A simplified method is presented for tryptic digestion of Coomassie brilliant blue(CBB)-stained proteins in polyacrylamide gels. Compared with conventional methods, the proposed method does not require a removal of th...A simplified method is presented for tryptic digestion of Coomassie brilliant blue(CBB)-stained proteins in polyacrylamide gels. Compared with conventional methods, the proposed method does not require a removal of the dye before digestion, and is thus faster and saves a lot of labor. The resulted digest can be analyzed by either RPLC/ESIMS or MALDI MS for identification of the protein in a conventional way. Model studies with bovine serum albumin(BSA) showed that 50 ng of the protein could be routinely identified. The simplified procedure displays a tendency to produce more incompletely cleaved peptides, which is favorable for improving the sequence coverage.展开更多
To identify acute renal allograft rejection biomarkers in human serum, two-dimensional differential in-gel electrophoresis （2-D DIGE） and reversed phase high-performance liquid chromatography （RP-HPLC） followed by...To identify acute renal allograft rejection biomarkers in human serum, two-dimensional differential in-gel electrophoresis （2-D DIGE） and reversed phase high-performance liquid chromatography （RP-HPLC） followed by electrospray ionization mass spectrometry （ESI-MS） were used. Serum samples from renal allograft patients and normal volunteers were divided into three groups： acute rejec- tion （AR）, stable renal function （SRF） and normal volunteer （N）. Serum samples were firstly processed using Multiple Affinity Removal Column to selectively remove the highest abundance proteins. Differentially expressed proteins were analyzed using 2-D DIGE. These differential protein spots were excised, digested by trypsin, and identified by RP-HPLC-ESI/MS. Twenty-two differentially expressed proteins were identified in serum from AR group. These proteins included complement C9 precursor, apolipoprotein A-IV precursor, vitamin D-binding protein precursor, beta-2-glycoprotein 1 precursor, etc. Vitamin D-binding protein, one of these proteins, was confirmed by ELISA in the independent set of serum samples. In conclusion, the differentially expressed proteins as serum biomarker candidates may provide the basis of acute rejection noninvasive diagnosis. Confirmed vitamin D-binding protein may be one of serum biomarkers of acute rejection. Furthermore, it may provide great insights into understanding the mechanisms and potential treatment strategy of acute rejection.展开更多
目的探讨宫腔粘连患者宫腔镜下宫腔粘连分离术(transcervical resection of adhesions,TCRA)术后应用Foley球囊+自交联透明质酸钠凝胶预防术后再粘连的临床效果。方法选取我院在2015年12月至2017年11月收治的中重度宫腔粘连患者120例,...目的探讨宫腔粘连患者宫腔镜下宫腔粘连分离术(transcervical resection of adhesions,TCRA)术后应用Foley球囊+自交联透明质酸钠凝胶预防术后再粘连的临床效果。方法选取我院在2015年12月至2017年11月收治的中重度宫腔粘连患者120例,按照随机数字表法分为观察组和对照组,每组各60例;观察组在TCRA术后应用Foley球囊+自交联透明质酸钠凝胶预防再粘连;对照组在TCRA术后应用Foley球囊+普通医用透明质酸钠凝胶预防再粘连。术后3个月观察两组患者临床疗效及AFS评分、粘连范围、粘连类型、月经状况及再粘连状况。结果治疗后观察组临床有效率为86.67%(52/60),对照组临床有效率为71.67%(43/60),比较具有统计学差异(χ2=4.093,P<0.05);术后3个月宫腔镜检查两组患者AFS总分、粘连范围评分、粘连类型及月经状况评分较术前均明显下降(P<0.05);观察组AFS总分、月经状况评分明显低于对照组,差异具有统计学意义(P<0.05);两组患者术后粘连范围、粘连类型比较差异无统计学意义(P>0.05);术后3个月宫腔镜检查,观察组术后再粘连发生率为1.92%(1/52),对照组术后再粘连发生率为16.28%(7/43),比较具有统计学差异(χ2=4.566,P<0.05)。结论宫腔粘连患者TCRA术后应用Foley球囊+自交联透明质酸钠凝胶预防粘连效果显著,能够有效降低AFS评分及月经状况,值得临床推广。展开更多
Background Neurocysticercosis is the infection of the nervous system by the larvae of Taenia solium （T. solium）. Despite continuous effort, the experimental diagnosis of neurocysticercosis remains unresolved. Since ...Background Neurocysticercosis is the infection of the nervous system by the larvae of Taenia solium （T. solium）. Despite continuous effort, the experimental diagnosis of neurocysticercosis remains unresolved. Since the cerebrospinal fluid （CSF） contacts with the brain, dynamic information about pathological processes of the brain is likely to be reflected in CSF. Therefore, CSF may serve as a rich source of putative biomarkers related to