SY5Y cell death, induced by a 24-h exposure to 6-OHDA (50 microm). Potential cell signaling candidates involved in this neuroprotective effect were further examined. EGCG restored the reduced protein kinase C (PKC) and extracellular signal-regulated kinases (ERK1/2) activities caused by 6-OHDA toxicity. However, the neuroprotective effect of EGCG on cell survival was abolished by pretreatment with PKC inhibitor GF 109203X (1 microm). Because EGCG increased phosphorylated PKC, we suggest that PKC isoenzymes are involved in the neuroprotective action of EGCG against 6-OHDA. In addition, gene expression analysis revealed that EGCG prevented both the 6-OHDA-induced expression of several mRNAs, such as Bax, Bad, and Mdm2, and the decrease in Bcl-2, Bcl-w, and Bcl-x(L). These results suggest that the neuroprotective mechanism of EGCG against oxidative stress-induced cell death includes stimulation of PKC and modulation of cell survival/cell cycle genes.     

Iron chelator Deferoxamine protects human neuroblastoma cell line SH-SY5Y from 6-Hydroxydopamine-induced apoptosis and autophagy dysfunction

BackgroundIntracellular iron involves in Fenton’s reaction-mediated Hydroxyl radical (OH·) generation by reacting with the neurotoxic agent 6-Hydroxydopamine (6-OHDA) autoxidation derivative Hydrogen Peroxide (H₂O₂). Several studies have been conducted so far on the neuroprotective activities of the iron chelator Deferoxamine (DFO) but little or no clear evidence about the underlying cellular mechanism is available.MethodsThe present study was conducted on Human neuroblastoma cell line SH-SY5Y in the absence or presence of 6-OHDA or H₂O₂ and / or DFO. Following incubation, cell viability assay, intracellular reactive oxygen species (ROS) determination, flow cytometric quantification of apoptotic cells followed by nuclear staining, intracellular tracking of transfected fusion construct of microtubule-associated protein 1B-light chain with Green fluorescent protein - Red fluorescent protein (LC3B-GFP-RFP reporters) and immunocytochemistry of intracellular Cathepsin protein by confocal microscopy, were conducted. In addition, western blotting was carried out to detect expressions of apoptotic and autophagy related proteins.ResultsThis study confirmed the neuroprotective potential of DFO by inhibiting 6-OHDA-mediated cell death and ROS generation. Reduced percentage of apoptotic cells and appearance of altered nuclei architecture followed by a reduced expression of cleaved PARP (Poly-ADP-ribose Polymerase) and cleaved Caspase-3 were observed upon DFO treatment against 6-OHDA, and as well as against H₂O₂ in SH-SY5Y cell lines. Besides, DFO induced the intracellular autophagolysosome formation (red puncta) rather than autophagosome (yellow puncta) only. Thereafter it was observed that DFO restored the expression of intracellular lysosomal protease Cathepsin and reduced the expression of the LC3-II.ConclusionTaken together, this study clearly demonstrated that the anti-Fenton activity of DFO inhibited apoptosis and caused blockade in ALP or autophagy dysfunction in SH-SY5Y cell lines. These outcomes further suggest that DFO provides neuroprotection by inhibiting apoptosis and inducing the progression of Autophagy- lysosomal pathway (ALP).     

Glycyrrhizic Acid Alleviates 6-Hydroxydopamine and Corticosterone-Induced Neurotoxicity in SH-SY5Y Cells Through Modulating Autophagy

Recent researches have shown that autophagy is associated with the pathogenesis of neurodegenerative disorders, but there is no paper to investigate the effects of autophagy modulation on Parkinson’s disease depression (PDD). In addition, glycyrrhizic acid (GA), the major bioactive ingredient of Radix glycyrrhizae, can induce autophagy and ease rotenone-induced Parkinson’s disease (PD). However, there is also no paper to study the action and molecular mechanisms of GA on PDD. In this research, we built the injury model of SH-SY5Y cells through 6-hydroxydopamine (6-OHDA) and corticosterone (CORT). Then, our results showed that GA markedly increased the viability and decreased the apoptosis in SH-SY5Y cells after pre-treating with 6-OHDA and CORT. Moreover, GA notably decreased the expressions of α-Syn and p-S1292-LRRK2 proteins, and significantly increased the levels of CREB and BDNF proteins. Previous papers have suggested that CORT contributed to dopaminergic neurodegeneration via the glucocorticoid (GC)/glucocorticoid receptor (GR) interaction, and our results showed that GA reduced GC level and hypothalamic–pituitary–adrenal (HPA) activity in SH-SY5Y cells by regulating GR signaling pathway. Furthermore, mechanism investigations also showed that GA had the ability to up-regulate the conversion of LC3B II/I and the expression of Beclin-1, and induce autophagy in SH-SY5Y cells, which were reversed by the autophagy inhibitor 3-methyladenine (3-MA). Collectively, these findings proved that GA exerted efficient activity against neurotoxicity in SH-SY5Y cells induced by 6-OHDA and CORT via activation of autophagy, which should be developed as an efficient candidate for treating PDD in the future.     

