Anemoside B4 inhibits SARS-CoV-2 replication in vitro and in vivo / 中草药·英文版

OBJECTIVE@#Anemoside B4 (AB4), the most abundant triterpenoidal saponin isolated from Pulsatilla chinensis, inhibited influenza virus FM1 or Klebsiella pneumoniae-induced pneumonia. However, the anti-SARS-CoV-2 effect of AB4 has not been unraveled. Therefore, this study aimed to determine the antiviral activity and potential mechanism of AB4 in inhibiting human coronavirus SARS-CoV-2 in vivo and in vitro.@*METHODS@#The cytotoxicity of AB4 was evaluated using the Cell Counting Kit-8 (CCK8) assay. SARS-CoV-2 infected HEK293T, HPAEpiC, and Vero E6 cells were used for in vitro assays. The antiviral effect of AB4 in vivo was evaluated by SARS-CoV-2-infected hACE2-IRES-luc transgenic mouse model. Furthermore, label-free quantitative proteomics and bioinformatic analysis were performed to explore the potential antiviral mechanism of action of AB4. Type I IFN signaling-associated proteins were assessed using Western blotting or immumohistochemical staining.@*RESULTS@#The data showed that AB4 reduced the propagation of SARS-CoV-2 along with the decreased Nucleocapsid protein (N), Spike protein (S), and 3C-like protease (3CLpro) in HEK293T cells. In vivo antiviral activity data revealed that AB4 inhibited viral replication and relieved pneumonia in a SARS-CoV-2 infected mouse model. We further disclosed that the antiviral activity of AB4 was associated with the enhanced interferon (IFN)-β response via the activation of retinoic acid-inducible gene I (RIG-1) like receptor (RLP) pathways. Additionally, label-free quantitative proteomic analyses discovered that 17 proteins were significantly altered by AB4 in the SARS-CoV-2 coronavirus infections cells. These proteins mainly clustered in RNA metabolism.@*CONCLUSION@#Our results indicated that AB4 inhibited SARS-CoV-2 replication through the RLR pathways and moderated the RNA metabolism, suggesting that it would be a potential lead compound for the development of anti-SARS-CoV-2 drugs.

Advances of peptide-centric data-independent acquisition analysis algorithms and software tools / 生物工程学报

Data-independent acquisition (DIA) is a high-throughput, unbiased mass spectrometry data acquisition method which has good quantitative reproducibility and is friendly to low-abundance proteins. It becomes the preferred choice for clinical proteomic studies especially for large cohort studies in recent years. The mass-spectrometry (MS)/MS spectra generated by DIA is usually heavily mixed with fragment ion information of multiple peptides, which makes the protein identification and quantification more difficult. Currently, DIA data analysis methods fall into two main categories, namely peptide-centric and spectrum-centric. The peptide-centric strategy is more sensitive for identification and more accurate for quantification. Thus, it has become the mainstream strategy for DIA data analysis, which includes four key steps: building a spectral library, extracting ion chromatogram, feature scoring and statistical quality control. This work reviews the peptide-centric DIA data analysis procedure, introduces the corresponding algorithms and software tools, and summarizes the improvements for the existing algorithms. Finally, the future development directions are discussed.

Application of quantitative proteomics in the study of acute mountain sickness / 生物工程学报

Acute mountain sickness (AMS) is a clinical syndrome of multi-system physiological disorder after acute exposure to low pressure and low oxygen at high altitude. Quantitative proteomics can systematically quantify and describe protein composition and dynamic changes. In recent years, quantitative proteomics has been widely used in the prevention, diagnosis, treatment and pathogenesis of many diseases. This review summarizes the progress of quantitative proteomics techniques and its application in the prevention, diagnosis, treatment of AMS and mechanisms of rapidly acclimatizing to plateau, in order to provide a reference for the pathogenesis, early intervention, clinical treatment and proteomic research of AMS.

