VDAC1 and RhoA in Plasma Exosomes Influence the Severity of Adolescent Idiopathic Scoliosis

Abstract Background: Adolescent idiopathic scoliosis (AIS) is the most common spine deformity, but biomarkers for its condition are lacking. Rhodopsin A (RhoA) and voltage-dependent anion-selective channel 1 (VDAC1) in plasma exosomes were defined as differentially expressed proteins between AIS patients and healthy controls. The purpose of this study was to assess exosomes as biomarkers for the occurrence and progression of AIS. Methods：We recruited 10 AIS patients and 8 healthy controls to detect expressed proteins from plasma by liquid chromatography coupled to tandem mass spectrometry. Plasma samples were analyzed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS). Pathway analysis identified that the VDAC1 and RhoA proteins were alterations expressed in the AIS patients, with the most different alteration was found in extracellular exosomes. Ultracentrifugation was carried out to isolate exosomes from plasma. Verification of the most differentially expressed protein was accessed by Western blot analysis and bioinformatics analysis was performed to predict the pathway of it.Results: 42 of significantly differentially expressed proteins were found in all subjects, and 17 proteins had significant difference. The differentially expressed proteins were enriched in plasma exosomes, and some proteins, such as FN1, were upregulated and others, such as VDAC1, RhoA and AHNAK, were downregulated in the AIS patients. Furthermore, ultracentrifugation was carried out to isolate exosomes from plasma, and RhoA and VDAC1 proteins in plasma exosomes were verified to downregulate by western blot. KEGG signaling pathways were used to predict potential pathways involved in the RhoA and VDAC1 proteins in the AIS patients. We found that the RhoA protein influences AIS probably through the chemokine signaling pathway, platelet activation and cAMP signaling pathway, and the VDAC1 protein is a key factor that participates in the necroptosis pathway, acting on the development of AIS.Conclusions: Consequently, this study mapped a profile of plasma protein, found the differentially expressed protein in AIS, which indicating that plasma exosomes, as a novel biomarker with high specificity, could be associated with the severity of AIS.

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Differential Proteomics Based on TMT and PRM Reveal the Resistance Response of Bambusa pervariabilis × Dendrocalamopisis grandis Induced by AP-Toxin

Metabolites ◽

10.3390/metabo9080166 ◽

2019 ◽

Vol 9 (8) ◽

pp. 166 ◽

Cited By ~ 2

Author(s):

Qianqian He ◽

Xinmei Fang ◽

Tianhui Zhu ◽

Shan Han ◽

Hanmingyue Zhu ◽

...

Keyword(s):

Mass Spectrometry ◽

Liquid Chromatography ◽

Metabolic Pathways ◽

Differentially Expressed ◽

Tandem Mass ◽

Differentially Expressed Proteins ◽

Liquid Chromatography Mass Spectrometry ◽

Chromatography Mass Spectrometry ◽

Tandem Mass Tag ◽

Mass Tag

Bambusa pervariabilis McClure × Dendrocalamopsis grandis (Q.H.Dai & X.l.Tao ex Keng f.) Ohrnb. blight is a widespread and dangerous forest fungus disease, and has been listed as a supplementary object of forest phytosanitary measures. In order to study the control of B. pervariabilis × D. grandis blight, this experiment was carried out. In this work, a toxin purified from the pathogen Arthrinium phaeospermum (Corda) Elli, which causes blight in B. pervariabilis × D. grandis, with homologous heterogeneity, was used as an inducer to increase resistance to B. pervariabilis × D. grandis. A functional analysis of the differentially expressed proteins after induction using a tandem mass tag labeling technique was combined with mass spectrometry and liquid chromatography mass spectrometry in order to effectively screen for the proteins related to the resistance of B. pervariabilis × D. grandis to blight. After peptide labeling, a total of 3320 unique peptides and 1791 quantitative proteins were obtained by liquid chromatography mass spectrometry analysis. Annotation and enrichment analysis of these peptides and proteins using the Gene ontology and Kyoto Encyclopedia of Genes and Genomes databases with bioinformatics software show that the differentially expressed protein functional annotation items are mainly concentrated on biological processes and cell components. Several pathways that are prominent in the Kyoto Encyclopedia of Genes and Genomes annotation and enrichment include metabolic pathways, the citrate cycle, and phenylpropanoid biosynthesis. In the Protein-protein interaction networks four differentially expressed proteins-sucrose synthase, adenosine triphosphate-citrate synthase beta chain protein 1, peroxidase, and phenylalanine ammonia-lyase significantly interact with multiple proteins and significantly enrich metabolic pathways. To verify the results of tandem mass tag, the candidate proteins were further verified by parallel reaction monitoring, and the results were consistent with the tandem mass tag data analysis results. It is confirmed that the data obtained by tandem mass tag technology are reliable. Therefore, the differentially expressed proteins and signaling pathways discovered here is the primary concern for subsequent disease resistance studies.

