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Purification and proteomic analysis of potent fibrinolytic enzymes extracted from Lumbricus rubellus. 风疹蚓强效纤溶酶的纯化及蛋白质组学分析。
IF 2 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-05-08 DOI: 10.1186/s12953-023-00206-9
Laurentia Stephani, Puji Rahayu, Debbie Retnoningrum, Maggy Thenawidjaja Suhartono, Heni Rachmawati, Raymond R Tjandrawinata

Background: Lumbrokinase derived from earthworms, Lumbricus rubellus is known to have fibrinolytic enzymes that have potential as therapeutic drugs due to its ability to dissolve fibrin. The current study is aimed to purify the Lumbrokinase from L. rubellus and identify its protein component.

Methods: Water extract of local earthworm Lumbricus rubellus revealed several proteins. Therefore, to identify its protein component, purification through HiPrep DEAE fast flow and proteomic analysis were conducted prior to identifications. A combination of two-dimension gel electrophoresis (2DE) and electrospray ionization mass spectrometry analysis was used to identify the purified fractions.

Results: The purified fractions contain five protein bands, namely F25-1, F25-2, F85-1, F85-2, and F85-3, which displayed strong fibrinogenolytic activity. F25 fractions showed fibrinogenolytic activity of 974.85 U/mg, while F85 fractions showed higher activity of 1,484.11 U/mg. Fractions F85-1, F85-2, and F85-3 showed molecular weights of 42.6 kDa, 27.03 kDa, and 14 kDa, respectively and were identified as Lumbrokinase iso-enzymes.

Conclusion: This preliminary study indicates that the F25 and F85 fractions are similar to published fibrinolytic protease-1 and lumbrokinase, respectively, in terms of their amino acid sequence.

背景:蚓激酶来源于蚯蚓,风疹蚓具有纤维蛋白溶解酶,由于其溶解纤维蛋白的能力,具有作为治疗药物的潜力。本研究旨在从风疹乳杆菌中纯化蚓激酶并鉴定其蛋白组分。方法:从当地蚯蚓rubellus水提液中提取多种蛋白质。因此,为了鉴定其蛋白质成分,在鉴定前通过HiPrep DEAE快速流纯化并进行蛋白质组学分析。采用二维凝胶电泳(2DE)和电喷雾电离质谱分析相结合的方法对纯化组分进行鉴定。结果:纯化后的部分含有F25-1、F25-2、F85-1、F85-2、F85-3 5条蛋白带,具有较强的纤维蛋白原溶解活性。F25组纤维蛋白原溶解活性为974.85 U/mg, F85组活性更高,为1484.11 U/mg。F85-1、F85-2和F85-3的分子量分别为42.6 kDa、27.03 kDa和14 kDa,鉴定为蚓激酶同工酶。结论:本初步研究表明,F25和F85组分的氨基酸序列分别与已发表的纤维蛋白溶解蛋白酶-1和蚓激酶相似。
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引用次数: 1
Proteomics identifies differentially expressed proteins in glioblastoma U87 cells treated with hederagenin. 蛋白质组学鉴定了hederagenin处理的胶质母细胞瘤U87细胞中差异表达的蛋白质。
IF 2 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-04-29 DOI: 10.1186/s12953-023-00208-7
Yesen Zhang, Yi Han, Yuchun Shang, Xiangyu Wang, Jiwei Sun

Objective: We investigated differentially expressed proteins (DEPs) in human glioblastoma U87 cells after treatment with hederagenin as a therapeutic screening mechanism and provided a theoretical basis for hederagenin in treating glioblastoma.

Methods: The Cell Counting Kit 8 assay was used to analyze the inhibitory effect of hederagenin on the proliferation of U87 cells. Protein was identified by tandem mass tags and LC-MS/MS analysis techniques. Annotation of DEPs, Gene Ontology enrichment and function, and Kyoto Encyclopedia of Genes and Genomes pathways and domains were all examined by bioinformatics. According to the TMT results, hub protein was selected from DEPs for WB verification.

Results: Protein quantitative analysis found 6522 proteins in total. Compared with the control group, 43 DEPs (P < 0.05) were involved in the highly enriched signaling pathway in the hederagenin group, among which 20 proteins were upregulated, and 23 proteins were downregulated. These different proteins are mainly involved in the longness regulating pathway-WORM, the hedgehog signaling pathway, Staphylococcus aureus infection, complement, coagulation cascades, and mineral absorption. KIF7 and ATAD2B expression were significantly down-regulated and PHEX and TIMM9 expression were significantly upregulated, according to WB analysis, supporting the TMT findings.

Conclusion: Hederagenin inhibition of GBM U87 cells may be related to KIF7, which is mainly involved in the hedgehog signaling pathway. Our findings lay a foundation for additional study of the therapeutic mechanism of hederagenin.

