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Neuroprotective effects of erucin against 6-hydroxydopamine-induced oxidative damage in a dopaminergic-like neuroblastoma cell line.

Tarozzi A, Morroni F, Bolondi C, Sita G, Hrelia P, Djemil A, Cantelli-Forti G - Int J Mol Sci (2012)

Bottom Line: The increase of GSH levels was associated with an increase in the resistance of SH-SY5Y cells to neuronal death, in terms of apoptosis, induced by 6-hydroxydopamine (6-OHDA).Last, the antiapoptotic and antioxidant effects of ER were abolished by buthionine sulfoximine, supporting the main role of GSH in the neuroprotective effects recorded by ER.These results suggest that ER may prevent the oxidative damage induced by 6-OHDA.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Alma Mater Studiorum-University of Bologna, via Irnerio 48, 40126 Bologna, Italy; E-Mails: fabiana.morroni@unibo.it (F.M.); cecilia.bolondi2@unibo.it (C.B.); giulia.sita2@unibo.it (G.S.); patrizia.hrelia@unibo.it (P.H.); alice.djemil@studio.unibo.it (A.D.); giorgio.cantelliforti@unibo.it (G.C.-F.).

ABSTRACT
Oxidative stress (OS) contributes to the cascade leading to the dysfunction or death of dopaminergic neurons during Parkinson's disease (PD). A strategy to prevent the OS of dopaminergic neurons may be the use of phytochemicals as inducers of endogenous antioxidants and phase 2 enzymes. In this study, we demonstrated that treatment of the dopaminergic-like neuroblastoma SH-SY5Y cell line with isothiocyanate erucin (ER), a compound of cruciferous vegetables, resulted in significant increases of both total glutathione (GSH) levels and total antioxidant capacity at the cytosolic level. The increase of GSH levels was associated with an increase in the resistance of SH-SY5Y cells to neuronal death, in terms of apoptosis, induced by 6-hydroxydopamine (6-OHDA). The pretreatment of SH-SY5Y cells with ER was also shown to prevent the redox status impairment, in terms of intracellular ROS and O(2) (•-) formation, and loss of mitochondrial membrane potential, early events that are initiators of the apoptotic process, induced by 6-OHDA. Last, the antiapoptotic and antioxidant effects of ER were abolished by buthionine sulfoximine, supporting the main role of GSH in the neuroprotective effects recorded by ER. These results suggest that ER may prevent the oxidative damage induced by 6-OHDA.

No MeSH data available.


Related in: MedlinePlus

ER counteracts 6-OHDA-induced intracellular O2•− formation in SH-SY5Y cells. (a) SH-SY5Y cells were incubated with ER (5 μmol/L) in absence or presence of BSO (400 μmol/L) for 24 h and then treated with 6-OHDA (200 μmol/L) for 2 h. At the end of incubation, O2−− formation was determined using a fluorescence probe, DHE, as described in the Experimental Section. Four randomly selected areas with 50–100 cells in each were analyzed under a fluorescence microscope and the values obtained are expressed as densitometry/cell. Values are shown as mean ± SEM of four independent experiments: ∘∘∘ p < 0.001 versus untreated cells, **p < 0.01 versus cells treated with 6-OHDA at ANOVA with Bonferroni post hoc test; (b) representative images of O2•− formation. Scale bars: 100 μm.
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f5-ijms-13-10899: ER counteracts 6-OHDA-induced intracellular O2•− formation in SH-SY5Y cells. (a) SH-SY5Y cells were incubated with ER (5 μmol/L) in absence or presence of BSO (400 μmol/L) for 24 h and then treated with 6-OHDA (200 μmol/L) for 2 h. At the end of incubation, O2−− formation was determined using a fluorescence probe, DHE, as described in the Experimental Section. Four randomly selected areas with 50–100 cells in each were analyzed under a fluorescence microscope and the values obtained are expressed as densitometry/cell. Values are shown as mean ± SEM of four independent experiments: ∘∘∘ p < 0.001 versus untreated cells, **p < 0.01 versus cells treated with 6-OHDA at ANOVA with Bonferroni post hoc test; (b) representative images of O2•− formation. Scale bars: 100 μm.

Mentions: To clarify the antioxidant and protective mechanisms underlying the antiapoptotic effects of ER, we also evaluated its ability to prevent the redox status impairment, in terms of MMP loss, and intracellular ROS and O2•− formation, early events that are initiators of the apoptotic process, induced by 6-OHDA. As reported in Figure 3, pre-treatment of SH-SY5Y cells with ER (5 μmol/L) for 24 h showed a significant inhibition of MMP loss elicited by 6-OHDA. Further, the same treatment with ER 5 μmol/L also showed a strong inhibitory effect on 6-OHDA-induced total ROS and O2•− formation in SH-SY5Y cells using fluorescent probe 2′,7′-dichlorodihydroflurescein diacetate (DCFH-DA) and dihydroethidium (DHE), respectively (Figures 4 and 5). Remarkably, this finding was abolished by adding BSO (400 μmol/L) to the treatment with ER and 6-OHDA, supporting the predominant role of GSH in the indirect antioxidant effects displayed by ER (Figures 3–5).


