BACKGROUND: Long-term remote ischemic conditioning (RIC) has been proven to be beneficial in multiple diseases, such as cerebral and cardiovascular diseases. However, the hyperacute and acute effects of a single RIC stimulus are still not clear. Quantitative proteomic analyses of plasma proteins following RIC application have been conducted in preclinical and clinical studies but exhibit high heterogeneity in results due to wide variations in experimental setups and sampling procedures. Hence, this study aimed to explore the immediate effects of RIC on plasma proteome in healthy young adults to exclude confounding factors of disease entity, such as medications and gender. METHODS: Young healthy male participants were enrolled after a systematic physical examination and 6-month lifestyle observation. Individual RIC sessions included five cycles of alternative ischemia and reperfusion, each lasting for 5 min in bilateral forearms. Blood samples were collected at baseline, 5 min after RIC, and 2 h after RIC, and then samples were processed for proteomic analysis using liquid chromatography-tandem mass spectrometry method. RESULTS: Proteins related to lipid metabolism (e.g., Apolipoprotein F), coagulation factors (hepatocyte growth factor activator preproprotein), members of complement cascades (mannan-binding lectin serine protease 1 isoform 2 precursor), and inflammatory responses (carboxypeptidase N catalytic chain precursor) were differentially altered at their serum levels following the RIC intervention. The most enriched pathways were protein glycosylation and complement/coagulation cascades. CONCLUSIONS One-time RIC stimulus may induce instant cellular responses like anti-inflammation, coagulation, and fibrinolysis balancing, and lipid metabolism regulation which are protective in different perspectives. Protective effects of single RIC in hyperacute and acute phases may be exploited in clinical emergency settings due to apparently beneficial alterations in plasma proteome profile. Furthermore, the beneficial effects of long-term (repeated) RIC interventions in preventing chronic cardiovascular diseases among general populations can also be expected based on our study findings.
Young Adult ; Humans ; Male ; Proteome ; Cardiovascular Diseases ; Proteomics ; Ischemia ; Blood Coagulation

OBJECTIVE: To analyze the differentially phosphorylated proteins in DENV-2-infected human umbilical venous endothelial cells (HUVECs) and explore the possible pathogenic mechanism of DENV-2 infection. METHODS: The total proteins were extracted from DENV-2-infected HUVECs and blank control HUVEC using SDT lysis method. The phosphorylated proteins were qualitatively and quantitatively analyzed using tandem mass spectrometry (TMT). The identified differentially phosphorylated proteins were analyzed by bioinformatics analyses such as subcellular localization analysis, GO enrichment analysis, KEGG pathway analysis and protein-protein interaction (PPI) analysis. Western blotting was used to detect the expressions of phosphorylated Jun, map2k2 and AKT1 proteins in DENV-2-infected HUVECs. RESULTS: A total of 2918 modified peptides on 1385 different proteins were detected, and among them 1346 were significantly upregulated (FC > 1.2, P < 0.05) and 1572 were significantly downregulated (FC < 0.83, P < 0.05). A total of 49 phosphorylated conserved motifs were obtained by amino acid conservative motif analysis. The most abundant differentially phosphorylated peptides in protein domain analysis included RNA recognition motif, protein kinase domain and PH domain. Subcellular localization analysis showed that the differentially modified peptides were mainly localized in the nucleus and cytoplasm. GO enrichment and KEGG pathway analysis showed that the differential peptides were mainly enriched in the regulation of stimulation response, biosynthesis of small molecules containing nuclear bases, and migration of phagosomes and leukocytes across the endothelium. PPI and KEGG joint analysis showed that the up-regulated and down-regulated differentially phosphorylated proteins were enriched in 15 pathways. In DENV-2-infected HUVECs, Western blotting detected differential expressions of phosphorylated proteins related with the autophagy pathway, namely JUN, MAP2K2 and AKT1, and among them p-JUN was significantly down-regulated and p-AKT1 and p-MAP2K2 were significantly upregulated (P < 0.01). CONCLUSION DENV-2 infected HUVECs show numerous differentially expressed proteins. The downregulation of p-JUN and upregulation of p-MAP2K2 and p-AKT1 suggest their potential roles in regulating autophagy, which is probably involved in the mechanism of DENV-2 infection.
Humans ; Autophagy ; Cell Death ; Cell Nucleus ; Human Umbilical Vein Endothelial Cells/virology* ; Dengue ; Proteome