目的:研究hederagenin治疗人胶质母细胞瘤U87细胞后差异表达蛋白(DEPs)的表达情况,为hederagenin治疗胶质母细胞瘤提供理论依据。方法:采用细胞计数试剂盒8 (Cell Counting Kit 8)检测hederagenin对U87细胞增殖的抑制作用。通过串联质量标签和LC-MS/MS分析技术鉴定蛋白质。利用生物信息学方法对DEPs的注释、基因本体的富集和功能、京都基因基因组百科全书的通路和结构域进行了检测。根据TMT结果,从DEPs中选择hub蛋白进行WB验证。结果:蛋白定量分析共检出6522个蛋白。结论:Hederagenin对GBM U87细胞的抑制作用可能与KIF7有关,KIF7主要参与hedgehog信号通路。本研究结果为进一步研究hederagenin的治疗机制奠定了基础。
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引用次数: 0
Proteomics profiling for the global and acetylated proteins of papillary thyroid cancers. 甲状腺乳头状癌的全蛋白和乙酰化蛋白蛋白质组学分析。
IF 2 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-04-26 DOI: 10.1186/s12953-023-00207-8
Wei Wei, Yuezhang Wu, Dong-Dong Chen, Yuntao Song, Guohui Xu, Qi Shi, Xiao-Ping Dong

Background: Papillary thyroid carcinoma (PTC) is the most common endocrine malignancy cancer among the malignancies of thyroid. Despite of wide usages of proteomics in PTC, the profile of acetylated proteins in PTC remains unsettled, which is helpful for understanding the carcinogenesis mechanism and identifying useful biomarkers for PTC.

Methods: The surgically removed specimens of cancer tissues (Ca-T) and adjacent normal tissues (Ca-N) from 10 female patients pathological diagnosed as PTC (TNM stage III) were enrolled in the study. After preparing the pooled extracts of the whole proteins and the acetylated proteins from 10 cases, TMT labeling and LC/MS/MS methods were applied to the assays of global proteomics and acetylated proteomics separately. Bioinformatics analysis, including KEGG, gene ontology (GO) and hierarchical clustering were performed. Some differentially expressed proteins (DEPs) and differentially expressed acetylated proteins (DEAPs) were validated by individual Western blots.

Results: Controlled with the normal tissues adjacent to the lesions, 147 out of 1923 identified proteins in tumor tissues were considered as DEPs in global proteomics, including 78 up-regulated and 69 down-regulated ones, while 57 out of 311 identified acetylated proteins in tumor tissues were DEAPs in acetylated proteomics, including 32 up-regulated and 25 down-regulated, respectively. The top 3 up- and down-regulated DEPs were fibronectin 1, KRT1B protein and chitinase-3-like protein 1, as well as keratin, type I cytoskeletal 16, A-gamma globin Osilo variant and Huntingtin interacting protein-1. The top 3 up- and down-regulated DEAPs were ribosomal protein L18a-like protein, alpha-1-acid glycoprotein 2 and eukaryotic peptide chain release factor GTP-binding subunit ERF3A, as well as trefoil factor 3, thyroglobulin and histone H2B. Functional GO annotation and KEGG pathway analysis based on the DEPs and DEAPs showed completely different changing pictures. Contrary to the top 10 up- and -down regulated DEPs, most of which were addressed in PTC and other types of carcinomas, changes of the majority DEAPs were not mentioned in the literatures.

Conclusions: Taken the profiling of the global and acetylated proteomics together will provide more broad view of protein alterations on the carcinogenesis and new direction for selecting biomarker for diagnosis of PTC.

背景:甲状腺乳头状癌是甲状腺恶性肿瘤中最常见的内分泌恶性肿瘤。尽管蛋白质组学在PTC中的应用广泛,但PTC中乙酰化蛋白的谱仍然不确定,这有助于了解PTC的致癌机制和识别有用的生物标志物。方法:选取病理诊断为PTC (TNM III期)的10例女性患者手术切除的癌组织(Ca-T)及邻近正常组织(Ca-N)标本。将10例患者的全蛋白和乙酰化蛋白混合提取后,分别采用TMT标记法和LC/MS/MS法进行全蛋白组学和乙酰化蛋白组学分析。生物信息学分析包括KEGG、基因本体(GO)和分层聚类。一些差异表达蛋白(DEPs)和差异表达乙酰化蛋白(DEAPs)通过单独的Western blots验证。结果:以病变旁正常组织为对照,肿瘤组织中鉴定的1923个蛋白中147个为全局蛋白质组学deap,其中上调78个,下调69个;鉴定的肿瘤组织中311个乙酰化蛋白中57个为乙酰化蛋白质组学deap,其中上调32个,下调25个。前3位上调和下调的dep分别是纤维连接蛋白1、KRT1B蛋白和几丁质酶样蛋白1,以及角蛋白、I型细胞骨架16、a - γ珠蛋白Osilo变体和亨廷顿蛋白相互作用蛋白1。上调和下调前3位的deap分别是核糖体蛋白l18a样蛋白、α -1-酸性糖蛋白2和真核肽链释放因子gtp结合亚基ERF3A,以及三叶因子3、甲状腺球蛋白和组蛋白H2B。基于DEPs和DEAPs的功能性GO注释和KEGG通路分析显示了完全不同的变化图。与前10位上下调节的deap(其中大部分在PTC和其他类型的癌症中得到解决)相反,大多数deap的变化在文献中未被提及。结论:将全局蛋白质组学和乙酰化蛋白质组学结合起来,将为PTC的癌变过程提供更广阔的蛋白质改变视角,并为选择诊断PTC的生物标志物提供新的方向。
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引用次数: 1
Effects of Atractylodes lancea extracts on intestinal flora and serum metabolites in mice with intestinal dysbacteriosis. 苍术提取物对肠道菌群及血清代谢物的影响。
IF 2 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-04-15 DOI: 10.1186/s12953-023-00204-x
BaiNian Zhang, Lan Bu, Hui Tian, ZhangQiang You, MingHai Zhao, Jie Tian, YuanYuan Zhang, Qian Wang, ChengJia Tan, Yu Cao, DaRen Feng, ZhenPeng Xi

Objective: This study aims to explore the effect of an extract of Atractylodes lancea (A. lancea) on antibiotics-induced intestinal tract disorder and the probable therapeutic mechanisms employed by this extract to ameliorate these disorders.

