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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 prevents 6-OHDA-induced apoptosis in SH-SY5Y cells. SH-SY5Y cells were incubated with various concentrations of ER for 24 h and then treated with 6-OHDA (200 μmol/L) for 2 h at 37 °C. After a further 15 h of incubation in medium without 6-OHDA, the apoptotic cells were determined by Annexin V staining and fluorescence microscopy as described in the Experimental Section. The values are expressed as a percentage of total number of apoptotic cells and shown as mean ± SEM of three/four independent experiments. **p < 0.01, ***p < 0.001 versus untreated cells at ANOVA with Dunnett post hoc test.
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f2-ijms-13-10899: ER prevents 6-OHDA-induced apoptosis in SH-SY5Y cells. SH-SY5Y cells were incubated with various concentrations of ER for 24 h and then treated with 6-OHDA (200 μmol/L) for 2 h at 37 °C. After a further 15 h of incubation in medium without 6-OHDA, the apoptotic cells were determined by Annexin V staining and fluorescence microscopy as described in the Experimental Section. The values are expressed as a percentage of total number of apoptotic cells and shown as mean ± SEM of three/four independent experiments. **p < 0.01, ***p < 0.001 versus untreated cells at ANOVA with Dunnett post hoc test.

Mentions: To ascertain whether the increases of the total GSH level and TAC recorded in SH-SY5Y cells could really translate into neuroprotective effects, we subsequently evaluated the ability of the SH-SY5Y cells under identical experimental conditions to prevent the increase of neuronal death, in terms of apoptosis, induced by 6-OHDA. As reported in Figure 2, the pre-treatment of SH-SY5Y cells for 24 h with ER led to a strong dose-dependent decrease of apoptotic cells elicited by 6-OHDA. The observed decrease of apoptosis was significant for all ER concentrations employed (1.25–5 μmol/L) and the maximum inhibition was ~59% with respect to cells treated with 6-OHDA alone. Further, the degree of apoptosis inhibition shown by pre-treatment of SH-SY5Y cells with similar concentrations of ER against 6-OHDA toxicity significantly correlated with elevation of total GSH levels (r = 0.8146, p < 0.05). In order to evaluate the role of GSH in ER neuroprotection, we used buthionine sulfoximine (BSO), which irreversibly inhibits gamma-glutamylcysteine synthetase, the first enzyme in the GSH biosynthesis pathway. The addition of BSO 400 μmol/L to treatment of SH-SY5Y cells with ER 5 μmol/L for 24 h abolished the antiapoptotic effects observed with ER alone against toxicity induced by 6-OHDA (sham: 3% ± 1%; BSO: 4% ± 1%; 6-OHDA: 16% ± 3%; 6-OHDA + ER: 7% ± 4%; 6-OHDA + ER + BSO: 13% ± 4%; results are expressed as percentage of apoptotic cells).


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 prevents 6-OHDA-induced apoptosis in SH-SY5Y cells. SH-SY5Y cells were incubated with various concentrations of ER for 24 h and then treated with 6-OHDA (200 μmol/L) for 2 h at 37 °C. After a further 15 h of incubation in medium without 6-OHDA, the apoptotic cells were determined by Annexin V staining and fluorescence microscopy as described in the Experimental Section. The values are expressed as a percentage of total number of apoptotic cells and shown as mean ± SEM of three/four independent experiments. **p < 0.01, ***p < 0.001 versus untreated cells at ANOVA with Dunnett post hoc test.
© Copyright Policy - open-access
Related In: Results  -  Collection

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

f2-ijms-13-10899: ER prevents 6-OHDA-induced apoptosis in SH-SY5Y cells. SH-SY5Y cells were incubated with various concentrations of ER for 24 h and then treated with 6-OHDA (200 μmol/L) for 2 h at 37 °C. After a further 15 h of incubation in medium without 6-OHDA, the apoptotic cells were determined by Annexin V staining and fluorescence microscopy as described in the Experimental Section. The values are expressed as a percentage of total number of apoptotic cells and shown as mean ± SEM of three/four independent experiments. **p < 0.01, ***p < 0.001 versus untreated cells at ANOVA with Dunnett post hoc test.
Mentions: To ascertain whether the increases of the total GSH level and TAC recorded in SH-SY5Y cells could really translate into neuroprotective effects, we subsequently evaluated the ability of the SH-SY5Y cells under identical experimental conditions to prevent the increase of neuronal death, in terms of apoptosis, induced by 6-OHDA. As reported in Figure 2, the pre-treatment of SH-SY5Y cells for 24 h with ER led to a strong dose-dependent decrease of apoptotic cells elicited by 6-OHDA. The observed decrease of apoptosis was significant for all ER concentrations employed (1.25–5 μmol/L) and the maximum inhibition was ~59% with respect to cells treated with 6-OHDA alone. Further, the degree of apoptosis inhibition shown by pre-treatment of SH-SY5Y cells with similar concentrations of ER against 6-OHDA toxicity significantly correlated with elevation of total GSH levels (r = 0.8146, p < 0.05). In order to evaluate the role of GSH in ER neuroprotection, we used buthionine sulfoximine (BSO), which irreversibly inhibits gamma-glutamylcysteine synthetase, the first enzyme in the GSH biosynthesis pathway. The addition of BSO 400 μmol/L to treatment of SH-SY5Y cells with ER 5 μmol/L for 24 h abolished the antiapoptotic effects observed with ER alone against toxicity induced by 6-OHDA (sham: 3% ± 1%; BSO: 4% ± 1%; 6-OHDA: 16% ± 3%; 6-OHDA + ER: 7% ± 4%; 6-OHDA + ER + BSO: 13% ± 4%; results are expressed as percentage of apoptotic cells).

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