The prevalence of obesity has increased worldwide in recent decades. Genetic factors are now known to play a substantial role in the predisposition to obesity and may contribute up to 70% of the risk for obesity. Technological advancements during the last decades have allowed the identification of many hundreds of genetic markers associated with obesity. However, the transformation of current genetic variant-obesity associations into biological knowledge has been proven challenging. Genomics and proteomics are complementary fields, as proteomics extends functional analyses. Integrating genomic and proteomic data can help to bridge a gap in knowledge regarding genetic variant-obesity associations and to identify new drug targets for the treatment of obesity. We provide an overview of the published papers on the integrated analysis of proteomic and genomic data in obesity and summarize four mainstream strategies: overlap, colocalization, Mendelian randomization, and proteome-wide association studies. The integrated analyses identified many obesity-associated proteins, such as leptin, follistatin, and adenylate cyclase 3. Despite great progress, integrative studies focusing on obesity are still limited. There is an increased demand for large prospective cohort studies to identify and validate findings, and further apply these findings to the prevention, intervention, and treatment of obesity. In addition, we also discuss several other potential integration methods.
Humans ; Proteome/metabolism* ; Proteomics ; Prospective Studies ; Obesity/genetics* ; Genomics ; Genome-Wide Association Study

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.
Humans ; Proteomics/methods* ; Reproducibility of Results ; Peptides/chemistry* ; Software ; Algorithms ; Tandem Mass Spectrometry/methods* ; Proteome/analysis*

Seed quality plays an important role in the agricultural and animal husbandry production, the effective utilization of genetic resources, the conservation of biodiversity and the restoration and reconstruction of plant communities. Seed aging is a common physiological phenomenon during storage. It is a natural irreversible process that occurs and develops along with the extension of seed storage time. It is not only related to the growth, yield and quality of seed and seedling establishment, but also has an important effect on the conservation, utilization and development of plant germplasm resources. The physiological mechanisms of seed aging are complex and diverse. Most studies focus on conventional physiological characterization, while systematic and comprehensive in-depth studies are lacking. Here we review the recent advances in understanding the physiology of seed aging process, including the methods of seed aging, the effect of aging on seed germination, and the physiological and molecular mechanisms of seed aging. The change of multiple physiological parameters, including seed vigor, electrical conductivity, malondialdehyde content and storage material in the seed, antioxidant enzyme activity and mitochondrial structure, were summarized. Moreover, insights into the mechanism of seed aging from the aspects of transcriptome, proteome and aging related gene function were summarized. This study may facilitate the research of seed biology and the conservation and utilization of germplasm resources.
Germination ; Plants ; Proteome ; Seedlings ; Seeds/genetics*

The effects of Jingui Shenqi Pills(Jingui) and Liuwei Dihuang Pills(Liuwei) which respectively tonify kidney Yang and kidney Yin on brain function have attracted great attention, while the differences of protein expression regulated by Jingui and Liuwei remain to be studied. This study explored the difference of protein expression profiles in the hippocampi of mice orally administrated with the two drugs for 7 days. The protein expression was quantified using LC-MS/MS. The results showed that among the 5 860 proteins tested, 151, 282 and 75 proteins responded to Jingui alone, Liuwei alone, and both drugs, respectively. The ratio of up-regulated proteins to down-regulated proteins was 1.627 in Jingui group while only 0.56 in Liuwei group. The proteins up-regulated by Jingui were mainly involved in membrane transport, synaptic vesicle cycle, serotonergic synapse, dopaminergic synapse and so on, suggesting that Jingui may play a role in promoting the transport of neurotransmitter in the nervous system. The proteins down-regulated by Liuwei were mainly involved in membrane transport, synapse, ion transport(potassium and sodium transport), neurotransmitter transport, innate and acquired immune responses, complement activation, inflammatory response, etc. In particular, Liuwei showed obvious down-regulation effect on the members of solute carrier(SLC) superfamily, which suggested that Liuwei had potential inhibitory effect on membrane excitation and transport. Finally, consistent results were obtained in the normal mouse and the mouse model with corticosterone-induced depressive-like behavior. This study provides an experimental basis for understanding the effect of Jingui and Liuwei on brain function from protein network.
Animals ; Chromatography, Liquid ; Drugs, Chinese Herbal/pharmacology* ; Hippocampus/metabolism* ; Mice ; Proteome/metabolism* ; Proteomics ; Tandem Mass Spectrometry