neurocysticercosis. Comparative proteomic analysis of CSF of neurocysticercosis patients and control subjects may find differentially expressed proteins.Methods Two-dimensional difference in gel electrophoresis （2D-DIGE） was used to investigate differentially expressed proteins in CSF of patients with neurocysticercosis by comparing the protein profile of CSF from neurocysticercosis patients with that from control subjects. The differentially expressed spots/proteins were recognized with matrix-assisted laser desorption/ionization-time of flight-time of flight （MALDI-TOF-TOF） mass spectrometry.Results Forty-four enzyme digested peptides were obtained from 4 neurocysticercotic patients. Twenty-three were identified through search of the NCBI protein database with Mascot software, showing 19 up-expressed and 4 down-expressed. Of these proteins, 26S proteosome related to ATP- and ubiquitin-dependent degradation of proteins and lipocalin type prostaglandin D synthase involved in PGD2-synthesis and extracellular transporter activities were up-expressed, while transferrin related to iron metabolism within the brain was down-expressed.Conclusions This study established the proteomic profile of pooled CSF from 4 patients with neurocysticercosis, suggesting the potential value of proteomic analysis for the study of candidate biomarkers involved in the diagnosis or pathogenesis of neurocysticercosis.展开更多
Superoxlde dlsmutase （SOD） is ubiquitous in aerobic organisms and constitutes the first link In the enzyme scavenging system of reactive oxygen species. In the present study, species and organ diversity of SOD activ...Superoxlde dlsmutase （SOD） is ubiquitous in aerobic organisms and constitutes the first link In the enzyme scavenging system of reactive oxygen species. In the present study, species and organ diversity of SOD activity In a solution and In an in-gel assay system, as well as the effects of hydrogen peroxide （H202） on SOD activity, were Investigated. In a solution assay system, SOD activity of jackfruIt root, shoot, leaves, axes, and cotyledons, of maize embryos and endosperms, of mung bean leaves and seeds, of sacred lotus axes and cotyledons, and of rice and wheat leaves was Increased by 1-15 mmol/L H2O2. However, SOD activity In rice root and seeds, maize roots and leaves, mung bean roots and shoots, and wheat seeds was decreased by 1-15 mmol/L H2O2. The SOD activity of wheat root and soybean roots, leaves, axes, and cotyledons was Increased by 1-4 mmol/L H2O2, but was decreased by concentrations of H2O2 〉4 mmol/L. The SOD activity of soybean shoots was not affected by 1-15 mmol/L H2O2. The SOD activity In crude mltochondrla of jackfruIt, maize, and upas seeds, as well as In purified mitochondria of jackfruIt, was also Increased by 1-15 mmol/L H2O2. In the In-gel assay system, the SOD In jackfruIt cotyledons was comprised of Mn-SOD, Cu/Zn-SOD, and Fe-SOD, the crude mltochondria of jackfruit seeds and maizes embryo was comprised of Mn-SOD and Cu/ Zn-SOD, and the crude mltochondria of maize seeds was comprised of Mn-SOD only. In the present study, H2O2 markedly Inhibited Cu/Zn-SOD and Fe-SOD activity.展开更多
Parthenogenetic embryonic stem(pES)cells isolated from parthenogenetic activation of oocytes and embryos,also called parthenogenetically induced pluripotent stem cells,exhibit pluripotency evidenced by both in vitro a...Parthenogenetic embryonic stem(pES)cells isolated from parthenogenetic activation of oocytes and embryos,also called parthenogenetically induced pluripotent stem cells,exhibit pluripotency evidenced by both in vitro and in vivo differentiation potential.Differential proteomic analysis was performed using differential in-gel electrophoresis and isotope-coded affinity tag-based quantitative proteomics to investigate the molecular mechanisms underlying the developmental pluripotency of pES cells and to compare the protein expression of pES cells generated from either the in vivo-matured ovulated(IVO)oocytes or from the in vitro-matured(IVM)oocytes with that of fertilized