Neuroprotective effects of (E)-3,4-diacetoxystyryl sulfone and sulfoxide derivatives in vitro models of Parkinson's disease

(E)-3,4-dihydroxystyryl aralkyl sulfones and sulfoxides have been reported as novel multifunctional neuroprotective agents in previous studies, which as phenolic compounds display antioxidative and antineuroinflammatory properties. To further enhance the neuroprotective effects and study structure-activity relationship of the derivatives, we synthesized their acetylated derivatives, (E)-3,4-diacetoxystyryl sulfones and sulfoxides, and examined their neuroprotective effects in vitro models of Parkinson’s disease. The results indicate that (E)-3,4-diacetoxystyryl sulfones and sulfoxides can significantly inhibit kinds of neuron cell injury induced by toxicities, including 6-OHDA, NO, and H₂O₂. More important, they show higher antineuroinflammatory properties and similar antioxidative properties to corresponding un-acetylated compounds. Thus, we suggest that (E)-3,4-diacetoxystyryl sulfones and sulfoxides may have potential for the treatment of neurodegenerative disorders, especially Parkinson’s disease.     

Toxicity of 6-hydroxydopamine: live cell imaging of cytoplasmic redox flux

Oxidative stress contributes to several debilitating neurodegenerative diseases. To facilitate direct monitoring of the cytoplasmic oxidation state in neuronal cells, we have developed roTurbo by including several mutations: F223R, A206K, and six of the mutations for superfolder green fluorescent protein. Thus we have generated an improved redox sensor that is much brighter in cells and oxidizes more readily than roGFP2. Cytoplasmic expression of the sensor demonstrated the temporal pattern of 6-hydroxydopamine (6-OHDA) induced oxidative stress in a neuroblastoma cell line (SH-SY5Y). Two distinct oxidation responses were identified in SH-SY5Y cells but a single response observed in cells lacking monoamine transporters (HEK293). While both cell lines exhibited a rapid transient oxidation in response to 6-OHDA, a second oxidative response coincident with cell death was observed only in SH-SY5Y cells, indicating an intracellular metabolism of 6-OHDA, and or its metabolites are involved. In contrast, exogenously applied hydrogen peroxide induced a cellular oxidative response similar to the first oxidation peak, and cell loss was minimal. Glucose deprivation enhanced the oxidative stress induced by 6-OHDA, confirming the pivotal role played by glucose in maintaining a reduced cytoplasmic environment. While these studies support previous findings that catecholamine auto-oxidation products cause oxidative stress, our findings also support studies indicating 6-OHDA induces lethal oxidative stress responses unrelated to production of hydrogen peroxide. Finally, temporal imaging revealed the sporadic nature of the toxicity induced by 6-OHDA in neuroblastoma cells.     

Exploring marine resources against neurological disorders– the neuroprotective and anti-inflammatory potential of the brown seaweed Bifurcaria bifurcata

Oxidative stress is strongly involved in the pathogenesis of neurodegenerative diseases, like Parkinson´s disease (PD). Particularly, an excess of reactive oxygen species (ROS) released by the cells promotes an oxidative stress condition, which is a main cause of tissue injury leading to nervous system dysfunction. In this work, the antioxidant, neuroprotective and anti-inflammatory activities of different fractions from the brown seaweed Bifurcaria bifurcata are presented and related with their chemical profile. The antioxidant capacity was evaluated by the Folin-Ciocalteu method, 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, ferric reducing antioxidant power (FRAP) and oxygen radical absorbance capacity (ORAC) assays. Neuroprotective capacity was evaluated to prevent neurological cell death mediated by the neurotoxin 6-hydroxydopamine (6-OHDA) on SH-SY5Y cells, and their anti-inflammatory effects on RAW 264.7 macrophages. The ethyl acetate fractions (100 µg mL^?1) exhibited significant antioxidant and neuroprotective activities in the in vitro models assayed. Furthermore, two of the most polar fractions obtained with methanol and water also evidenced a significant neuroprotective potential. Bifurcaria bifurcata fractions treatment decreased ROS production, mitochondrial dysfunction, and Caspase-3 activity. Regarding the anti-inflammatory potential, five fractions (100 µg mL^?1) inhibited nitric oxide (NO) production and reduced the interleukin – 6 (IL-6) and tumor necrosis factor (TNF-α) levels. Mannitol, identified as the major component of the most bioactive fraction, protected SH-SY5Y cells against the 6-OHDA neurotoxicity mediating ROS generation mitigation, mitochondrial dysfunction, and DNA damage, together with the Caspase-3 activity inhibition. Results suggest that B. bifurcata is a relevant source of neuroprotective agents, with particular interest for preventive therapeutics.