Thermal proteome profiling (TPP) is a combination of cellular thermal shift assay (CETSA) and quantitative mass spectrometry (MS), also termed as MS-CETSA. TPP determines the stability of the entire proteome by measuring the content of soluble proteins in cells or cell lysates at different heating temperatures. Proteins can change their thermostability when interacting with small molecules (e.g., drugs or metabolites), nucleic acids, or other proteins or posttranslational modification, while TPP can identify target proteins based on the difference in thermostability with or without ligand-binding. At present, TPP has been applied to identify the targets and off-targets of drugs and interrogate protein-metabolite and protein-protein interactions. Due to limited understanding of this technology, this review introduced the principles, methods, applications, advantages and limitations of TPP.
Proteome ; Mass Spectrometry

The systematic and in-depth study of phosphoproteome rely on highly reproducible and specific phosphopeptide enrichment methods. At present, a variety of enrichment methods have been developed based on different principles, and these methods often display different selectivity and specificity. It is therefore very important to select the most suitable enrichment method according to different research purposes. This review summarized the phosphopeptide enrichment based on affinity chromatography, immunoprecipitation, chemical derivatization, chromatography and other newly developed methods. The advantages and disadvantages of these methods, as well as the related optimization and improvement strategies, were discussed in detail. In addition, we also briefly summarized the progress of the combination of phosphopeptide enrichment and fractionation methods developed in recent years.
Phosphopeptides/metabolism* ; Proteomics/methods* ; Titanium/chemistry* ; Chromatography, Affinity ; Proteome ; Phosphorylation

The purpose of this study was to explore the effect of acute hypoxia on urine proteome in rats. In this study, rats were placed in a hypoxic chamber simulating a plateau environment at an altitude of 5 000 m for 24 hours. Urine samples were collected at 0, 12, and 24 h after hypoxia. Urinary proteins were profiled using liquid chromatography coupled with tandem mass spectrometry (LC-MS/MS). Compared with the control (before hypoxia), a total of 144 differentially expressed proteins were identified in the hypoxia 12 h group, and 129 differentially expressed proteins were identified in the hypoxia 24 h group. Functional annotation analysis revealed that these differentially expressed proteins were involved in a series of biological pathways related to hypoxic stress, such as anti-oxidative stress, glycolysis, complement and coagulation cascade. Our results suggest that the urinary proteome can reflect significant changes upon acute hypoxic stimulation. These findings may provide an approach to judge the hypoxia state of the body and help to assist the detection of hypoxia state.
Animals ; Rats ; Proteome/analysis* ; Chromatography, Liquid/methods* ; Tandem Mass Spectrometry/methods* ; Proteomics/methods* ; Hypoxia

Methotrexate (MTX) is a widely used immunosuppressive drug. Large-dose of MTX is used for the treatment of cancer while low-dose is used for the treatment of rheumatoid arthritis (RA). This study aimed to explore the effect of MTX on the urinary proteome of rats. MTX was given to rats orally to construct an MTX intragastric administration rat model. The urine of the rats were collected within 10 hours after giving MTX, and the urine proteins of the rats were analyzed by liquid chromatography tandem mass spectrometry (LC-MS/MS). A total of 31 differential proteins were identified, of which 7 proteins were related to the effect MTX and the symptom of RA. The biological processes of some rats reflected the effect of MTX on the body's glutathione metabolism and the JAK/STAT signaling pathway, which indicated that urine proteins have the ability to reflect the effects of MTX on the body of rats. The spectrum of the differential proteins of each single rat showed that different individuals respond to the drug quite differently.
Rats ; Animals ; Methotrexate/metabolism* ; Proteome ; Chromatography, Liquid/methods* ; Tandem Mass Spectrometry/methods* ; Arthritis, Rheumatoid/drug therapy*