Methods: Three days after acclimatization, nine male and nine female specific-pathogen-free (SPF) mice were randomly assigned into three groups: Group C (normal saline), Group M (antibiotic: cefradine + gentamicin), and Group T (antibiotic + A. lancea extract). Each mouse in Groups M and T received intragastric (i.g.) gavage antibiotics containing cefradine and gentamicin sulfate (0.02 ml/g-1/D-1) for 7 days. A. lancea extract (0.02 ml/g-1/D-1) was administered by i.g. gavage to Group T mice for 7 days following the cessation of antibiotic therapy. Group M received an equivalent volume of normal saline for 7 days, while Group C received an equivalent volume of normal saline for 14 days. Afterwards, we collected mouse feces to assess changes in intestinal microbiota by 16S ribosomal ribonucleic acid (rRNA) sequencing and metabolomics. In addition, serum samples were gathered and analyzed using liquid chromatography-mass spectrometry (LS-MS). Finally, we performed a correlation analysis between intestinal microbiota and metabolites.

Results: After treatment with antibiotic, the richness and diversity of the flora, numbers of wall-breaking bacteria and Bacteroidetes, and the numbers of beneficial bacteria decreased, while the numbers of harmful bacteria increased. After i.g. administration of A. lancea extract, the imbalance of microbial flora began to recover. Antibiotics primarily influence the metabolism of lipids, steroids, peptides, organic acids, and carbohydrates, with lipid compounds ranking first. Arachidonic acid (AA), arginine, and proline have relatively strong effects on the metabolisms of antibiotic-stressed mice. Our findings revealed that A. lancea extract might restore the metabolism of AA and L-methionine. The content of differential metabolites detected in the serum of Group T mice was comparable to that in the serum of Group C mice, but significantly different from that of Group M mice. Compared to putative biomarkers in the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, it was found that altered metabolites, such as amino acids, glycerol, and phospholipids, were primarily associated with the metabolism.

Conclusions: The effective mechanisms of A. lancea extract in regulating the disorder of intestinal flora in mice are related to the mechanisms of A. lancea. It could relate to lipid metabolism, bile acid metabolism, and amino acid metabolism. These results will provide a basis for further explaining the mechanism by which A. lancea regulats intestinal flora.