Quantitative proteomics revealed extensive microenvironmental changes after stem cell transplantation in ischemic stroke / 医学前沿

The local microenvironment is essential to stem cell-based therapy for ischemic stroke, and spatiotemporal changes of the microenvironment in the pathological process provide vital clues for understanding the therapeutic mechanisms. However, relevant studies on microenvironmental changes were mainly confined in the acute phase of stroke, and long-term changes remain unclear. This study aimed to investigate the microenvironmental changes in the subacute and chronic phases of ischemic stroke after stem cell transplantation. Herein, induced pluripotent stem cells (iPSCs) and neural stem cells (NSCs) were transplanted into the ischemic brain established by middle cerebral artery occlusion surgery. Positron emission tomography imaging and neurological tests were applied to evaluate the metabolic and neurofunctional alterations of rats transplanted with stem cells. Quantitative proteomics was employed to investigate the protein expression profiles in iPSCs-transplanted brain in the subacute and chronic phases of stroke. Compared with NSCs-transplanted rats, significantly increased glucose metabolism and neurofunctional scores were observed in iPSCs-transplanted rats. Subsequent proteomic data of iPSCs-transplanted rats identified a total of 39 differentially expressed proteins in the subacute and chronic phases, which are involved in various ischemic stroke-related biological processes, including neuronal survival, axonal remodeling, antioxidative stress, and mitochondrial function restoration. Taken together, our study indicated that iPSCs have a positive therapeutic effect in ischemic stroke and emphasized the wide-ranging microenvironmental changes in the subacute and chronic phases.

Proteomics: a tool to investigate of composition and function of HDL in hyperlipidemia / Proteômica: uma ferramenta para a investigação da composição e função da HDL em hiperlipidemia

The inverse relationship between HDL-C (high-density lipoprotein cholesterol) and cardiovascular disease is well established. However, it is consensus that the cholesterol content present in HDL does not capture its complexity, and other metrics need to be explored. HDL is a heterogeneous, protein-enriched particle with functions going beyond lipid metabolism. In this way, its protein content seems to be attractive to investigate its behavior in the face of pathologies. Many of the proteins with important function in HDL are in low abundance (<1% of total proteins), which makes their detection challenging. Quantitative proteomics allows detecting proteins with high precision and robustness in complex matrix. However, quantitative proteomics is still poorly explored in the context of HDL. In this sense, in the second chapter of this thesis, the analytical performance of two quantitative methodologies was carefully investigated. These methods achieved adequate linearity and high precision using labeled peptides in a pool HDL, in addition to comparable ability to differentiate proteins from HDL subclasses of healthy subjects. Another bottleneck that waits for a solution in proteomics is the lack of standardization in data processing and analysis after mass spectrometry acquisition. In addition, interest in the cardioprotective properties of omega-3 is growing, but little is known about its effects on the HDL proteome. Thus, in the third chapter of this thesis, we compared five protein quantification strategies using Skyline and MaxDIA software platforms in order to investigate the HDL proteome from mice submitted to a high-fat diet supplemented or not with omega-3. MaxDIA with label-free quantification (MaxLFQ) achieved high precision to show that polyunsaturated fatty acids remodel the HDL proteome to a less inflammatory profile. Therefore, the two studies presented in this thesis begin to open new paths for a deeper and more reliable understanding of HDL, both at the level of protein quantification by mass spectrometry and after data acquisition

