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ITRAQ-Based Proteomics Analysis Reveals the Effect of Rhubarb in Rats with Ischemic Stroke.

ABSTRACT: Background. Rhubarb, a traditional Chinese medicine, promotes viscera and remove blood stasis. Rhubarb is skilled in smoothening meridians, improving blood circulation which exhibits better efficacy on cerebral ischemic stroke. In this study, we aimed to analyze the underlying mechanisms of rhubarb which treated rats of middle cerebral artery occlusion (MCAO) model according to an iTRAQ-based proteomics and bioinformatics analysis. 30 rats were randomly allocated into three groups including sham group (SG), model group (MG), and rhubarb group (RG). Rhubarb group was given a gavage of rhubarb decoction at dose of 3 g/kg and the remaining groups were prepared with normal saline by gavage. Rats from MG and RG were induced into MCAO model. The effects of rhubarb were estimated by Modified Neurological Severity Score (mNSS) and cerebral infarct volume. The brain tissues were measured via the quantitative proteomic approach of iTRAQ coupled to liquid chromatography-tandem mass spectrometry (LC-MS/MS). Furthermore, the bioinformatics analysis of overlapping differentially expression proteins (DEPs) was conducted by DAVID, KEGG, and Cytoscape. Specific selective DEPs were validated by Western blotting. Rats treated with rhubarb after MCAO showed a significant reduction on mNSS and cerebral infarct volume compared with MG. In MG versus SG and RG versus MG, we identified a total of 4578 proteins, of which 287 were DEPs. There were 76 overlapping DEPs between MG versus SG and RG versus MG. Through bioinformatics analysis, 14 associated pathways were searched including cGMP-PKG signaling pathway, tuberculosis, synaptic vesicle cycle, amyotrophic lateral sclerosis, long-term potentiation, and so on. 76 overlapping DEPs mainly involved synaptic vesicle cycling biological processes based on GO annotation. Further, the selective overlapping DEPs were verified at the protein level by using Western blotting. Our present study reveals that rhubarb highlights promising neuroprotective effect. Rhubarb exerts novel therapeutic action via modulating multiple proteins, targets, and pathways.

SUBMITTER: Lin X

PROVIDER: S-EPMC6077657 | biostudies-literature | 2018

REPOSITORIES: biostudies-literature

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iTRAQ-Based Proteomics Analysis Reveals the Effect of Rhubarb in Rats with Ischemic Stroke.

Lin Xiangping X Liu Tao T Li Pengfei P He Zehui Z Zhong Yuanyuan Y Cui Hanjin H Luo Jiekun J Wang Yang Y Tang Tao T

BioMed research international 20180716

<i>Background.</i> Rhubarb, a traditional Chinese medicine, promotes viscera and remove blood stasis. Rhubarb is skilled in smoothening meridians, improving blood circulation which exhibits better efficacy on cerebral ischemic stroke. In this study, we aimed to analyze the underlying mechanisms of rhubarb which treated rats of middle cerebral artery occlusion (MCAO) model according to an iTRAQ-based proteomics and bioinformatics analysis. 30 rats were randomly allocated into three groups includi ...[more]

PMID: 30112417

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Project description:Smooth muscle (SM) contraction is one of the important physiological functions of the human body, and SM abnormal contraction will induce many diseases. The phosphorylated regulatory light chains (p-RLC) play a decisive role in SM contraction, and dephosphorylation of p-RLC is an effective way to relax SM. Our previous study showed that the novel benzylisoquinoline alkaloid, neoliensinine (Neo), could relax microvascular SM contracted by KCl hyperpolarization. In this study, mesenteric capillaries isolated from 45 mice were divided into normal tension group (Control), 124 mM KCl induced contraction model group (Model), and KCl and Neo-treatment group (Drug). The dephosphorylation levels of RLC in the three groups were measured. Compared with the model group, the phosphorylation of RLC in the drug group was decreased dramatically as expected, suggesting that the relaxation effect of Neo was caused by downregulating p-RLC of microvessel SM. In order to fully understand its fundamental mechanism, our research focused on the identification of target proteins in mice with KCl-induced contractile mesenteric capillary. Isobaric tags for relative and absolute quantification (ITRAQ) tagging was carried out by nanospray liquid chromatography-tandem mass spectrometry. The results allowed the upregulation of 164 differential abundance proteins (DAPs) among the 3,474 protein abundance disturbances identified from the model/control samples. Further comparison showed that there were 16 DAP convergences associated with vascular SM contraction between the drug/model and the drug/control samples. Among them, two proteins with known function, PLCβ and RhoGEF12, were selected as target proteins of the relaxation effect of Neo. The two selective target DAPs were verified by Western blot at protein level. The results suggested that changes of the two proteins were consistent with that of the iTRAQ results. Our present work reveals that Neo relaxes vascular smooth muscle via inhibition of RLC phosphorylation, and PLCβ and RhoGEF12 may be potential biomarkers for evaluating the effects mediated by Neo.

Dataset Information

ITRAQ-Based Proteomics Analysis Reveals the Effect of Rhubarb in Rats with Ischemic Stroke.

Publications

iTRAQ-Based Proteomics Analysis Reveals the Effect of Rhubarb in Rats with Ischemic Stroke.

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