目的:探讨苍术提取物对抗生素性肠道疾病的治疗作用及其可能的作用机制。方法:适应3 d后,将9只雄性和9只雌性无特定病原体(SPF)小鼠随机分为3组:C组(生理盐水)、M组(抗生素:头孢拉定+庆大霉素)和T组(抗生素+ A)。lancea提取)。M组和T组小鼠灌胃含头孢拉定和硫酸庆大霉素的抗生素(0.02 ml/g-1/D-1),连续7 d。停止抗生素治疗后,T组小鼠ig灌胃荷叶提取物0.02 ml/g-1/D-1。M组给予等量生理盐水7 d, C组给予等量生理盐水14 d。随后,我们收集小鼠粪便,通过16S核糖体核糖核酸(rRNA)测序和代谢组学来评估肠道微生物群的变化。此外,收集血清样本并使用液相色谱-质谱法(LS-MS)进行分析。最后,我们进行了肠道微生物群与代谢物之间的相关性分析。结果:抗生素治疗后,肠道菌群的丰富度和多样性、破壁菌和拟杆菌门数量、有益菌数量减少,有害菌数量增加。给药后,微生物菌群的失衡开始恢复。抗生素主要影响脂类、类固醇、多肽、有机酸和碳水化合物的代谢,以脂类化合物为主。花生四烯酸(AA)、精氨酸和脯氨酸对抗生素应激小鼠的代谢有较强的影响。结果表明,刺花提取物具有恢复机体AA和l -蛋氨酸代谢的作用。在T组小鼠血清中检测到的差异代谢物含量与C组小鼠相当,但与M组小鼠差异显著。与京都基因与基因组百科全书(KEGG)数据库中假定的生物标志物相比,发现改变的代谢物,如氨基酸、甘油和磷脂,主要与代谢相关。结论:刺叶提取物调节小鼠肠道菌群紊乱的作用机制与刺叶提取物的作用机制有关。可能与脂质代谢、胆汁酸代谢、氨基酸代谢有关。这些结果将为进一步解释刺草调控肠道菌群的机制提供依据。
{"title":"Effects of Atractylodes lancea extracts on intestinal flora and serum metabolites in mice with intestinal dysbacteriosis.","authors":"BaiNian Zhang,&nbsp;Lan Bu,&nbsp;Hui Tian,&nbsp;ZhangQiang You,&nbsp;MingHai Zhao,&nbsp;Jie Tian,&nbsp;YuanYuan Zhang,&nbsp;Qian Wang,&nbsp;ChengJia Tan,&nbsp;Yu Cao,&nbsp;DaRen Feng,&nbsp;ZhenPeng Xi","doi":"10.1186/s12953-023-00204-x","DOIUrl":"https://doi.org/10.1186/s12953-023-00204-x","url":null,"abstract":"<p><strong>Objective: </strong>This study aims to explore the effect of an extract of Atractylodes lancea (A. lancea) on antibiotics-induced intestinal tract disorder and the probable therapeutic mechanisms employed by this extract to ameliorate these disorders.</p><p><strong>Methods: </strong>Three days after acclimatization, nine male and nine female specific-pathogen-free (SPF) mice were randomly assigned into three groups: Group C (normal saline), Group M (antibiotic: cefradine + gentamicin), and Group T (antibiotic + A. lancea extract). Each mouse in Groups M and T received intragastric (i.g.) gavage antibiotics containing cefradine and gentamicin sulfate (0.02 ml/g<sup>-1</sup>/D<sup>-1</sup>) for 7 days. A. lancea extract (0.02 ml/g<sup>-1</sup>/D<sup>-1</sup>) was administered by i.g. gavage to Group T mice for 7 days following the cessation of antibiotic therapy. Group M received an equivalent volume of normal saline for 7 days, while Group C received an equivalent volume of normal saline for 14 days. Afterwards, we collected mouse feces to assess changes in intestinal microbiota by 16S ribosomal ribonucleic acid (rRNA) sequencing and metabolomics. In addition, serum samples were gathered and analyzed using liquid chromatography-mass spectrometry (LS-MS). Finally, we performed a correlation analysis between intestinal microbiota and metabolites.</p><p><strong>Results: </strong>After treatment with antibiotic, the richness and diversity of the flora, numbers of wall-breaking bacteria and Bacteroidetes, and the numbers of beneficial bacteria decreased, while the numbers of harmful bacteria increased. After i.g. administration of A. lancea extract, the imbalance of microbial flora began to recover. Antibiotics primarily influence the metabolism of lipids, steroids, peptides, organic acids, and carbohydrates, with lipid compounds ranking first. Arachidonic acid (AA), arginine, and proline have relatively strong effects on the metabolisms of antibiotic-stressed mice. Our findings revealed that A. lancea extract might restore the metabolism of AA and L-methionine. The content of differential metabolites detected in the serum of Group T mice was comparable to that in the serum of Group C mice, but significantly different from that of Group M mice. Compared to putative biomarkers in the Kyoto Encyclopedia of Genes and Genomes (KEGG) database, it was found that altered metabolites, such as amino acids, glycerol, and phospholipids, were primarily associated with the metabolism.</p><p><strong>Conclusions: </strong>The effective mechanisms of A. lancea extract in regulating the disorder of intestinal flora in mice are related to the mechanisms of A. lancea. It could relate to lipid metabolism, bile acid metabolism, and amino acid metabolism. These results will provide a basis for further explaining the mechanism by which A. lancea regulats intestinal flora.</p>","PeriodicalId":20857,"journal":{"name":"Proteome Science","volume":"21 1","pages":"5"},"PeriodicalIF":2.0,"publicationDate":"2023-04-15","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10105428/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9685288","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 1
Proteomics analysis of methionine enkephalin upregulated macrophages against infection by the influenza-A virus. 蛋氨酸脑啡肽上调巨噬细胞抗流感a型病毒感染的蛋白质组学分析。
IF 2 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-04-11 DOI: 10.1186/s12953-023-00205-w
Wenrui Fu, Zifeng Xie, Mei Bai, Zhen Zhang, Yuanlong Zhao, Jing Tian

Macrophages have a vital role in phagocytosis and antiviral effect against invading influenza viruses. Previously, we found that methionine enkephalin (MENK) inhibited influenza virus infection by upregulating the "antiviral state" of macrophages. To investigate the immunoregulatory mechanism of action of MENK on macrophages, we employed proteomic analysis to identify differentially expressed proteins (DEPs) between macrophages infected with the influenza-A virus and cells infected with the influenza-A virus after pretreatment with MENK. A total of 215 DEPs were identified: 164 proteins had upregulated expression and 51 proteins had downregulated expression. Proteomics analysis showed that DEPs were highly enriched in "cytokine-cytokine receptor interaction", "phagosome", and "complement and coagulation cascades pathway". Proteomics analysis revealed that MENK could be an immune modulator or prophylactic for the prevention and treatment of influenza. MENK promoted the polarization of M1 macrophages, activated inflammatory responses, and enhanced phagocytosis and killing function by upregulating opsonizing receptors.

巨噬细胞对入侵的流感病毒具有重要的吞噬作用和抗病毒作用。先前,我们发现蛋氨酸脑啡肽(MENK)通过上调巨噬细胞的“抗病毒状态”来抑制流感病毒感染。为了研究MENK对巨噬细胞的免疫调节机制,我们采用蛋白质组学分析方法鉴定了经MENK预处理后感染流感a- a病毒的巨噬细胞与感染流感a- a病毒的细胞之间的差异表达蛋白(DEPs)。共鉴定出215个DEPs,其中164个蛋白表达上调,51个蛋白表达下调。蛋白质组学分析显示,DEPs在“细胞因子-细胞因子受体相互作用”、“吞噬体”和“补体与凝血级联通路”中高度富集。蛋白质组学分析表明,MENK可能是预防和治疗流感的免疫调节剂或预防剂。MENK通过上调调理受体,促进M1巨噬细胞极化,激活炎症反应,增强吞噬和杀伤功能。
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引用次数: 1
Isolation and proteomic profiling of urinary exosomes from patients with colorectal cancer. 结直肠癌患者尿外泌体的分离及蛋白质组学分析。
IF 2 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-02-09 DOI: 10.1186/s12953-023-00203-y
Ling Ma, Haijiao Yu, Yubing Zhu, Kaiyu Xu, Aimin Zhao, Lei Ding, Hong Gao, Man Zhang

