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Comparative Study
. 2009 Jul;157(6):1072-84.
doi: 10.1111/j.1476-5381.2009.00213.x. Epub 2009 May 21.

Clovamide and rosmarinic acid induce neuroprotective effects in in vitro models of neuronal death (VSports)

S Fallarini  1 , G Miglio, T Paoletti, A Minassi, A Amoruso, C Bardelli, S Brunelleschi, G Lombardi
Affiliations
Comparative Study

Clovamide and rosmarinic acid induce neuroprotective effects in in vitro models of neuronal death

S Fallarini et al. Br J Pharmacol. 2009 Jul.

Abstract

Background and purpose: Phenolic compounds exert cytoprotective effects; our purpose was to investigate whether the isosteric polyphenolic compounds clovamide and rosmarinic acid are neuroprotective VSports手机版. .

Experimental approach: Three in vitro models of neuronal death were selected: (i) differentiated SH-SY5Y human neuroblastoma cells exposed to tert-butylhydroperoxide (t-BOOH), for oxidative stress; (ii) differentiated SK-N-BE(2) human neuroblastoma cells treated with L-glutamate, for excitotoxicity; and (iii) differentiated SH-SY5Y human neuroblastoma cells exposed to oxygen-glucose deprivation/reoxygenation, for ischaemia-reperfusion. Cell death was evaluated by lactate dehydrogenase measurements in the cell media, while the mechanisms underlying the effects by measuring: (i) t-BOOH-induced glutathione depletion and increase in lipoperoxidation; and (ii) L-glutamate-induced intracellular Ca(2+) overload (fura-2 method) and inducible gene expression (c-fos, c-jun), by reverse transcriptase-PCR. The ability of compounds to modulate nuclear factor-kappaB and peroxisome proliferator-activated receptor-gamma activation was evaluated by Western blot in SH-SY5Y cells not exposed to harmful stimuli V体育安卓版. .

Key results: Both clovamide and rosmarinic acid (10-100 micromol x L(-1)) significantly protected neurons against insults with similar potencies and efficacies. The EC(50) values were in the low micromolar range (0 V体育ios版. 9-3. 7 micromol x L(-1)), while the maximal effects ranged from 40% to -60% protection from cell death over untreated control at 100 micromol x L(-1). These effects are mediated by the prevention of oxidative stress, intracellular Ca(2+) overload and c-fos expression. In addition, rosmarinic acids inhibited nuclear factor-kappaB translocation and increased peroxisome proliferator-activated receptor-gamma expression in SH-SY5Y cells not exposed to harmful stimuli. .

Conclusion and implications: Clovamide and rosmarinic acid are neuroprotective compounds of potential use at the nutritional/pharmaceutical interface VSports最新版本. .

