同位素标记相对和绝对定量(isobaric tags for relative and absolute quantitation,iTRAQ)技术—利用多种同位素试剂标记蛋白多肽N末端或赖氨酸侧链基团,经高精度质谱仪串联分析,可同时比较多达8种样品之间的蛋白表达量,是近年来定量...同位素标记相对和绝对定量(isobaric tags for relative and absolute quantitation,iTRAQ)技术—利用多种同位素试剂标记蛋白多肽N末端或赖氨酸侧链基团,经高精度质谱仪串联分析,可同时比较多达8种样品之间的蛋白表达量,是近年来定量蛋白质组学常用的高通量筛选技术,正迅速被广大学者所接受。主要从iTRAQ实验技术概要、iTRAQ多重化学标记串联质谱技术在生命科学领域中的应用、问题与展望等三个方面进行简要总结。展开更多
Objective The aim of the study was to investigate the expression of proteins in colonic tissues of mice with ulcerative colitis(UC) by using isobaric tags for relative and absolute quantitation(iTRAQ), probe into the ...Objective The aim of the study was to investigate the expression of proteins in colonic tissues of mice with ulcerative colitis(UC) by using isobaric tags for relative and absolute quantitation(iTRAQ), probe into the pathogenesis of UC, and find potential biomarkers of UC. Methods Forty C57 mice were randomly divided into the control and model groups(20 mice in each group). The mice in the model group were administered dextran sulphate sodium(DSS) for 7 consecutive days ad libitum to induce acute colitis, and the colon tissue was extracted on the 8 th day after the successful establishment of the UC model. Proteins were identified by the i TRAQ and tandem mass spectrometry techniques,and the identified proteins were analyzed by bioinformatics. Results A total of 4019 proteins were identified among the two groups. Among them, 317 significant differentially expressed proteins(DEPs) were detected according to the screening criteria for selecting DEPs, i.e. fold change ratios ≥ 1.5 or ≤ 0.67 and P-values < 0.05, of which 156 were upregulated and 161 were downregulated. In the Gene Ontology(GO) analysis, the DEPs were classified into 48 functional categories, which contained biological process, cellular component, and molecular function. Based on the 317 DEPs, the KEGG pathway analysis identified 160 vital pathways.Conclusion DEPs in colonic tissues of mice with UC were screened using the iTRAQ technique, which laid a foundation for further studies regarding the pathogenesis of UC.展开更多
A growing body of evidence has linked the gut microbiota to liver metabolism.The manipulation of intestinal microflora has been considered as a promising avenue to promote liver health.However,the effects of Lactobaci...A growing body of evidence has linked the gut microbiota to liver metabolism.The manipulation of intestinal microflora has been considered as a promising avenue to promote liver health.However,the effects of Lactobacillus gasseri LA39,a potential probiotic,on liver metabolism remain unclear.Accumulating studies have investigated the proteomic profile for mining the host biological events affected by microbes,and used the germ-free(GF)mouse model to evaluate host-microbe interaction.Here,we explored the effects of L.gasseri LA39 gavage on the protein expression profiles of the liver of GF mice.Our results showed that a total of 128 proteins were upregulated,whereas a total of 123 proteins were downregulated by treatment with L.gasseri LA39.Further bioinformatics analyses suggested that the primary bile acid(BA)biosynthesis pathway in the liver was activated by L.gasseri LA39.Three differentially expressed proteins(cytochrome P450 family 27 subfamily A member 1(CYP27A1),cytochrome P450 family 7 subfamily B member 1(CYP7B1),and cytochrome P450 family 8 subfamily B member 1(CYP8B1))involved in the primary BA biosynthesis pathway were further validated by western blot assay.In addition,targeted metabolomic analyses demonstrated that serum and fecalβ-muricholic acid(a primary BA),dehydrolithocholic acid(a secondary BA),and glycolithocholic acid-3-sulfate(a secondary BA)were significantly increased by L.gasseri LA39.Thus,our data revealed that L.gasseri LA39 activates the hepatic primary BA biosynthesis and promotes the intestinal secondary BA biotransformation.Based on these findings,we suggest that L.gasseri LA39 confers an important function in the gut‒liver axis through regulating BA metabolism.展开更多