embryonic stem(fES)cells derived from fertilized embryos.A total of 76 proteins were upregulated and 16 proteins were downregulated in the IVM pES cells,whereas 91 proteins were upregulated and 9 were downregulated in the IVO pES cells based on a minimal 1.5-fold change as the cutoff value.No distinct pathways were found in the differentially expressed proteins except for those involved in metabolism and physiological processes.Notably,no differences were found in the protein expression of imprinted genes between the pES and fES cells,suggesting that genomic imprinting can be corrected in the pES cells at least at the early passages.The germline competent IVM pES cells may be applicable for germ cell renewal in aging ovaries if oocytes are retrieved at a younger age.展开更多
基金Supported by the National BasicResearch Priorities Program me of China( No.0 0 1CB5 10 2 0 2),National High- TechProgramm e of China( No.2 0 0 1AA2 30 31)andShanghai Science and Technology Developing Program me( No.0 1JC14 0 11)
文摘A simplified method is presented for tryptic digestion of Coomassie brilliant blue(CBB)-stained proteins in polyacrylamide gels. Compared with conventional methods, the proposed method does not require a removal of the dye before digestion, and is thus faster and saves a lot of labor. The resulted digest can be analyzed by either RPLC/ESIMS or MALDI MS for identification of the protein in a conventional way. Model studies with bovine serum albumin(BSA) showed that 50 ng of the protein could be routinely identified. The simplified procedure displays a tendency to produce more incompletely cleaved peptides, which is favorable for improving the sequence coverage.
基金supported by a grant from National Basic Research 973 Program of China (No.2009CB522407)
文摘To identify acute renal allograft rejection biomarkers in human serum, two-dimensional differential in-gel electrophoresis （2-D DIGE） and reversed phase high-performance liquid chromatography （RP-HPLC） followed by electrospray ionization mass spectrometry （ESI-MS） were used. Serum samples from renal allograft patients and normal volunteers were divided into three groups： acute rejec- tion （AR）, stable renal function （SRF） and normal volunteer （N）. Serum samples were firstly processed using Multiple Affinity Removal Column to selectively remove the highest abundance proteins. Differentially expressed proteins were analyzed using 2-D DIGE. These differential protein spots were excised, digested by trypsin, and identified by RP-HPLC-ESI/MS. Twenty-two differentially expressed proteins were identified in serum from AR group. These proteins included complement C9 precursor, apolipoprotein A-IV precursor, vitamin D-binding protein precursor, beta-2-glycoprotein 1 precursor, etc. Vitamin D-binding protein, one of these proteins, was confirmed by ELISA in the independent set of serum samples. In conclusion, the differentially expressed proteins as serum biomarker candidates may provide the basis of acute rejection noninvasive diagnosis. Confirmed vitamin D-binding protein may be one of serum biomarkers of acute rejection. Furthermore, it may provide great insights into understanding the mechanisms and potential treatment strategy of acute rejection.
文摘目的探讨宫腔粘连患者宫腔镜下宫腔粘连分离术(transcervical resection of adhesions,TCRA)术后应用Foley球囊+自交联透明质酸钠凝胶预防术后再粘连的临床效果。方法选取我院在2015年12月至2017年11月收治的中重度宫腔粘连患者120例,按照随机数字表法分为观察组和对照组,每组各60例;观察组在TCRA术后应用Foley球囊+自交联透明质酸钠凝胶预防再粘连;对照组在TCRA术后应用Foley球囊+普通医用透明质酸钠凝胶预防再粘连。术后3个月观察两组患者临床疗效及AFS评分、粘连范围、粘连类型、月经状况及再粘连状况。结果治疗后观察组临床有效率为86.67%(52/60),对照组临床有效率为71.67%(43/60),比较具有统计学差异(χ2=4.093,P<0.05);术后3个月宫腔镜检查两组患者AFS总分、粘连范围评分、粘连类型及月经状况评分较术前均明显下降(P<0.05);观察组AFS总分、月经状况评分明显低于对照组,差异具有统计学意义(P<0.05);两组患者术后粘连范围、粘连类型比较差异无统计学意义(P>0.05);术后3个月宫腔镜检查,观察组术后再粘连发生率为1.92%(1/52),对照组术后再粘连发生率为16.28%(7/43),比较具有统计学差异(χ2=4.566,P<0.05)。结论宫腔粘连患者TCRA术后应用Foley球囊+自交联透明质酸钠凝胶预防粘连效果显著,能够有效降低AFS评分及月经状况,值得临床推广。
