Article

Proteomic screening of antioxidant effects exhibited by Radix Salvia miltiorrhiza aqueous extract in cultured rat aortic smooth muscle cells under homocysteine treatment

Graduate Institute of Clinical Medical Sciences, Kaohsiung Division, Chang Gung University, Kaohsiung, Taiwan.
Journal of ethnopharmacology (Impact Factor: 2.32). 05/2009; 124(3):463-74. DOI: 10.1016/j.jep.2009.05.020
Source: PubMed

ABSTRACT Still little is known about the cellular mechanisms that contribute to the attenuated proliferation of aortic smooth muscle cells under the influence of the oxidative stress factors such as homocysteine (Hcy). Thus, we aimed to evaluate whether Salvia miltiorrhiza Bunge (Labiatae), a Chinese medicinal herb widely used in folk medicine for therapy of variety of human cardiovascular disorders would modulate this Hcy promoted growth effect in model animal aortic cells system.
The Salvia miltiorrhiza roots aqueous extract (SMAE) containing 3,4-dihydroxybenzoic acid, 3,4-dihydroxyphenyl lactic acid and salvianolic acid B, as confirmed by narrow-bore HPLC analyses with binary gradient elution was used in variable concentrations for the treatment of the rat aortic smooth muscle A10 cells under Hcy stimulation. Two-dimensional electrophoresis (2-DE) coupled with MALDI-TOF mass spectrometry was applied for the elucidation of protein changes characterizing the response of the rat A10 cells into the Hcy-induced oxidative stress.
This study showed that a low dose (0.015 mg/mL) of the SMAE significantly inhibited growth (>60%, p<0.05) of the Hcy stimulated rat A10 cells. In addition, concentration of intracellular reactive oxygen species (ROS) obviously decreased in the rat A10 cells after its incubation with SMAE in terms of catalase increasing activity. Next, marked down-regulation of protein kinase C beta-1 (PKC beta-1) and phosphorylated mitogen-activated protein kinase (p-MAPK) expression suggest that observed inhibitory effect of the polyphenol-rich SMAE on the Hcy-induced growth of rat A10 cells was realized via the PKC/p44/42 MAPK-dependent pathway. The intensity changes of 10 protein spots in response of the rat A10 cells to the Hcy-induced oxidative damage as alpha-4-tropomyosin, vimentin, F1F0-ATP synthase (beta subunit), glucose regulated protein 75 (GRP75), actin (fragment), prohibitin, capping protein, plakoglobin, endoplasmic reticulum protein (ERp29), and peptidylprolyl isomerase A (PPIase A), were detected with statistical significance (p<0.05). Meanwhile, it was showed that used here SMAE resist carbonylation of specific cytoskeleton and chaperone proteins as vimentin, alpha-4-tropomyosin and GRP75, respectively, leading to phenotype transformations in the rat A10 cells.
These data suggest that applied here SMAE exerts its protective effect through circulating ROS suppression and subsequent modulation of protein carbonylation in rat aortic smooth muscle A10 cells. Redox-proteomics protocol highlighted in this study may be applicable in facilitating the assessing potential novel molecular therapeutic targets to reduce cardiovascular risk related with elevated Hcy levels in various human populations and elucidating new mechanisms through which protein functions can be regulated by the redox status with the use of naturally occurring antioxidants.

0 Bookmarks
 · 
117 Views
  • Source
    [Show abstract] [Hide abstract]
    ABSTRACT: Hepatic ischemia-reperfusion (IR) injury is a common clinical problem and reactive oxygen species (ROS) may be a contributing factor on IR injury. The current study evaluates the potential protective effect of saffron ethanol extract (SEE) in a rat model upon hepatic IR injury. Caspases 3 and TUNEL's results showed increased cell death in the IR samples; reversely, minor apoptosis was detected in the SEE/IR group. Pretreatment with SEE significantly restored the content of antioxidant enzymes (SOD1 and catalase) and remarkably inhibited the intracellular ROS concentration in terms of reducing p47phox translocation. Proteome tools revealed that 20 proteins were significantly modulated in protein intensity between IR and SEE/IR groups. Particularly, SEE administration could attenuate the carbonylation level of several chaperon proteins. Network analysis suggested that saffron extract could alleviate IR-induced endoplasmic reticulum (ER) stress and protein ubiquitination which finally lead to cell apoptosis. Taken together, SEE could reduce hepatic IR injury through modulating protein oxidation and our results might help to develop novel therapeutic strategies against ROS-caused diseases. This article is protected by copyright. All rights reserved.
    Proteomics 05/2013; 13(15). DOI:10.1002/pmic.201200551 · 3.97 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: It is well documented that oxidative stress has been implicated as one of the leading causes for brain damage induced by cerebral ischemia/reperfusion (I/R). In this study, we assessed the potential cerebraprotective and antioxidant effects of the polysaccharides (DSP) from the roots of Salvia miltiorrhiza against global cerebral I/R injury in an animal model established by blocking bilateral common carotid arteries (BCCA) for 30minutes followed by reperfusion for 24hours. The rats were treated with their respective treatments for 10 days prior to the BCCA occlusion. After that, animals were sacrificed by decapitation, brain was removed, and various biochemical estimations, cerebral edema and assessment of cerebral infarct size were carried out. We found that pretreatment with DSP significantly decreased the neurological deficit scores, percentage of infarction and brain edema and the generation of mitochondrial reactive oxygen species (ROS). Moreover, DSP also increased mitochondria superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GSH-Px) activities and reduced malondialdehyde (MDA) production in cerebral ischemia brain. In conclusion, these studies suggest that pretreatment with DSP provides significant protection against cerebral I/R injury in rats most probably by virtue of its antioxidant property.
    International journal of biological macromolecules 06/2013; 60. DOI:10.1016/j.ijbiomac.2013.05.035 · 2.37 Impact Factor
  • [Show abstract] [Hide abstract]
    ABSTRACT: Endoplasmic reticulum protein 29 (ERp29) belongs to the redox-inactive PDI-Dβ-subfamily of PDI-proteins. ERp29 is expressed in all mammalian tissues examined. Especially high levels of expression were observed in secretory tissues and in some tumors. However, the biological role of ERp29 remains unclear. In the present study we show, by using thyrocytes and primary dermal fibroblasts from adult ERp29(-/-) mice, that ERp29 deficiency affects the activation of the ATF6-CHOP-branch of unfolded protein response (UPR) without influencing the function of other UPR branches, like the ATF4-eIF2α-XBP1 signaling pathway. As a result of impaired ATF6 activation, dermal fibroblasts and adult thyrocytes from ERp29(-/-) mice display significantly lower apoptosis sensitivities when treated with tunicamycin and hydrogen peroxide. However, in contrast to previous reports, we could demonstrate that ERp29 deficiency does not alter thyroglobulin expression levels. Therefore, our study suggests that ERp29 acts as an escort factor for ATF6 and promotes its transport from ER to Golgi apparatus under ER stress conditions.
    Apoptosis 12/2013; DOI:10.1007/s10495-013-0961-0 · 3.61 Impact Factor