Objective:Using isobaric tags for relative and absolute quantitation(i TRAQ)technology to study differential protein expression in the retinal tissue of DR mouse models,providing proteomic evidence at the protein leve...Objective:Using isobaric tags for relative and absolute quantitation(i TRAQ)technology to study differential protein expression in the retinal tissue of DR mouse models,providing proteomic evidence at the protein level for the pathogenesis of DR.Methods:Firstly,establish diabetic mice and DR mice models,use i TRAQ technology to detect the retinal tissue samples of normal control group and DR model group mice,label the total retinal proteins of mice with i TRAQ reagent,and analyze them using mass spectrometry technology.Evaluate the differential proteins of the two groups using BioWorks TM 3.0 software,and conduct interaction feature analysis on the different proteins using the STRING website.Results:The i TRAQ technology detected a total of 406 different proteins between the diabetes and normal control groups,with 19 of them closely related to retinal cell apoptosis.Among them,significantly different proteins include acyl-CoA dehydrogenase short chain specific(ACADS),ataxin-10(ATXN10),BCL-2-associated X protein(BAX),caspase-3(CASP3),collagen type IIα1 chain(COL4A2),glycyl-tRNA synthetase(GARS),glial fibrillary acidic protein(GFAP),legumain(LGMN),mucin-4(MUC4),N-myc downstream-regulated gene 1 protein(NDRG1),with ratios to internal controls in the normal group of 1.67,2.06,1.76,2.16,1.53,1.87,1.24,1.61,0.42,0.56,respectively;and there is a potential functional association between GFAP,CASP3,and BAX proteins.Conclusion:In DR mice retinas,there are abnormal changes in the expression of a large number of apoptosis-related proteins.i TRAQ technology can effectively screen out key apoptosis proteins,among which GFAP,CASP3,and BAX may have adverse effects on the progression of DR by participating in the apoptosis process.The application of i TRAQ technology can provide new technical support for proteomic research on apoptosis proteins.展开更多
The molecular mechanism of triple-negative breast cancer（TNBC） remains unclear, and there has been no effective targeted therapy for it. A better understanding of the mechanisms of TNBC is urgently needed to identif...The molecular mechanism of triple-negative breast cancer（TNBC） remains unclear, and there has been no effective targeted therapy for it. A better understanding of the mechanisms of TNBC is urgently needed to identify new therapeutic targets. In this study, eight cases of premenopausal TNBC patients were collected, and a comparative proteomic analysis of their breast cancer tissues and matched paraneoplastic ones was performed via isobaric tags for relative and absolute quantitation（iTRAQ） technology coupled with two-dimensional liquid chromatography-tandem mass spectrumetry（2D LC-MS/MS）. The researches result in the identification of 1254 nonredundant proteins, of which 1243 proteins reached the strict quantitative standard. The quantitative comparison reveal