A inversa relação entre HDL-C (do inglês, high-density lipoprotein cholesterol) e doenças cardiovasculares é bem estabelecida. No entanto, é consenso que o conteúdo de colesterol presente na HDL não captura sua complexidade, e outras métricas precisam ser exploradas. A HDL é uma partícula heterogênea, enriquecida em proteínas, com funções que vão além do metabolismo de lipídeos. Dessa forma, seu conteúdo proteico parece ser mais atrativo para exprimir seu comportamento frente às patologias. Muitas das proteínas com função importante estão em baixa abundância (<1% do total de proteínas), o que torna a detecção desafiadora. Métodos quantitativos de proteômica permitem detectar proteínas com alta precisão e robustez em matrizes complexas. No entanto, a proteômica quantitativa ainda é pouco explorada no contexto da HDL. Nesse sentido, no segundo capítulo dessa tese, a performance analítica de dois métodos quantitativos foi criteriosamente investigada, os quais alcançaram adequada linearidade e alta precisão usando peptídeos marcados em um pool de HDL, além de comparável habilidade em diferenciar as proteínas das subclasses da HDL de indivíduos saudáveis. Outro gargalo que aguarda por solução em proteômica é a falta de padronização no processamento e análise de dados após a aquisição por espectrometria de massas. Além disso, é crescente o interesse das propriedades cardioprotetivas do ômega-3, porém pouco se conhece sobre seus efeitos no proteoma da HDL. Então, no terceiro capítulo dessa tese, comparamos cinco estratégias de quantificação de proteínas utilizando os softwares Skyline e MaxDIA com o intuito de comparar o proteoma da HDL de camundongos submetidos a uma dieta hiperlipídica suplementados ou não com ômega-3. MaxDIA com quantificação label-free (MaxLFQ) apresentou alta precisão para mostrar que o ômega-3 remodela o proteoma da HDL para um perfil menos inflamatório. Portanto, os dois estudos apresentados nessa tesa começam a abrir novos caminhos para o entendimento mais profundo e confiável da HDL tanto por meio da quantificação das proteínas por espectrometria de massas quanto após à aquisição dos dados

Label-free quantitative proteomic study of RUNX3 regulating Herceptin resistance in gastric cancer cells / 药学学报

Resistance of tumor cells is a complex biological process involving multiple mechanisms and factors, in which anti-apoptosis is the most important cause of drug resistance. Previous studies have shown that the DNA binding activity of Runt related transcription factor 3 (RUNX3) increased prominently in Herceptin resistant gastric cancer cells (NCI N87R) while the relevance of which to drug resistance has not yet been confirmed. In this study, we employed CRISPR/Cas9 to establish RUNX3 knock-out cell line (△RUNX3/NCI N87R) to investigate the functions of RUNX3 in Herceptin resistance of NCI N87R cells and its potential mechanisms. We investigated proteomics profiling of △RUNX3/NCI N87R cells based on label free quantitative proteomics. Differentially expressed proteins were screened out according to fold change and significance level between △RUNX3/NCI N87R and NCI N87R cells. Pathway enrichment analysis was done using GeneAnalytics database, and gene ontology analysis was conducted by DAVID Bioinformatics Resources database. Protein-protein interaction networks were constructed based on STRING database. The results showed that △RUNX3/NCI N87R cells increased the sensitivity to Herceptin. Proteomic data demonstrated that the expression of 577 genes changed significantly in △RUNX3/NCI N87R cells, among which 191 genes were up-regulated while 386 ones down-regulated comparing with NCI N87R cells. Pathway analysis showed that autophagy, cell cycle, apoptosis, mitochondrial fatty acid β oxidation, neurogenic locus notch homolog protein 1 (NOTCH1), mammalian target of rapamycin (mTOR), Hedgehog and DNA damage response pathways exhibited notable changes based on pathway enrichment ratio and significance level (P < 0.05). These results indicated that RUNX3 knock-out altered multiple signaling pathways of NCI N87R cells. Western blotting manifested that the expression of autophagy regulatory molecules autophagy-related protein (ATG) 13, 7 and BECN1 increased remarkably while cell cycle molecules serine/threonine-protein kinase Chk2 (CHEK2) and apoptosis regulator Bcl-2 (BCL2) decreased prominently in △RUNX3/NCI N87R cells. The p-AKT expression decreased significantly in △RUNX3/NCI N87R cells compared with NCI N87R cells (P < 0.01) and was suppressed by Herceptin. These results indicated that RUNX3 knock-out altered cell cycle, increased inhibition to p-AKT by Herceptin, promoted autophagy and induced cell apoptosis of NCI N87R cells. These results suggested that RUNX3 may be a potential therapeutic target for reversing or reducing Herceptin resistance in gastric cancer cells.