     

The neuroprotective effect of Activin A and B: implication for neurodegenerative diseases

Activin is a member of the transforming growth factor-beta superfamily which comprises a growing list of multifunctional proteins that function as modulators of cell proliferation, differentiation, hormone secretion and neuronal survival. This study examined the neuroprotective effect of both Activin A and B in serum withdrawal and oxidative stress apoptotic cellular models and investigated the expression of pro- and anti-apoptotic proteins, which may account for the mechanism of Activin-induced neuroprotection. Here, we report that recombinant Activin A and B are neuroprotective against serum deprivation- and toxin- [either the parkinsonism-inducing neurotoxin, 6-hydroxydopamine (6-OHDA) or the peroxynitrite donor, 3-(4-morpholinyl) sydnonimine hydrochloride (SIN-1)] induced neuronal death in human SH-SY5Y neuroblastoma cells. Furthermore, we demonstrate for the first time that transient transfection with Activin betaA or betaB significantly protect SH-SY5Y and rat pheochromocytoma PC12 cells against serum withdrawal-induced apoptosis. This survival effect is mediated by the Bcl-2 family members and involves inhibition of caspase-3 activation; reduction of cleaved poly-ADP ribose polymerase and phosphorylated H2A.X protein levels and elevation of tyrosine hydroxylase expression. These results indicate that both Activin-A and -B share the potential to induce neuroprotective activity and thus may have positive impact on aging and neurodegenerative diseases to retard the accelerated rate of neuronal degeneration.     

Baicalein attenuates 6-hydroxydopamine-induced neurotoxicity in SH-SY5Y cells

It has been suggested that baicalein, a flavonoid obtained from the Scutellaria root, exerts a protective role on neurons against several neuronal insults. However, the protective mechanisms underlying this protective effect remain largely unknown. Our results indicate that baicalein protects SH-SY5Y cells, a dopaminergic neuronal cell line, from 6-hydroxydopamine (6-OHDA)-induced damage by the attenuation of reactive oxygen species (ROS). In order to determine the effects of baicalein on mitochondrial events, mitochondrial membrane potentials (deltapsim) and caspase cascades downstream of mitochondria were assessed. Baicalein inhibited the collapse of deltapsim, suggesting that baicalein reduces the mitochondrial dysfunction associated with 6-OHDA treatment. Baicalein also inhibited caspase-9 and caspase-3 activation, which can be triggered by mitochondrial malfunctions. Furthermore, baicalein induced a significant reduction in the level of phospho-JNK, which is known as an apoptotic mediator in 6-OHDA-induced neuronal cell death. Our results indicate that baicalein protects neurons from the deleterious effects of 6-OHDA via the attenuation of oxidative stress, mitochondrial dysfunction, caspase activity, and JNK activation.     

Assessment of novel azaanthraquinone derivatives as potent multi-target inhibitors of inflammation and amyloid-β aggregation in Alzheimer’s disease

A series of 6-substituted azaanthraquinone derivatives have been designed, synthesized, and their anti-inflammatory activities, antiaggregation effects on β-amyloid proteins, anticholinesterase and neuroprotective activity were tested. The new derivatives strongly suppressed NO and iNOS production and modulate the production of cytokines by decreasing TNF-a, IL-1β and IL-6 formation in lipopolysaccharide (LPS)-activated RAW 264.7 macrophages. Meanwhile, the derivatives exhibited a significant in vitro inhibitory activity toward the self-induced Aβ aggregation. While, treatment of SH-SY5Y cells overexpressing the Swedish mutant form of human b-amyloid precursor protein (APPsw) with derivatives was associated with significant reduction of Aβ42 secretion levels. Moreover, the derivatives exhibited moderate inhibitory potency toward acetylcholinesterase (AChE) and butyrylcholinesterase (BChE). Further investigations indicated that compound 7b could attenuate H₂O₂-induced neurotoxicity toward SH-SY5Y neuroblastoma cells and half of the synthetic compounds were predicted to be able to cross the blood–brain barrier (BBB) to reach their targets in the central nervous system (CNS) according to a parallel artificial membrane permeation assay for BBB. Taken together, azaanthraquinone derivatives targeting multiple pathogenetic factors deserves further investigation for prevention and treatment of AD.