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Quantification of Globotriaosylsphingosine in Plasma and Urine of Fabry Patients by Stable Isotope Ultraperformance Liquid Chromatography–Tandem Mass Spectrometry

Clinical Chemistry ◽

10.1373/clinchem.2012.192138 ◽

2013 ◽

Vol 59 (3) ◽

pp. 547-556 ◽

Cited By ~ 63

Author(s):

Henrik Gold ◽

Mina Mirzaian ◽

Nick Dekker ◽

Maria Joao Ferraz ◽

Johan Lugtenburg ◽

...

Keyword(s):

Mass Spectrometry ◽

Liquid Chromatography ◽

Tandem Mass Spectrometry ◽

Fabry Disease ◽

Multiple Reaction Monitoring ◽

Internal Standard ◽

Response To Treatment ◽

Tandem Mass ◽

Healthy Controls ◽

Enzyme Replacement

BACKGROUND Biochemical markers that accurately reflect the severity and progression of disease in patients with Fabry disease and their response to treatment are urgently needed. Globotriaosylsphingosine, also called lysoglobotriaosylceramide (lysoGb3), is a promising candidate biomarker. METHODS We synthesized lysoGb3 and isotope-labeled [5,6,7,8,9] 13C5-lysoGb3 (internal standard). After addition of the internal standard to 25 μL plasma or 400 μL urine from patients with Fabry disease and healthy controls, samples were extracted with organic solvents and the lysoGb3 concentration was quantified by UPLC-ESI-MS/MS (ultraperformance liquid chromatography–electrospray ionization–tandem mass spectrometry). Calibration curves were constructed with control plasma and urine supplemented with lysoGb3. In addition to lysoGb3, lyso-ene-Gb3 was quantified. Quantification was achieved by multiple reaction monitoring of the transitions m/z 786.4 > 282.3 [M+H]+ for lysoGb3, m/z 791.4 > 287.3 [M+H]+ for [5,6,7,8,9] 13C5-lysoGb3, and 784.4 > 280.3 [M+H]+ for lyso-ene-Gb3. RESULTS The mean (SD) plasma lysoGb3 concentration from 10 classically affected Fabry hemizygotes was 94.4 (25.8) pmol/mL (range 52.7–136.8 pmol/mL), from 10 classically affected Fabry heterozygotes 9.6 (5.8) pmol/mL (range 4.1–23.5 pmol/mL), and from 20 healthy controls 0.4 (0.1) pmol/mL (range 0.3–0.5 pmol/mL). Lyso-ene-Gb3 concentrations were 10%–25% of total lysoGb3. The urine concentration of lysoGb3 was 40–480 times lower than in corresponding plasma samples. Lyso-ene-Gb3 concentrations in urine were comparable or even higher than the corresponding lysoGb3 concentrations. CONCLUSIONS This assay for the quantification of lysoGb3 and lyso-ene-Gb3 in human plasma and urine samples will be an important tool in the diagnosis of Fabry disease and for monitoring the effect of enzyme replacement therapy in patients with Fabry disease.

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Molecular Markers of MDR of Chemotherapy for HSCC: Proteomic Screening With High-Throughput Liquid Chromatography-Tandem Mass Spectrometry

Frontiers in Oncology ◽

10.3389/fonc.2021.687320 ◽

2021 ◽

Vol 11 ◽

Author(s):

Bin Shen ◽

Xuelin Dong ◽

Bo Yuan ◽

Zhijun Zhang

Keyword(s):

Mass Spectrometry ◽

Liquid Chromatography ◽

Molecular Markers ◽

Tandem Mass Spectrometry ◽

Signaling Pathway ◽

Tandem Mass ◽

Differentially Expressed Proteins ◽

Chromatography Tandem Mass Spectrometry ◽

Liquid Chromatography Tandem Mass ◽

Proteomic Screening

BackgroundHypopharyngeal squamous cell cancer (HSCC) is a head and neck tumor with a poor prognosis. Chemotherapy lacks effectiveness because of multidrug resistance (MDR), which has increased toxic side effects. Thus, there is an urgent need to identify the molecular markers of MDR of chemotherapy for HSCC.MethodsFifty clinical samples of HSCC were derived from patients including 12 sensitive or resistant to chemotherapy drugs. Proteomic screening was performed using liquid chromatography-tandem mass spectrometry (LC-MS), which was based on data-independent acquisition. Molecular markers of MDR of chemotherapy in patients with HSCC were identified with clinical data and validated with ELISA.ResultsA total of 673 differentially expressed proteins were identified in HSCC samples, where 172 were upregulated and 501 were downregulated. A total of 183 differentially expressed proteins including 102 upregulated and 81 downregulated proteins, were identified by comparing cancer sensitive to chemotherapy with cancer resistant to chemotherapy. Clinical HSCC samples had significantly higher expression of FADD and significantly lower expression of RIPK1. Expressions of FADD and RIPK1 proteins were significantly lower in the chemotherapy-sensitive group. These expression differences were not correlated with clinical data. RIPK1 and FADD are involved in necroptosis and the signaling pathway of PRRs. Using ELISA, the low expression of RIPK1 and FADD was found in the patients sensitive to chemotherapy.ConclusionLC-MS proteomics is an effective method to identify the molecular markers of HSCC. FADD and RIPK1 can act as molecular markers of MDR of chemotherapy in patients with HSCC and may function through necroptosis and the PRR signaling pathway.

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