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Figures

Figure 1
Figure 1
Chemical structure of the compounds used in this study.
Figure 2
Figure 2
Concentration–response curves of the neuroprotective effects of clovamide or rosmarinic acid. (A) Effects of increasing concentrations of clovamide or rosmarinic acid (0.1–100 µmol·L−1) on t-BOOH (100 µmol·L−1; 3 h)-induced cell death, expressed as percentage of LDH released from damaged cells over that released from cells treated with DMSO alone (control). (B) Effects of increasing concentrations of clovamide or rosmarinic acid (0.1–100 µmol·L−1) on L-glutamate (1 mmol·L−1; 24 h)-induced cell death, expressed as percentage of LDH released from damaged cells over that released from cells treated with DMSO alone (control). (C and D) Effects of increasing concentrations of clovamide or rosmarinic acid (0.1–100 µmol·L−1) on OGD- (5 h) (C) or OGD (5 h)/reoxygenation (20 h) (D)-induced cell death, expressed as percentage of LDH released from damaged cells over that released from cells treated with DMSO alone (control). Vitamin E (VitE; 50 µmol·L−1) and (+)–MK 801 (MK-801; 1 µmol·L−1) were used as internal positive controls. The data represent mean ± SEM of at least six experiments run in triplicate. #P < 0.01 versus cells treated with DMSO alone (control); **P < 0.01, *P < 0.05 versus t-BOOH (A)-, L-glutamate (B)-, or OGD (C and D)-treated cells. DMSO, dimethylsulphoxide; LDH, lactate dehydrogenase; OGD, oxygen-glucose deprivation; t-BOOH, tert-butylhydroperoxide.
Figure 3
Figure 3
Antioxidant effects of clovamide or rosmarinic acid. (A) Ability of increasing concentrations (0.1–100 µmol·L−1) of either clovamide or rosmarinic acid in reducing t-BOOH-induced GSH decrease, expressed as percentage of GSH over cells treated with DMSO alone (control). (B) Ability of increasing concentrations (0.1–100 µmol·L−1) of either clovamide or rosmarinic acid in reducing lipoperoxidation (TBARS increase), expressed as percentage of the TBARS level in cells treated with DMSO alone (control). Vitamin E (VitE; 50 µmol·L−1) was used as internal positive control. The data represent mean ± SEM of at least five experiments run in triplicate. #P < 0.01 versus cells treated with DMSO alone (control); **P < 0.01 versus t-BOOH-treated cells. DMSO, dimethylsulphoxide; GSH, glutathione; t-BOOH, tert-butylhydroperoxide; TBARS, thiobarbituric acid-reacting substances.
Figure 4
Figure 4
Effects of clovamide or rosmarinic acid on the [Ca2+]i increase induced by L-glutamate treatment. [Ca2+]i was measured at single cell level in fura-2/acetoxymethyl ester-loaded cells exposed to L-glutamate (1 mmol·L−1; 24 h) in the absence or presence of 10 µmol·L−1 clovamide or rosmarinic acid. The values are expressed as Fr (MFI) mean of 36–53 cells monitored. The data represent mean ± SEM of at least six experiments run in triplicate. #P < 0.01 versus cells treated with DMSO alone (control); *P < 0.05 versus L-glutamate-treated cells. [Ca2+]i, intracellular calcium concentration; DMSO, dimethylsulphoxide; MFI, mean fluorescence ratio.
Figure 5
Figure 5
Effects of clovamide or rosmarinic acid on L-glutamate-induced c-fos and c-jun gene expression. (A) c-fos and c-jun gene expression was quantified by semi-quantitative RT-PCR in SK-N-BE(2) cells untreated or treated with L-glutamate (1 mmol·L−1; 24 h) in the absence or presence of increasing concentration (0.1–100 µmol·L−1) of clovamide or rosmarinic acid. The extracted total mRNA was reverse transcribed into its related cDNA, and PCR was carried out to amplify c-fos and c-jun cDNA by using specific primers (see Table 1). Expression of GAPDH was used as a loading control. PCR products were visualized with ethidium bromide on a 1% agarose gel. (B) The signals are densitometrically analysed; data, calculated as mean ± SEM of at least four determinations, are expressed as the ratio of the signal obtained for each sample divided by that obtained for GAPDH in the same sample to permit between-sample comparisons. #P < 0.01 versus cells treated with DMSO alone (control); **P < 0.01 versus L-glutamate-treated cells. DMSO, dimethylsulphoxide; GAPDH, glyceraldehyde 3-phosphate dehydrogenase; RT-PCR, reverse transcriptase-PCR.
Figure 6
Figure 6
Effects of clovamide or rosmarinic acid on NF-κB activation in SH-SY5Y cells not exposed to harmful stimuli. Both compounds were used at 10 µmol·L−1; results are presented as the nuclear/cytoplasmic ratio of NF-κB p50 and p65 subunits (see Methods) of at least four determinations. *P < 0.05 versus cells treated with DMSO alone (control). DMSO, dimethylsulphoxide; NF-κB, nuclear factor-κB.
Figure 7
Figure 7
Effects of clovamide or rosmarinic acid on PPARγ expression in SH-SY5Y cells not exposed to harmful stimuli. (A) The endogenous PPARγ agonist 15-deoxy-Δ12,14-PGJ2, used as positive internal control, increases PPARγ expression in a concentration-dependent manner (2–20 µmol·L−1) in comparison with control. (B) Effects of 10 µmol·L−1 clovamide or rosmarinic acid and 20 µmol·L−1 15-deoxy-Δ12,14-PGJ2 on PPARγ expression. Results are presented as PPARγ/β-actin ratio (see Methods) of at least five determinations. *P < 0.05; **P < 0.01 versus cells treated with DMSO alone (control). 15-deoxy-Δ12,14-PGJ2, 15-deoxy-Δ12,14-prostaglandin J2; PPARγ, peroxisome proliferator-activated receptor-γ.
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References

    1. van Acker SA, Koymans LM, Bast A. Molecular pharmacology of vitamin E: structural aspects of antioxidant activity. Free Radic Biol Med. 1993;15:311–328. - PubMed
    1. Amoroso S, Gioielli A, Cataldi M, Di Renzo G, Annunziato L. In the neuronal cell line SH-SY5Y, oxidative stress-induced free radical overproduction causes cell death without any participation of intracellular Ca (2+) increase. Biochem Biophys Acta. 1999;1452:151–160. - PubMed
    1. Amoruso A, Bardelli C, Gunella G, Fresu LG, Ferrero V, Brunelleschi S. Quantification of PPAR-gamma protein in monocyte/macrophages from healthy smokers and non-smokers: a possible direct effect of nicotine. Life Sci. 2007;81:906–915. - PubMed
    1. Arlorio M, Locatelli M, Travaglia F, Coïsson J-D, Del Grosso E, Minassi A, et al. Roasting impact on the contents of clovamide (N-caffeoyl-L-DOPA) and the antioxidant activity of cocoa beans (Theobroma cacao L.) Food Chem. 2008;106:967–975.
    1. Baba S, Osakabe N, Natsume M, Terao J. Orally administered rosmarinic acid is present as the conjugated and/or methylated forms in plasma, and is degraded and metabolized to conjugated forms of caffeic acid, ferulic acid and m-coumaric acid. Life Sci. 2004;75:165–178. - PubMed (V体育官网入口)

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