Exosomes in the body fluid are effective cell-derived membranous structures transferring various molecules to mediate intercellular communication. The expression of protein in the urinary exosomes from the colorectal cancer (CRC) patients could reflect the characteristics of tumorigenesis. The urinary exosomes with globular membrane structure, the size of 30 ~ 100 nm and positive expression of CD9, CD63 and CD81 were successfully isolated from 9 CRC patients and 3 heathy adults using the density gradient ultracentrifugation. Proteome profiles revealed by label-free liquid chromatography-tandem mass spectrometry (LC-MS/MS) indicated that several proteins were differentially expressed among different stages of CRC. Compared with normal controls, 67 proteins in CRC urinary exosomes were upregulated and 74 proteins were downregulated. The bioinformatics analysis revealed the decreased proteins were related to ESCRT III complex disassembly. The CHMP family was further determined to be the hub of interaction network of proteins enriched in ESCRT signaling. The significant decrease of CHMP4A, CHMP4B, CHMP2A, CHMP2B and CHMP1B were respectively found in the total CRC group and distant metastasis group compared with NC group. Moreover, the CEACAM family also showed significant aberrant changes in the urinary exosomes of CRC patients. The CEACAM7 and CEACAM1 were increased in the CRC patients compared with healthy individuals (P < 0.05). Significant changes of proteomic profile could be found in the urinary exosomes in the CRC patients. The differential expressed urinary exosomes derived proteins showed potential usage in diagnosis and prognosis of CRC.

体液中的外泌体是有效的细胞源性膜结构,传递各种分子以介导细胞间通讯。结直肠癌(CRC)患者尿外泌体中蛋白的表达可以反映肿瘤发生的特点。采用密度梯度超离心技术,从9例结直肠癌患者和3例健康成人中成功分离到CD9、CD63和CD81阳性表达的球状膜结构尿外泌体。无标记液相色谱-串联质谱(LC-MS/MS)分析的蛋白质组谱显示,几种蛋白质在不同阶段的CRC中表达存在差异。与正常对照相比,结直肠癌尿外泌体中67个蛋白上调,74个蛋白下调。生物信息学分析显示,减少的蛋白与ESCRT III复合物的分解有关。进一步确定CHMP家族是ESCRT信号富集蛋白相互作用网络的枢纽。与NC组相比,总结直肠癌组和远处转移组的CHMP4A、CHMP4B、CHMP2A、CHMP2B和CHMP1B的表达均显著降低。此外,CEACAM家族在结直肠癌患者尿外泌体中也表现出显著的异常变化。结直肠癌患者的CEACAM7和CEACAM1与健康人相比增高(P
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引用次数: 3
Quantitative proteomics analysis revealed the potential role of lncRNA Ftx in cardiomyocytes. 定量蛋白质组学分析揭示了lncRNA Ftx在心肌细胞中的潜在作用。
IF 2 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-01-05 DOI: 10.1186/s12953-022-00201-6
Xiangfei Sun, Ying Jiang, Qingbao Li, Qi Tan, Mingliang Dong, Bi'e Cai, Di Zhang, Qi Zhao

Objective: This study aims to decode the proteomic signature of cardiomyocytes in response to lncRNA Ftx knockdown and overexpression via proteomic analysis, and to study the biological role of lncRNA Ftx in cardiomyocytes.  METHODS: The expression level of the lncRNA Ftx in cardiomyocytes cultured in vitro was intervened, and the changes in protein levels in cardiomyocytes were quantitatively detected by liquid chromatography-mass spectrometry. The key molecules and pathways of the lncRNA-Ftx response were further examined by GO, KEGG, and protein interaction analysis.

Results: A total of 2828 proteins are quantified. With a 1.5-fold change threshold, 32 upregulated proteins and 49 downregulated proteins are identified in the lncRNA Ftx overexpression group, while 67 up-regulated proteins and 54 down-regulated proteins are identified in the lncRNA Ftx knockdown group. Functional clustering analysis of differential genes revealed that the lncRNA Ftx is involved in regulating cardiomyocyte apoptosis and ferroptosis and improving cellular energy metabolism. In addition, Hub genes such as ITGB1, HMGA2, STAT3, GSS, and LPCAT3 are regulated downstream by lncRNA Ftx.

Conclusion: This study demonstrates that lncRNA Ftx plays a vital role in cardiomyocytes and may be involved in the occurrence and development of various myocardial diseases. It provides a potential target for clinical protection of the myocardium and reversal of myocardial fibrosis.