Quantitative Proteomics Using Isobaric Labeling: A Practical Guide / 基因组蛋白质组与生物信息学报·英文版

In the past decade, relative proteomic quantification using isobaric labeling technology has developed into a key tool for comparing the expression of proteins in biological samples. Although its multiplexing capacity and flexibility make this a valuable technology for addressing various biological questions, its quantitative accuracy and precision still pose significant challenges to the reliability of its quantification results. Here, we give a detailed overview of the different kinds of isobaric mass tags and the advantages and disadvantages of the isobaric labeling method. We also discuss which precautions should be taken at each step of the isobaric labeling workflow, to obtain reliable quantification results in large-scale quantitative proteomics experiments. In the last section, we discuss the broad applications of the isobaric labeling technology in biological and clinical studies, with an emphasis on thermal proteome profiling and proteogenomics.

Commonly used quantitative proteomics research techniques and their application in the study of pathogenesis of liver-derived diseases / 中国临床药理学与治疗学

Proteomics is a subject with proteome as its research object. It is an important omics technology in the post-genomic era. Protein is the final product of gene expression. The overall study of protein expression is of great significance to the study of disease mechanisms and drug efficacy. Among them, quantitative proteomics has been widely used in recent years. The liver is the most important drug metabolism organ in the body, and more than 70% of drugs are metabolized by the liver. This article introduces the commonly used proteomics technology so far, as well as the progress of quantitative proteomics in drug-induced liver injury, non-alcoholic fatty liver disease, liver fibrosis and cirrhosis, hepatocellular carcinoma in the past five years.

Method and application of proteomics in study of targets of traditional Chinese medicines / 中国中药杂志

The study on the targets of traditional Chinese medicine is an important part of researchers using modern scientific language to clarify the mechanism of traditional Chinese medicine. However, the research on the targets of Chinese medicine is full of challenges due to the complexity of active ingredients. As a branch of systems biology, proteomics focus on specific proteins in living organisms from a holistic perspective, which significantly improves the efficiency of targets discovery and has obvious advantages in the research of targets of Chinese medicine. Based on relevant literature and different methods used in targets of Chinese medicine, proteomics can be divided into chemical proteomics, differential proteomics and quantitative proteomics. The applications of the above three methods are illustrated in this paper as well, which will provide new methods and ideas for the study of the mechanism of Chinese medicine in the future.

Quantitative Proteomic Analysis Reveals Impaired Axonal Guidance Signaling in Human Postmortem Brain Tissues of Chronic Traumatic Encephalopathy

Chronic traumatic encephalopathy (CTE) is a distinct neurodegenerative disease that associated with repetitive head trauma. CTE is neuropathologically defined by the perivascular accumulation of abnormally phosphorylated tau protein in the depths of the sulci in the cerebral cortices. In advanced CTE, hyperphosphorylated tau protein deposits are found in widespread regions of brain, however the mechanisms of the progressive neurodegeneration in CTE are not fully understood. In order to identify which proteomic signatures are associated with CTE, we prepared RIPA-soluble fractions and performed quantitative proteomic analysis of postmortem brain tissue from individuals neuropathologically diagnosed with CTE. We found that axonal guidance signaling pathwayrelated proteins were most significantly decreased in CTE. Immunohistochemistry and Western blot analysis showed that axonal signaling pathway-related proteins were down regulated in neurons and oligodendrocytes and neuron-specific cytoskeletal proteins such as TUBB3 and CFL1 were reduced in the neuropils and cell body in CTE. Moreover, oligodendrocyte-specific proteins such as MAG and TUBB4 were decreased in the neuropils in both gray matter and white matter in CTE, which correlated with the degree of axonal injury and degeneration. Our findings indicate that deregulation of axonal guidance proteins in neurons and oligodendrocytes is associated with the neuropathology in CTE. Together, altered axonal guidance proteins may be potential pathological markers for CTE.

