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Eriocaulon buergerianum extract protects PC12 cells and neurons in zebrafish against 6-hydroxydopamine-induced damage.

Wang M, Zhang Z, Cheang LC, Lin Z, Lee SM - Chin Med (2011)

Bottom Line: EBE inhibited the 6-OHDA-induced decrease in total distance of movement in zebrafish.EBE exhibited significant neuroprotective activities in zebrafish, including recovery of dopaminergic neuron loss caused by 6-OHDA in a dose-dependent manner in vivo, inhibition of 6-OHDA-induced decrease of total distance in movement in zebrafish.The iNOS-NO pathway may be involved.

View Article: PubMed Central - HTML - PubMed

Affiliation: State Key Laboratory of Quality Research in Chinese Medicine, Institute of Chinese Medical Sciences, University of Macau, Av, Padre Tomás Pereira, Taipa, Macao, China. SimonLee@umac.mo.

ABSTRACT

Background: Ericaulon buergerianum (Gujingcao) is an ophthalmic, anti-inflammatory and antimicrobial Chinese medicinal herb. This study aims to investigate the neuroprotective effects of Ericaulon buergerianum ethanol extract (EBE) and to elucidate its underlying action mechanism.

Methods: The viability of dopaminergic (DA) neuron in zebrafish was examined by anti-tyrosine hydroxylase (TH) immunostaining. The locomotor activity of zebrafish was assessed with a digital video tracking system. The viability and cellular damage of the PC12 cells were determined by MTT and LDH assays respectively. The nuclear morphological changes in apoptotic cells were evaluated with DNA staining by Hoechst 33342 dye. Intracellular nitric oxide (NO) was quantified by DAF-FM diacetate staining. The expression of inducible nitric oxide synthase (iNOS) was determined by Western blot.

Results: EBE inhibited the 6-OHDA-induced decrease in total distance of movement in zebrafish. Pretreatments of EBE (25, 50, 100 and 200 μg/ml) increased the viability of 6-OHDA-damaged PC12 cells in a dose dependent manner. Protection against 6-OHDA-induced nuclear fragmentation and accumulation of apoptotic bodies was also observed in EBE pretreated cells. Anti-oxidative (inhibition of NO production and iNOS expression in PC12 cells in vitro) activities of EBE are related to its neuroprotective effects in 6-OHDA-induced DA neuron damage.

Conclusion: EBE exhibited significant neuroprotective activities in zebrafish, including recovery of dopaminergic neuron loss caused by 6-OHDA in a dose-dependent manner in vivo, inhibition of 6-OHDA-induced decrease of total distance in movement in zebrafish. The iNOS-NO pathway may be involved.

No MeSH data available.


Related in: MedlinePlus

EBE down-regulated iNOS over-expression in PC12 cells stimulated by 6-OHDA. Cells were incubated with EBE (25, 50, 100 and 200 μg/ml) for 12 hours prior to a 6-hour 6-OHDA stimulation. (A) Western blot analysis showing 6-OHDA-induced iNOS over-expression was inhibited by pretreatment of different concentrations of EBE. (B) Densitometric analysis of iNOS expression with measurements of each blot expressed relative to that of the control. Three times independent experiments showed the same tendency of the iNOS expression.
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Figure 8: EBE down-regulated iNOS over-expression in PC12 cells stimulated by 6-OHDA. Cells were incubated with EBE (25, 50, 100 and 200 μg/ml) for 12 hours prior to a 6-hour 6-OHDA stimulation. (A) Western blot analysis showing 6-OHDA-induced iNOS over-expression was inhibited by pretreatment of different concentrations of EBE. (B) Densitometric analysis of iNOS expression with measurements of each blot expressed relative to that of the control. Three times independent experiments showed the same tendency of the iNOS expression.

Mentions: Figure 7 shows that 6-OHDA exposure led to a roughly 1.5-fold increase in NO production compared with the control group; similar elevation was also observed when SNP (sodium nitroprusside dehydrate, NO generator) was added. This increase in NO level was reduced in a concentration-dependent manner by pretreatment with EBE in a 25-200 μg/mL range for 12 hours (Figure 7E-H). Pretreatment with 250 μM L-NAME for 12 hours also significantly (P = 0.044) reduced this 6-OHDA-induced NO over-production. Pretreatment with higher concentrations (50, 100, 200 μg/ml) of EBE suppressed the elevated NO production more efficiently than L-NAME (Figure 7D). Furthermore, DAF-FM diacetate staining was photographed with a fluorescent microscope (Figure 7A-H). The expression of iNOS was up-regulated by 6-OHDA exposure in Western blot analysis and such up-regulation was prevented by pretreatment with EBE (Figure 8A).