目的:本研究旨在通过蛋白质组学分析解码心肌细胞响应lncRNA Ftx敲低和过表达的蛋白质组学特征,研究lncRNA Ftx在心肌细胞中的生物学作用。方法:干预体外培养的心肌细胞中lncRNA Ftx的表达水平,采用液相色谱-质谱联用技术定量检测心肌细胞中蛋白水平的变化。通过GO、KEGG和蛋白相互作用分析进一步研究lncRNA-Ftx反应的关键分子和途径。结果:共定量2828个蛋白。在1.5倍的变化阈值下,lncRNA Ftx过表达组鉴定出32个上调蛋白和49个下调蛋白,lncRNA Ftx敲低组鉴定出67个上调蛋白和54个下调蛋白。差异基因功能聚类分析发现lncRNA Ftx参与调控心肌细胞凋亡和铁凋亡,改善细胞能量代谢。此外,Hub基因如ITGB1、HMGA2、STAT3、GSS和LPCAT3都受lncRNA Ftx的下游调控。结论:本研究表明lncRNA Ftx在心肌细胞中起着至关重要的作用,可能参与了多种心肌疾病的发生发展。它为临床保护心肌和逆转心肌纤维化提供了潜在的靶点。
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引用次数: 2
Proteomic Analysis of Protective Effects of Dl-3-n-Butylphthalide against mpp + -Induced Toxicity via downregulating P53 pathway in N2A Cells. dl -3-n-丁苯酞通过下调P53通路对N2A细胞mpp +诱导毒性的保护作用的蛋白质组学分析。
IF 2 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2023-01-03 DOI: 10.1186/s12953-022-00199-x
Yuan Zhao, Jian Zhang, Yidan Zhang, Shuyue Li, Ya Gao, Cui Chang, Xiang Liu, Lei Xu, Guofeng Yang

Background: Dl-3-n-butylphthalide (NBP) is an important medial therapy for acute ischemic stroke in China. Recent studied have revealed that NBP not only rescued the loss of dopaminergic neurons in cellular and animal models of Parkinson's disease (PD), but also could improve motor symptoms in PD patients. However, the protective mechanism is not fully understood. P53 is a multifunctional protein implicated in numerous cellular processes, including apoptosis, DNA repair, mitochondrial functions, redox homeostasis, autophagy and protein aggregations. In PD, p53 integrated with various neurodegeneration-related signals inducing neuronal loss, indicating the suppression of P53 might be a promising target for PD treatment. Therefore, the purpose of the current study was to systemically screen new therapeutic targets of NBP in PD.

Method: In our study, we constructed mpp + induced N2A cells to investigate the benefit effect of NBP in PD. MTT assay was performed to evaluate the cell viability; TMT-based LC-MS/MS was applied to determine the different expressed proteins (DEPs) of NBP pretreatment; online bioinformatics databases such as DAVID, STRING, and KEGG was used to construe the proteomic data. After further analyzed and visualized the protein-protein interactions (PPI) by Cytoscape, DEPs were verified by western blot.

Result: A total of 5828 proteins were quantified in the comparative proteomics experiments and 417 proteins were considered as DEPs (fold change > 1.5 and p < 0.05). Among the 417 DEPs, 140 were upregulated and 277 were downregulated in mpp + -induced N2A cells with NBP pretreatment. KEGG pathway analysis indicated that lysosome, phagosome, apoptosis, endocytosis and ferroptosis are the mainly enriched pathways. By using MCL clustering in PPI analysis, 48 clusters were generated and the subsequent KEGG analysis of the top 3 clusters revealed that P53 signaling pathway was recognized as the dominant pathway for NBP treatment.

Conclusion: NBP significantly relived mpp + -induced cell toxicity. The neuroprotective role of NBP was implicated with P53 signaling pathway in some extent. These findings will reinforce the understanding of the mechanism of NBP in PD and identify novel therapeutic targets.