Quantitative proteomics and differential signal enrichment in nasopharyngeal carcinoma cells with or without gene knockout / 南方医科大学学报

OBJECTIVE@#To analyze the effects of alterations in the expressions of methyltransferase on protein expression profiles in human nasopharyngeal carcinoma (NPC) cells and enrich the differential signaling pathways.@*METHODS@#The total protein was extracted from -knockout cell line CNE1 and the wild-type cell line CNE1, and the differentially expressed proteins were screened by tandem mass tag (TMT) labeled protein quantification technique and tandem mass spectrometry. GO analysis was used to annotate and enrich the differentially expressed proteins, and the KEGG database was used to enrich and analyze the pathways of the differential proteins.@*RESULTS@#With a fold change (FC)≥1.2 and < 0.05 as the screening standard, 2049 differentially expressed proteins were identified in CNE1 cells, among which 904 were up-regulated and 1145 were down-regulated. GO functional annotation results indicated that knockout caused characteristic changes in multiple biological processes (cell processes and regulation, cell movement, metabolic processes, and biosynthesis of cellular components), molecular functions (catalytic activity and molecular binding, transcription factor activity), and cellular components (cell membrane, organelle, macromolecular complex). KEGG analysis showed that the differentially expressed proteins were involved in an array of signaling pathways closely related to tumors, including MAPK, PI3K-Akt, Ras, Rap1, mTOR, Hippo, HIF-1, Wnt, AMPK, FoxO, ErbB, P53 and JAK-STAT.@*CONCLUSIONS@# knockout significantly changes the protein expression characteristics of NPC cells and affects a number of signal pathways closely related to tumors. The results provide evidence for investigation of the pathogenesis and therapeutic target screening of NPC.

Proteomic-based studies on hypolipidemic mechanism of total phenylpropanoid glycosides from ligustrum robustum (roxb. ) blume in hyperlipidemic hamsters / 中国药理学通报

Aim To explore the hypolipidemic mechanism of the total phenylpropanoid glycoside from Ligustrum robustum (Roxb. ) Blume (LRTPG) on hyperlipidemic hamsters using label-free quantitative proteomic technique. Methods The total protein was extracted from livers of model group and the group treated with LRTPG for label-free quantitative proteomics research. Results The proteomic data showed that a total of 2231 proteins were identified. And 549 proteins were found to be differentially expressed between model group and group treated with LRTPG. Among the 549 proteins, 93 proteins were up-regulated and 59 proteins were down-regulated, and 397 proteins had quantitative values only in model group or drug-administered group. Further, gene ontology (GO) analysis indicated that those differentially expressed proteins were primarily involved in an array of biological processes including metabolism, transport, oxidation-reduction, phosphorylation, signal transduction and lipid metabolism. KEGG pathway analysis revealed that these proteins were involved in several signal pathways including oxidative phosphorylation, non-alcoholic fatty liver dis-ease, PI3K-Akt, cAMP, and cGMP-PKG pathway. And some of these proteins were much related to the lipid metabolism, such as CD36, PK, HSS, GCK, ApoA I, Acly and FABP5. Conclusion The hypolipidemic effect of LRTPG may be related to CD36, PK, HSS, GCK, ApoA I, Acly and FABP5.