Neuroprotective effects of erucin against 6-hydroxydopamine-induced oxidative damage in a dopaminergic-like neuroblastoma cell line.

Tarozzi A, Morroni F, Bolondi C, Sita G, Hrelia P, Djemil A, Cantelli-Forti G - Int J Mol Sci (2012)

ER counteracts 6-OHDA-induced intracellular O2•− formation in SH-SY5Y cells. (a) SH-SY5Y cells were incubated with ER (5 μmol/L) in absence or presence of BSO (400 μmol/L) for 24 h and then treated with 6-OHDA (200 μmol/L) for 2 h. At the end of incubation, O2−− formation was determined using a fluorescence probe, DHE, as described in the Experimental Section. Four randomly selected areas with 50–100 cells in each were analyzed under a fluorescence microscope and the values obtained are expressed as densitometry/cell. Values are shown as mean ± SEM of four independent experiments: ∘∘∘ p < 0.001 versus untreated cells, **p < 0.01 versus cells treated with 6-OHDA at ANOVA with Bonferroni post hoc test; (b) representative images of O2•− formation. Scale bars: 100 μm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC3472719&req=5

f5-ijms-13-10899: ER counteracts 6-OHDA-induced intracellular O2•− formation in SH-SY5Y cells. (a) SH-SY5Y cells were incubated with ER (5 μmol/L) in absence or presence of BSO (400 μmol/L) for 24 h and then treated with 6-OHDA (200 μmol/L) for 2 h. At the end of incubation, O2−− formation was determined using a fluorescence probe, DHE, as described in the Experimental Section. Four randomly selected areas with 50–100 cells in each were analyzed under a fluorescence microscope and the values obtained are expressed as densitometry/cell. Values are shown as mean ± SEM of four independent experiments: ∘∘∘ p < 0.001 versus untreated cells, **p < 0.01 versus cells treated with 6-OHDA at ANOVA with Bonferroni post hoc test; (b) representative images of O2•− formation. Scale bars: 100 μm.
Mentions: To clarify the antioxidant and protective mechanisms underlying the antiapoptotic effects of ER, we also evaluated its ability to prevent the redox status impairment, in terms of MMP loss, and intracellular ROS and O2•− formation, early events that are initiators of the apoptotic process, induced by 6-OHDA. As reported in Figure 3, pre-treatment of SH-SY5Y cells with ER (5 μmol/L) for 24 h showed a significant inhibition of MMP loss elicited by 6-OHDA. Further, the same treatment with ER 5 μmol/L also showed a strong inhibitory effect on 6-OHDA-induced total ROS and O2•− formation in SH-SY5Y cells using fluorescent probe 2′,7′-dichlorodihydroflurescein diacetate (DCFH-DA) and dihydroethidium (DHE), respectively (Figures 4 and 5). Remarkably, this finding was abolished by adding BSO (400 μmol/L) to the treatment with ER and 6-OHDA, supporting the predominant role of GSH in the indirect antioxidant effects displayed by ER (Figures 3–5).

Bottom Line: The increase of GSH levels was associated with an increase in the resistance of SH-SY5Y cells to neuronal death, in terms of apoptosis, induced by 6-hydroxydopamine (6-OHDA).Last, the antiapoptotic and antioxidant effects of ER were abolished by buthionine sulfoximine, supporting the main role of GSH in the neuroprotective effects recorded by ER.These results suggest that ER may prevent the oxidative damage induced by 6-OHDA.

View Article: PubMed Central - PubMed

Affiliation: Department of Pharmacology, Alma Mater Studiorum-University of Bologna, via Irnerio 48, 40126 Bologna, Italy; E-Mails: fabiana.morroni@unibo.it (F.M.); cecilia.bolondi2@unibo.it (C.B.); giulia.sita2@unibo.it (G.S.); patrizia.hrelia@unibo.it (P.H.); alice.djemil@studio.unibo.it (A.D.); giorgio.cantelliforti@unibo.it (G.C.-F.).

ABSTRACT
Oxidative stress (OS) contributes to the cascade leading to the dysfunction or death of dopaminergic neurons during Parkinson's disease (PD). A strategy to prevent the OS of dopaminergic neurons may be the use of phytochemicals as inducers of endogenous antioxidants and phase 2 enzymes. In this study, we demonstrated that treatment of the dopaminergic-like neuroblastoma SH-SY5Y cell line with isothiocyanate erucin (ER), a compound of cruciferous vegetables, resulted in significant increases of both total glutathione (GSH) levels and total antioxidant capacity at the cytosolic level. The increase of GSH levels was associated with an increase in the resistance of SH-SY5Y cells to neuronal death, in terms of apoptosis, induced by 6-hydroxydopamine (6-OHDA). The pretreatment of SH-SY5Y cells with ER was also shown to prevent the redox status impairment, in terms of intracellular ROS and O(2) (•-) formation, and loss of mitochondrial membrane potential, early events that are initiators of the apoptotic process, induced by 6-OHDA. Last, the antiapoptotic and antioxidant effects of ER were abolished by buthionine sulfoximine, supporting the main role of GSH in the neuroprotective effects recorded by ER. These results suggest that ER may prevent the oxidative damage induced by 6-OHDA.

No MeSH data available.


Related in: MedlinePlus