背景:dl -3-正丁苯酞(NBP)在中国是治疗急性缺血性脑卒中的重要药物。最近的研究表明,NBP不仅可以挽救帕金森病(PD)细胞和动物模型中多巴胺能神经元的损失,而且可以改善PD患者的运动症状。然而,保护机制尚不完全清楚。P53是一种涉及多种细胞过程的多功能蛋白,包括细胞凋亡、DNA修复、线粒体功能、氧化还原稳态、自噬和蛋白质聚集。在PD中,p53与多种神经退行性相关信号结合,诱导神经元丢失,表明抑制p53可能是PD治疗的一个有希望的靶点。因此,本研究的目的是系统筛选NBP治疗PD的新靶点。方法:构建mpp +诱导的N2A细胞,研究NBP对PD的益处作用。MTT法测定细胞活力;采用基于tmt的LC-MS/MS检测NBP预处理的不同表达蛋白(DEPs);利用DAVID、STRING和KEGG等在线生物信息学数据库构建蛋白质组学数据。通过Cytoscape进一步分析和可视化蛋白相互作用(PPI)后,用western blot验证DEPs。结果:在比较蛋白质组学实验中,共有5828个蛋白被量化,其中417个蛋白被认为是DEPs (fold change > 1.5和p)。结论:NBP可显著缓解mpp +诱导的细胞毒性。NBP的神经保护作用可能与P53信号通路有关。这些发现将加强对NBP在PD中的作用机制的理解,并确定新的治疗靶点。
{"title":"Proteomic Analysis of Protective Effects of Dl-3-n-Butylphthalide against mpp + -Induced Toxicity via downregulating P53 pathway in N2A Cells.","authors":"Yuan Zhao,&nbsp;Jian Zhang,&nbsp;Yidan Zhang,&nbsp;Shuyue Li,&nbsp;Ya Gao,&nbsp;Cui Chang,&nbsp;Xiang Liu,&nbsp;Lei Xu,&nbsp;Guofeng Yang","doi":"10.1186/s12953-022-00199-x","DOIUrl":"https://doi.org/10.1186/s12953-022-00199-x","url":null,"abstract":"<p><strong>Background: </strong>Dl-3-n-butylphthalide (NBP) is an important medial therapy for acute ischemic stroke in China. Recent studied have revealed that NBP not only rescued the loss of dopaminergic neurons in cellular and animal models of Parkinson's disease (PD), but also could improve motor symptoms in PD patients. However, the protective mechanism is not fully understood. P53 is a multifunctional protein implicated in numerous cellular processes, including apoptosis, DNA repair, mitochondrial functions, redox homeostasis, autophagy and protein aggregations. In PD, p53 integrated with various neurodegeneration-related signals inducing neuronal loss, indicating the suppression of P53 might be a promising target for PD treatment. Therefore, the purpose of the current study was to systemically screen new therapeutic targets of NBP in PD.</p><p><strong>Method: </strong>In our study, we constructed mpp + induced N2A cells to investigate the benefit effect of NBP in PD. MTT assay was performed to evaluate the cell viability; TMT-based LC-MS/MS was applied to determine the different expressed proteins (DEPs) of NBP pretreatment; online bioinformatics databases such as DAVID, STRING, and KEGG was used to construe the proteomic data. After further analyzed and visualized the protein-protein interactions (PPI) by Cytoscape, DEPs were verified by western blot.</p><p><strong>Result: </strong>A total of 5828 proteins were quantified in the comparative proteomics experiments and 417 proteins were considered as DEPs (fold change > 1.5 and p < 0.05). Among the 417 DEPs, 140 were upregulated and 277 were downregulated in mpp + -induced N2A cells with NBP pretreatment. KEGG pathway analysis indicated that lysosome, phagosome, apoptosis, endocytosis and ferroptosis are the mainly enriched pathways. By using MCL clustering in PPI analysis, 48 clusters were generated and the subsequent KEGG analysis of the top 3 clusters revealed that P53 signaling pathway was recognized as the dominant pathway for NBP treatment.</p><p><strong>Conclusion: </strong>NBP significantly relived mpp + -induced cell toxicity. The neuroprotective role of NBP was implicated with P53 signaling pathway in some extent. These findings will reinforce the understanding of the mechanism of NBP in PD and identify novel therapeutic targets.</p>","PeriodicalId":20857,"journal":{"name":"Proteome Science","volume":"21 1","pages":"1"},"PeriodicalIF":2.0,"publicationDate":"2023-01-03","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9809048/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10855487","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 3
Responses of the tree peony (Paeonia suffruticosa, Paeoniaceae) cultivar 'Yu Hong' to heat stress revealed by iTRAQ-based quantitative proteomics. 基于iTRAQ的定量蛋白质组学揭示树牡丹(芍药科)栽培品种'玉红'对热胁迫的响应
IF 2.1 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2022-12-29 DOI: 10.1186/s12953-022-00202-5
Jin Ma, Qun Wang, Ling-Ling Wei, Yu Zhao, Guo-Zhe Zhang, Jie Wang, Cui-Hua Gu

Horticulture productivity has been increasingly restricted by heat stress from growing global warming, making it far below the optimum production capacity. As a popular ornamental cultivar of tree peony, Paeonia suffruticosa 'Yu Hong' has also been suffering from heat stress not suitable for its optimal growth. To better understand the response mechanisms against heat stress of tree peony, investigations of phenotypic changes, physiological responses, and quantitative proteomics were conducted. Phenotypic and physiological changes indicated that 24 h of exposure to heat stress (40 °C) was the critical duration of heat stress in tree peony. The proteomic analyses revealed a total of 100 heat-responsive proteins (HRPs). According to bioinformatic analysis of HRPs, the heat tolerance of tree peony might be related to signal transduction, synthesis/degradation, heat kinetic proteins, antioxidants, photosynthesis, energy conversion, and metabolism. Our research will provide some new insights into the molecular mechanism under the response against the heat stress of tree peony, which will benefit the future breeding of heat-resistant ornamental plants.

由于全球气候日益变暖,园艺生产越来越受到热胁迫的限制,远远达不到最佳生产能力。作为深受人们喜爱的树牡丹观赏栽培品种,芍药'玉红'也受到了不适合其最佳生长的热胁迫的影响。为了更好地了解树牡丹对热胁迫的响应机制,研究人员对其表型变化、生理响应和定量蛋白质组学进行了研究。表型和生理变化表明,暴露于热胁迫(40 °C)24 小时是树牡丹热胁迫的临界期。蛋白质组学分析共发现了 100 个热响应蛋白(HRPs)。根据对HRPs的生物信息学分析,树牡丹的耐热性可能与信号转导、合成/降解、热动力学蛋白、抗氧化剂、光合作用、能量转换和新陈代谢有关。我们的研究将为树牡丹应对热胁迫的分子机制提供一些新的见解,这将有利于未来耐热观赏植物的育种。
{"title":"Responses of the tree peony (Paeonia suffruticosa, Paeoniaceae) cultivar 'Yu Hong' to heat stress revealed by iTRAQ-based quantitative proteomics.","authors":"Jin Ma, Qun Wang, Ling-Ling Wei, Yu Zhao, Guo-Zhe Zhang, Jie Wang, Cui-Hua Gu","doi":"10.1186/s12953-022-00202-5","DOIUrl":"10.1186/s12953-022-00202-5","url":null,"abstract":"<p><p>Horticulture productivity has been increasingly restricted by heat stress from growing global warming, making it far below the optimum production capacity. As a popular ornamental cultivar of tree peony, Paeonia suffruticosa 'Yu Hong' has also been suffering from heat stress not suitable for its optimal growth. To better understand the response mechanisms against heat stress of tree peony, investigations of phenotypic changes, physiological responses, and quantitative proteomics were conducted. Phenotypic and physiological changes indicated that 24 h of exposure to heat stress (40 °C) was the critical duration of heat stress in tree peony. The proteomic analyses revealed a total of 100 heat-responsive proteins (HRPs). According to bioinformatic analysis of HRPs, the heat tolerance of tree peony might be related to signal transduction, synthesis/degradation, heat kinetic proteins, antioxidants, photosynthesis, energy conversion, and metabolism. Our research will provide some new insights into the molecular mechanism under the response against the heat stress of tree peony, which will benefit the future breeding of heat-resistant ornamental plants.</p>","PeriodicalId":20857,"journal":{"name":"Proteome Science","volume":"20 1","pages":"18"},"PeriodicalIF":2.1,"publicationDate":"2022-12-29","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9798725/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10453244","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}
引用次数: 0
TMT-based quantitative proteomics analysis of the effects of Jiawei Danshen decoction myocardial ischemia-reperfusion injury. 基于tmt的加味丹参汤对心肌缺血再灌注损伤的定量蛋白质组学分析。
IF 2 3区 生物学 Q3 BIOCHEMICAL RESEARCH METHODS Pub Date : 2022-12-14 DOI: 10.1186/s12953-022-00200-7
Xiang-Mei Zhu, Yang Tan, Yu-He Shi, Qing Li, Jue Zhu, Xiang-Dan Liu, Qiao-Zhen Tong