Study on the mechanisms underlying overwintering of Culex pipiens pallens using iTRAQ-based quantitative proteomic analysis / 中国血吸虫病防治杂志

Objective To compare the difference of protein expression between the post-overwintering stage and the diapauses preparation stage in Culex pipiens pallens, so as to reveal the mechanisms underlying the overwintering diapause of Cx. pipienspallens. Methods A quantitative proteomic analysis was performed in Cx. pipiens pallens before and after overwintering diapause by using isobaric tags for relative and absolute quantification (iTRAQ) labeling. Results A total of 244 differentially expressed proteins were identified in Cx. pipiens pallens before and after overwintering diapause, including 126 up-regulated proteins and 118 down-regulated proteins. iTRAQ-based quantitative proteomic analysis revealed that these differentially expressed proteins were linked to function and energy production and conversion, lipid metabolism, remodeling of cytoskeleton, carbohydrate metabolism, protein transport, molecular chaperones, stress tolerance and metabolic enzymes. Conclusions This is the first study to identify the overwintering diapause-related proteins in Cx. pipiens pallens using proteomics tools, which reveals KEGG pathways and GO terms associated with the overwintering diapauses of Cx. pipiens pallens. Our findings provide additional understandings pertaining to the mechanisms underlying the overwintering diapauses of Cx. pipiens pallens.

Study on the mechanisms underlying overwintering of Culex pipiens pallens using iTRAQ-based quantitative proteomic analysis / 中国血吸虫病防治杂志

Objective To compare the difference of protein expression between the post-overwintering stage and the diapauses preparation stage in Culex pipiens pallens, so as to reveal the mechanisms underlying the overwintering diapause of Cx. pipienspallens. Methods A quantitative proteomic analysis was performed in Cx. pipiens pallens before and after overwintering diapause by using isobaric tags for relative and absolute quantification (iTRAQ) labeling. Results A total of 244 differentially expressed proteins were identified in Cx. pipiens pallens before and after overwintering diapause, including 126 up-regulated proteins and 118 down-regulated proteins. iTRAQ-based quantitative proteomic analysis revealed that these differentially expressed proteins were linked to function and energy production and conversion, lipid metabolism, remodeling of cytoskeleton, carbohydrate metabolism, protein transport, molecular chaperones, stress tolerance and metabolic enzymes. Conclusions This is the first study to identify the overwintering diapause-related proteins in Cx. pipiens pallens using proteomics tools, which reveals KEGG pathways and GO terms associated with the overwintering diapauses of Cx. pipiens pallens. Our findings provide additional understandings pertaining to the mechanisms underlying the overwintering diapauses of Cx. pipiens pallens.

Quantitative proteomics of inhibitory mechanism of Dendrobium denneanum ether extract on lung cancer cells / 中国中药杂志

Dendrobium denneanum have been used for a long time as rare medicinal herbs in traditional Chinese medicine. Our previous works found that ether extract of D. denneanum had higher anticancer activities than alcohol or water extract,thus with better development prospects. Quantitative proteomics based on SILAC technique was used to investigate the anticancer mechanism of D. denneanum on lung tumor cell line A549,and 4 855 proteins were detected in A549 cells. Quantitative proteomics experiments found that 193 proteins of A549 cells were up-regulated,and 44 proteins were down-regulated by ether extract of D. denneanum. Those proteins are associated with synthesis,transport and metabolism of biological macromolecules,chaperone,DNA repair,oxidoreductase,cell adhesion,cell cycle,apoptosis and autophagy. Through the function analysis of differentially expressed proteins,it was inferred that ether extract of D. denneanum caused cell protein metabolism disorder,endoplasmic reticulum stress response,abnormal self-repair mechanism of cells,damage of cell adhesion and proliferation; besides,it caused a dramatic increase in ROS level in A549 cells,and upset the balance of intracellular oxidation reduction system. Affected by the above factors,lung cancer cells initiated apoptosis and autophagy,which accelerated cell death. This research explains the anticancer mechanism of D. denneanum from the perspective of quantitative proteomics,and lays a foundation for future research and development of new anticancer drugs based on ether extract of D. denneanum.