that among the 214 proteins, 81 proteins significantly increased and 133 proteins decreased in TNBC tissues compared to corresponding ones in control. The Gene Ontology（GO） annotations and pathway analysis show their distributions in GO and the marked functions, as well as the closely related signal transduction pathways involved in extra cellular matrix （ECM）-receptor interaction, protein digestion and absorption, renin-angiotensin system, complement and coagulation cascades and focal adhesion. This pilot study will lay a foundation for further searching for therapeutic targets of TNBC and exploring the molecular mechanism, which can also be extended as a part of a large scale biomarker discovery plan.展开更多
文摘同位素标记相对和绝对定量(isobaric tags for relative and absolute quantitation,iTRAQ)技术—利用多种同位素试剂标记蛋白多肽N末端或赖氨酸侧链基团,经高精度质谱仪串联分析,可同时比较多达8种样品之间的蛋白表达量,是近年来定量蛋白质组学常用的高通量筛选技术,正迅速被广大学者所接受。主要从iTRAQ实验技术概要、iTRAQ多重化学标记串联质谱技术在生命科学领域中的应用、问题与展望等三个方面进行简要总结。
基金Supported by a grant from the Natural Science Foundation of Hubei Province(No.2011CHB025)
文摘Objective The aim of the study was to investigate the expression of proteins in colonic tissues of mice with ulcerative colitis(UC) by using isobaric tags for relative and absolute quantitation(iTRAQ), probe into the pathogenesis of UC, and find potential biomarkers of UC. Methods Forty C57 mice were randomly divided into the control and model groups(20 mice in each group). The mice in the model group were administered dextran sulphate sodium(DSS) for 7 consecutive days ad libitum to induce acute colitis, and the colon tissue was extracted on the 8 th day after the successful establishment of the UC model. Proteins were identified by the i TRAQ and tandem mass spectrometry techniques,and the identified proteins were analyzed by bioinformatics. Results A total of 4019 proteins were identified among the two groups. Among them, 317 significant differentially expressed proteins(DEPs) were detected according to the screening criteria for selecting DEPs, i.e. fold change ratios ≥ 1.5 or ≤ 0.67 and P-values < 0.05, of which 156 were upregulated and 161 were downregulated. In the Gene Ontology(GO) analysis, the DEPs were classified into 48 functional categories, which contained biological process, cellular component, and molecular function. Based on the 317 DEPs, the KEGG pathway analysis identified 160 vital pathways.Conclusion DEPs in colonic tissues of mice with UC were screened using the iTRAQ technique, which laid a foundation for further studies regarding the pathogenesis of UC.
文摘目的研究桃核承气汤延缓肾间质纤维化(renal intersitial fibrosis,RIF)的分子机制。方法30只SPF级健康wistar雄性大鼠,随机分为假手术组、模型组、药物组,每组10只;假手术组游离单侧输尿管,模型组及药物组采用单侧输尿管梗阻方法建立RIF模型。假手术组、模型组、药物组自术后第一天开始,分别灌服5 mL蒸馏水、5 mL蒸馏水、5 mL桃核承气汤,每日一次。14天后取梗阻侧肾组织,用绝对和相对定量同位素标记(isobaric tags for relative and absolute quantification,iTRAQ)技术筛选差异蛋白,进行蛋白质组学检测,基因本体(gene ontology,GO)分析、京都基因和基因组百科全书(Kyoto Encyclopedia of Genes and Genomes,KEGG)比对及聚类分析。结果桃核承气汤具有延缓单侧输卵管梗阻造成RIF的作用;药物组与模型组相比筛选的差异蛋白共343种,其中与RIF密切相关的蛋白16种,11种蛋白表达上调,5种蛋白表达下调。GO分析主要涉及氧化还原、药物代谢等生物学过程;线粒体基质、线粒体内膜等细胞组分;氧化还原酶活性等分子功能,KEGG分析涉及AMPK、FoxO/Wnt、PPAR、HIF-1等多条信号通路。结论桃核承气汤延缓肾间质纤维化的分子机制可能与调节磷酸烯醇式丙酮酸羧基酶1(phosphoenolpyruvate carboxykinase 1,PCK1)等蛋白,影响AMPK、FoxO/Wnt、PPAR、HIF-1等信号通路相关。
基金supported by the National Natural Science Foundation of China(Nos.31925037,31730090,and 32102499)the National Postdoctoral Program for Innovative Talents of China(No.BX20190133)+1 种基金the Postdoctoral Science Foundation of China(No.2019M662671)the Natural Science Foundation of Hubei Province(Nos.2022CFB358 and 2021CFA018).