Background: Every year, approximately 17 million people worldwide die due to coronary heart disease, with China ranking second in terms of the death toll. Myocardial ischemia-reperfusion injury (MIRI) significantly influences cardiac function and prognosis in cardiac surgery patients. Jiawei Danshen Decoction (JWDSD) is a traditional Chinese herbal prescription that has been used clinically for many years in China to treat MIRI. The underlying molecular mechanisms, however, remain unknown. To investigate the proteomic changes in myocardial tissue of rats given JWDSD for MIRI therapy-based proteomics.

Methods: MIRI rat model was created by ligating/releasing the left anterior descending coronary artery. For seven days, the drugs were administered twice daily. The model was created following the last drug administration. JWDSD's efficacy in improving MIRI was evaluated using biochemical markers and cardiac histology. Tandem mass tag-based quantitative proteomics (TMT) technology was also used to detect proteins in the extracted heart tissue. To analyze differentially expressed proteins (DEPs), bioinformatics analysis, including gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) pathways, were employed. Furthermore, western blotting confirmed the potential targets regulated by JWDSD.

Results: The histopathologic characteristics and biochemical data showed JWDSD's protective effects on MIRI rats. A total of 4549 proteins were identified with FDR (false discovery rate) ≤1%. Twenty overlapping were identified (162 DEPs and 45 DEPs in Model/Control or JWDSD/Model group, respectively). Of these DEPs, 16 were regulated by JWDSD. GO analysis provided a summary of the deregulated protein expression in the categories of biological process (BP), cell component (CC), and molecular function (MF). KEGG enrichment analysis revealed that the signaling pathways of neutrophil extracellular trap formation, RNA polymerase, serotonergic synapse, and linoleic acid metabolism are all closely related to JWDSD effects in MIRI rats. Furthermore, T-cell lymphoma invasion and metastasis 1 (TIAM1) was validated using western blotting, and the results were consistent with proteomics data.

Conclusions: Our study suggests that JWDSD may exert therapeutic effects through multi-pathways regulation in MIRI treatment. This work may provide proteomics clues for continuing research on JWDSD in treating MIRI.

背景:全世界每年约有1700万人死于冠心病,其中中国的死亡人数排名第二。心肌缺血再灌注损伤(MIRI)对心脏手术患者心功能和预后有显著影响。加味丹参汤(JWDSD)是中国临床上用于治疗MIRI多年的传统中药方剂。然而,其潜在的分子机制尚不清楚。目的:研究JWDSD给药大鼠心肌组织蛋白质组学的变化,并进行基于MIRI治疗的蛋白质组学研究。方法:结扎/释放左冠状动脉前降支,建立MIRI大鼠模型。在七天的时间里,每天给药两次。这个模型是在最后一次给药后创建的。采用生化指标和心脏组织学指标评价JWDSD改善MIRI的疗效。串联质量标记定量蛋白质组学(TMT)技术也被用于检测提取的心脏组织中的蛋白质。为了分析差异表达蛋白(DEPs),采用了生物信息学分析,包括基因本体(GO)和京都基因与基因组百科全书(KEGG)途径。此外,western blotting证实了JWDSD调控的潜在靶点。结果:JWDSD对MIRI大鼠具有一定的保护作用。共鉴定出4549个蛋白,FDR(错误发现率)≤1%。共发现20个重叠位点(模型/控制组162个,JWDSD/模型组45个)。在这些dep中,有16个受JWDSD调控。氧化石墨烯分析提供了在生物过程(BP)、细胞成分(CC)和分子功能(MF)类别中解除调节的蛋白质表达的总结。KEGG富集分析显示,中性粒细胞胞外陷阱形成、RNA聚合酶、血清素能突触和亚油酸代谢等信号通路均与MIRI大鼠JWDSD效应密切相关。此外,使用western blotting验证t细胞淋巴瘤侵袭和转移1 (TIAM1),结果与蛋白质组学数据一致。结论:本研究提示JWDSD可能通过多途径调控在MIRI治疗中发挥作用。这项工作可能为继续研究JWDSD治疗MIRI提供蛋白质组学线索。
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引用次数: 0
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Proteome Science
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