Label-free quantitative proteomics reveals fibrinopeptide B and heparin cofactor II as potential serum biomarkers in respiratory syncytial virus-infected mice treated with Qingfei oral liquid formula / 中国天然药物

Respiratory syncytial virus (RSV) is a leading cause of acute lower respiratory tract infections. Qingfei oral liquid (QFOL), a traditional Chinese medicine, is widely used in clinical treatment for RSV-induced pneumonia. The present study was designed to reveal the potential targets and mechanism of action for QFOL by exploring its influence on the host cellular network following RSV infection. We investigated the serum proteomic changes and potential biomarkers in an RSV-infected mouse pneumonia model treated with QFOL. Eighteen BALB/c mice were randomly divided into three groups: RSV pneumonia model group (M), QFOL-treated group (Q) and the control group (C). Serum proteomes were analyzed and compared using a label-free quantitative LC-MS/MS approach. A total of 172 protein groups, 1009 proteins, and 1073 unique peptides were successfully identified. 51 differentially expressed proteins (DEPs) were identified (15 DEPs when M/C and 43 DEPs when Q/M; 7 DEPs in common). Classification and interaction network showed that these proteins participated in various biological processes including immune response, blood coagulation, complement activation, and so forth. Particularly, fibrinopeptide B (FpB) and heparin cofactor II (HCII) were evaluated as important nodes in the interaction network, which was closely involved in coagulation and inflammation. Further, the FpB level was increased in Group M but decreased in Group Q, while the HCII level exhibited the opposite trend. These findings not only indicated FpB and HCII as potential biomarkers and targets of QFOL in the treatment of RSV pneumonia, but also suggested a regulatory role of QFOL in the RSV-induced disturbance of coagulation and inflammation-coagulation interactions.

Label-free quantitative proteomics reveals fibrinopeptide B and heparin cofactor II as potential serum biomarkers in respiratory syncytial virus-infected mice treated with Qingfei oral liquid formula / 中国天然药物

Respiratory syncytial virus (RSV) is a leading cause of acute lower respiratory tract infections. Qingfei oral liquid (QFOL), a traditional Chinese medicine, is widely used in clinical treatment for RSV-induced pneumonia. The present study was designed to reveal the potential targets and mechanism of action for QFOL by exploring its influence on the host cellular network following RSV infection. We investigated the serum proteomic changes and potential biomarkers in an RSV-infected mouse pneumonia model treated with QFOL. Eighteen BALB/c mice were randomly divided into three groups: RSV pneumonia model group (M), QFOL-treated group (Q) and the control group (C). Serum proteomes were analyzed and compared using a label-free quantitative LC-MS/MS approach. A total of 172 protein groups, 1009 proteins, and 1073 unique peptides were successfully identified. 51 differentially expressed proteins (DEPs) were identified (15 DEPs when M/C and 43 DEPs when Q/M; 7 DEPs in common). Classification and interaction network showed that these proteins participated in various biological processes including immune response, blood coagulation, complement activation, and so forth. Particularly, fibrinopeptide B (FpB) and heparin cofactor II (HCII) were evaluated as important nodes in the interaction network, which was closely involved in coagulation and inflammation. Further, the FpB level was increased in Group M but decreased in Group Q, while the HCII level exhibited the opposite trend. These findings not only indicated FpB and HCII as potential biomarkers and targets of QFOL in the treatment of RSV pneumonia, but also suggested a regulatory role of QFOL in the RSV-induced disturbance of coagulation and inflammation-coagulation interactions.