文摘A growing body of evidence has linked the gut microbiota to liver metabolism.The manipulation of intestinal microflora has been considered as a promising avenue to promote liver health.However,the effects of Lactobacillus gasseri LA39,a potential probiotic,on liver metabolism remain unclear.Accumulating studies have investigated the proteomic profile for mining the host biological events affected by microbes,and used the germ-free(GF)mouse model to evaluate host-microbe interaction.Here,we explored the effects of L.gasseri LA39 gavage on the protein expression profiles of the liver of GF mice.Our results showed that a total of 128 proteins were upregulated,whereas a total of 123 proteins were downregulated by treatment with L.gasseri LA39.Further bioinformatics analyses suggested that the primary bile acid(BA)biosynthesis pathway in the liver was activated by L.gasseri LA39.Three differentially expressed proteins(cytochrome P450 family 27 subfamily A member 1(CYP27A1),cytochrome P450 family 7 subfamily B member 1(CYP7B1),and cytochrome P450 family 8 subfamily B member 1(CYP8B1))involved in the primary BA biosynthesis pathway were further validated by western blot assay.In addition,targeted metabolomic analyses demonstrated that serum and fecalβ-muricholic acid(a primary BA),dehydrolithocholic acid(a secondary BA),and glycolithocholic acid-3-sulfate(a secondary BA)were significantly increased by L.gasseri LA39.Thus,our data revealed that L.gasseri LA39 activates the hepatic primary BA biosynthesis and promotes the intestinal secondary BA biotransformation.Based on these findings,we suggest that L.gasseri LA39 confers an important function in the gut‒liver axis through regulating BA metabolism.
文摘Objective:Using isobaric tags for relative and absolute quantitation(i TRAQ)technology to study differential protein expression in the retinal tissue of DR mouse models,providing proteomic evidence at the protein level for the pathogenesis of DR.Methods:Firstly,establish diabetic mice and DR mice models,use i TRAQ technology to detect the retinal tissue samples of normal control group and DR model group mice,label the total retinal proteins of mice with i TRAQ reagent,and analyze them using mass spectrometry technology.Evaluate the differential proteins of the two groups using BioWorks TM 3.0 software,and conduct interaction feature analysis on the different proteins using the STRING website.Results:The i TRAQ technology detected a total of 406 different proteins between the diabetes and normal control groups,with 19 of them closely related to retinal cell apoptosis.Among them,significantly different proteins include acyl-CoA dehydrogenase short chain specific(ACADS),ataxin-10(ATXN10),BCL-2-associated X protein(BAX),caspase-3(CASP3),collagen type IIα1 chain(COL4A2),glycyl-tRNA synthetase(GARS),glial fibrillary acidic protein(GFAP),legumain(LGMN),mucin-4(MUC4),N-myc downstream-regulated gene 1 protein(NDRG1),with ratios to internal controls in the normal group of 1.67,2.06,1.76,2.16,1.53,1.87,1.24,1.61,0.42,0.56,respectively;and there is a potential functional association between GFAP,CASP3,and BAX proteins.Conclusion:In DR mice retinas,there are abnormal changes in the expression of a large number of apoptosis-related proteins.i TRAQ technology can effectively screen out key apoptosis proteins,among which GFAP,CASP3,and BAX may have adverse effects on the progression of DR by participating in the apoptosis process.The application of i TRAQ technology can provide new technical support for proteomic research on apoptosis proteins.
基金Supported by the National Natural Science Foundation of China（No.81041098）.
文摘The molecular mechanism of triple-negative breast cancer（TNBC） remains unclear, and there has been no effective targeted therapy for it. A better understanding of the mechanisms of TNBC is urgently needed to identify new therapeutic targets. In this study, eight cases of premenopausal TNBC patients were collected, and a comparative proteomic analysis of their breast cancer tissues and matched paraneoplastic ones was performed via isobaric tags for relative and absolute quantitation（iTRAQ） technology coupled with two-dimensional liquid chromatography-tandem mass spectrumetry（2D LC-MS/MS）. The researches result in the identification of 1254 nonredundant proteins, of which 1243 proteins reached the strict quantitative standard. The quantitative comparison reveal that among the 214 proteins, 81 proteins significantly increased and 133 proteins decreased in TNBC tissues compared to corresponding ones in control. The Gene Ontology（GO） annotations and pathway analysis show their distributions in GO and the marked functions, as well as the closely related signal transduction pathways involved in extra cellular matrix （ECM）-receptor interaction, protein digestion and absorption, renin-angiotensin system, complement and coagulation cascades and focal adhesion. This pilot study will lay a foundation for further searching for therapeutic targets of TNBC and exploring the molecular mechanism, which can also be extended as a part of a large scale biomarker discovery plan.