Eriocaulon buergerianum extract protects PC12 cells and neurons in zebrafish against 6-hydroxydopamine-induced damage.

Wang M, Zhang Z, Cheang LC, Lin Z, Lee SM - Chin Med (2011)

EBE down-regulated iNOS over-expression in PC12 cells stimulated by 6-OHDA. Cells were incubated with EBE (25, 50, 100 and 200 μg/ml) for 12 hours prior to a 6-hour 6-OHDA stimulation. (A) Western blot analysis showing 6-OHDA-induced iNOS over-expression was inhibited by pretreatment of different concentrations of EBE. (B) Densitometric analysis of iNOS expression with measurements of each blot expressed relative to that of the control. Three times independent experiments showed the same tendency of the iNOS expression.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3108929&req=5

Figure 8: EBE down-regulated iNOS over-expression in PC12 cells stimulated by 6-OHDA. Cells were incubated with EBE (25, 50, 100 and 200 μg/ml) for 12 hours prior to a 6-hour 6-OHDA stimulation. (A) Western blot analysis showing 6-OHDA-induced iNOS over-expression was inhibited by pretreatment of different concentrations of EBE. (B) Densitometric analysis of iNOS expression with measurements of each blot expressed relative to that of the control. Three times independent experiments showed the same tendency of the iNOS expression.
Mentions: Figure 7 shows that 6-OHDA exposure led to a roughly 1.5-fold increase in NO production compared with the control group; similar elevation was also observed when SNP (sodium nitroprusside dehydrate, NO generator) was added. This increase in NO level was reduced in a concentration-dependent manner by pretreatment with EBE in a 25-200 μg/mL range for 12 hours (Figure 7E-H). Pretreatment with 250 μM L-NAME for 12 hours also significantly (P = 0.044) reduced this 6-OHDA-induced NO over-production. Pretreatment with higher concentrations (50, 100, 200 μg/ml) of EBE suppressed the elevated NO production more efficiently than L-NAME (Figure 7D). Furthermore, DAF-FM diacetate staining was photographed with a fluorescent microscope (Figure 7A-H). The expression of iNOS was up-regulated by 6-OHDA exposure in Western blot analysis and such up-regulation was prevented by pretreatment with EBE (Figure 8A).

Bottom Line: EBE inhibited the 6-OHDA-induced decrease in total distance of movement in zebrafish.EBE exhibited significant neuroprotective activities in zebrafish, including recovery of dopaminergic neuron loss caused by 6-OHDA in a dose-dependent manner in vivo, inhibition of 6-OHDA-induced decrease of total distance in movement in zebrafish.The iNOS-NO pathway may be involved.

View Article: PubMed Central - HTML - PubMed

Affiliation: State Key Laboratory of Quality Research in Chinese Medicine, Institute of Chinese Medical Sciences, University of Macau, Av, Padre Tomás Pereira, Taipa, Macao, China. SimonLee@umac.mo.

ABSTRACT

Background: Ericaulon buergerianum (Gujingcao) is an ophthalmic, anti-inflammatory and antimicrobial Chinese medicinal herb. This study aims to investigate the neuroprotective effects of Ericaulon buergerianum ethanol extract (EBE) and to elucidate its underlying action mechanism.

Methods: The viability of dopaminergic (DA) neuron in zebrafish was examined by anti-tyrosine hydroxylase (TH) immunostaining. The locomotor activity of zebrafish was assessed with a digital video tracking system. The viability and cellular damage of the PC12 cells were determined by MTT and LDH assays respectively. The nuclear morphological changes in apoptotic cells were evaluated with DNA staining by Hoechst 33342 dye. Intracellular nitric oxide (NO) was quantified by DAF-FM diacetate staining. The expression of inducible nitric oxide synthase (iNOS) was determined by Western blot.

Results: EBE inhibited the 6-OHDA-induced decrease in total distance of movement in zebrafish. Pretreatments of EBE (25, 50, 100 and 200 μg/ml) increased the viability of 6-OHDA-damaged PC12 cells in a dose dependent manner. Protection against 6-OHDA-induced nuclear fragmentation and accumulation of apoptotic bodies was also observed in EBE pretreated cells. Anti-oxidative (inhibition of NO production and iNOS expression in PC12 cells in vitro) activities of EBE are related to its neuroprotective effects in 6-OHDA-induced DA neuron damage.

Conclusion: EBE exhibited significant neuroprotective activities in zebrafish, including recovery of dopaminergic neuron loss caused by 6-OHDA in a dose-dependent manner in vivo, inhibition of 6-OHDA-induced decrease of total distance in movement in zebrafish. The iNOS-NO pathway may be involved.

No MeSH data available.


Related in: MedlinePlus