Xiao-Xu-Ming decoction protects against chronic cerebral ischemia injury in rats and profile of differentially expression proteins analysis in hippocampus / 中国药理学与毒理学杂志

OBJECTIVE Vascular dementia (VD) refers to a progressive decline in memory and cognitive function caused by chronic cerebral ischemia. 2-Vessels occlusion (2-VO) has been widely used as a model of VD. Xiao-Xu-Ming decoction, a well-known traditional Chinese medicine prescrip-tion,has been widely used to treat stroke and sequelae of stroke.The present study was to investigate the mechanism of Xiao-Xu-Ming decoction(XXM) against chronic cerebral ischemia injury in rats. METHODS After XXM treatment, rats were performed a memory testing with Morris water maze and motor ability testing using prehensile test and inclined screen test.Neuronal plasticity was observed by immunofluorescent staining with MAP2 antibody. Differentially expressed proteins of rat hippocampus were analyzed by Label-free quantitative proteomics. RESULTS XXM significantly alleviated 2-VO-induced learning and memory deficits, motor ability dysfunction, and neuronal plasticity injury in rats. The mechanism might be involved in up-regulation of 39 proteins and down-regulation of 13 proteins in the hippocampus of rats after XXM treatment vs 2-VO group rats.Gene ontology and pathway analysis showed that the regulated proteins are mainly involved in oxidation reduction process, intracellular signaling cascade process, and protein catabolic process, etc. The signal pathways are mainly involved in ubiquitin mediated proteolysis and phosphatidylinositol signaling system. CONCLUSION Current findings provide new insights into the molecular mechanisms of XXM on chronic cerebral ischemia.

Comparative untargeted proteomic analysis of ADME proteins and tumor antigens for tumor cell lines

In the present study, total membrane proteins from tumor cell lines including HepG2, Hep3B2, H226, Ovcar3 and N87 were extracted and digested with LysC and trypsin. The resulting peptide lysate were pre-fractionated and subjected to untargeted quantitative proteomics analysis using a high resolution mass spectrometer. The mass spectra were processed by the MaxQuant and the protein abundances were estimated using total peak area (TPA) method. A total of 6037 proteins were identified, and the analysis resulted in the identification of 2647 membrane proteins. Of those, tumor antigens and absorption, metabolism, disposition and elimination (ADME) proteins including UDP-glucuronosyltransferase, cytochrome P450, solute carriers and ATP-binding cassette transporters were detected and disclosed significant variations among the cell lines. The principal component analysis was performed for the cluster of cell lines. The results demonstrated that H226 is closely related with N87, while Hep3B2 aligned with HepG2. The protein cluster of Ovcar3 was apart from that of other cell lines investigated. By providing for the first time quantitative untargeted proteomics analysis, the results delineated the expression profiles of membrane proteins. These findings provided a useful resource for selecting targets of choice for anticancer therapy through advancing data obtained from preclinical tumor cell line models to clinical outcomes.

Differentially quantitative proteome of MHCC97H after Rab27B knock-down:a preliminary study / 军事医学

Objective To discover the vital role of Rab27B in tumor cells and its potential molecular mechanism by means of quantitative proteomics analysis of Rab27B knockdown in MHCC97H.Methods The expression of Rab27B in MHCC97H cells was knocked down by the combination of Tet-on advanced inducible expression system and RNA interference technology.Then, proteins extracted from the cells were identified by LC-MS/MS system after FASP digestion and iTRAQ 4-plex labeling. Finally, the properties of differentially expressed proteins, including the subcellular localizations, biological processes and molecular functions, were analyzed by the bioinformatics method.Results There were 448 differentially expressed proteins (|Ratio|>1.21, P<0.05) identified in MHCC97H cells after Rab27B knock-down.The expression levels of 229 or 219 proteins were positively or negatively correlated with Rab27B, respectively. These differentially expressed proteins were mainly involved in vesicle transport, macromolecule localization, cellular response to stimulus.Furthermore, there were 26 differentially expressed proteins participating in 8 tumor-related signal pathways, eleven of which were in the focal adhesion signal pathway.Conclusion The analysis of quantitative proteomics in Rab27B-knockdown MHCC97H cell line by iTRAQ suggests that Rab27B not only has an impact on the exosomal secretion of tumor cells, but also regulates master proteins in signal pathways involved in cell proliferation and migration.