文摘Background Neurocysticercosis is the infection of the nervous system by the larvae of Taenia solium （T. solium）. Despite continuous effort, the experimental diagnosis of neurocysticercosis remains unresolved. Since the cerebrospinal fluid （CSF） contacts with the brain, dynamic information about pathological processes of the brain is likely to be reflected in CSF. Therefore, CSF may serve as a rich source of putative biomarkers related to neurocysticercosis. Comparative proteomic analysis of CSF of neurocysticercosis patients and control subjects may find differentially expressed proteins.Methods Two-dimensional difference in gel electrophoresis （2D-DIGE） was used to investigate differentially expressed proteins in CSF of patients with neurocysticercosis by comparing the protein profile of CSF from neurocysticercosis patients with that from control subjects. The differentially expressed spots/proteins were recognized with matrix-assisted laser desorption/ionization-time of flight-time of flight （MALDI-TOF-TOF） mass spectrometry.Results Forty-four enzyme digested peptides were obtained from 4 neurocysticercotic patients. Twenty-three were identified through search of the NCBI protein database with Mascot software, showing 19 up-expressed and 4 down-expressed. Of these proteins, 26S proteosome related to ATP- and ubiquitin-dependent degradation of proteins and lipocalin type prostaglandin D synthase involved in PGD2-synthesis and extracellular transporter activities were up-expressed, while transferrin related to iron metabolism within the brain was down-expressed.Conclusions This study established the proteomic profile of pooled CSF from 4 patients with neurocysticercosis, suggesting the potential value of proteomic analysis for the study of candidate biomarkers involved in the diagnosis or pathogenesis of neurocysticercosis.
基金Supported by the Knowledge Innovation Program of the Chinese Academy of Sciences(KSCX2-SW-117)Hundreds Talent Program of the Chinese Academy of Sciences,Natural Science Foundation of Yunnan,China (2003C0068M)the Ministry of Sciences and Technology of China (2004DKA30430)
文摘Superoxlde dlsmutase （SOD） is ubiquitous in aerobic organisms and constitutes the first link In the enzyme scavenging system of reactive oxygen species. In the present study, species and organ diversity of SOD activity In a solution and In an in-gel assay system, as well as the effects of hydrogen peroxide （H202） on SOD activity, were Investigated. In a solution assay system, SOD activity of jackfruIt root, shoot, leaves, axes, and cotyledons, of maize embryos and endosperms, of mung bean leaves and seeds, of sacred lotus axes and cotyledons, and of rice and wheat leaves was Increased by 1-15 mmol/L H2O2. However, SOD activity In rice root and seeds, maize roots and leaves, mung bean roots and shoots, and wheat seeds was decreased by 1-15 mmol/L H2O2. The SOD activity of wheat root and soybean roots, leaves, axes, and cotyledons was Increased by 1-4 mmol/L H2O2, but was decreased by concentrations of H2O2 〉4 mmol/L. The SOD activity of soybean shoots was not affected by 1-15 mmol/L H2O2. The SOD activity In crude mltochondrla of jackfruIt, maize, and upas seeds, as well as In purified mitochondria of jackfruIt, was also Increased by 1-15 mmol/L H2O2. In the In-gel assay system, the SOD In jackfruIt cotyledons was comprised of Mn-SOD, Cu/Zn-SOD, and Fe-SOD, the crude mltochondria of jackfruit seeds and maizes embryo was comprised of Mn-SOD and Cu/ Zn-SOD, and the crude mltochondria of maize seeds was comprised of Mn-SOD only. In the present study, H2O2 markedly Inhibited Cu/Zn-SOD and Fe-SOD activity.
基金supported by MOST National Major Basic Research Program(Grant Nos.2010CB94500,2009CB941000(to LL),and 2010CB833703(to FY)).
文摘Parthenogenetic embryonic stem(pES)cells isolated from parthenogenetic activation of oocytes and embryos,also called parthenogenetically induced pluripotent stem cells,exhibit pluripotency evidenced by both in vitro and in vivo differentiation potential.Differential proteomic analysis was performed using differential in-gel electrophoresis and isotope-coded affinity tag-based quantitative proteomics to investigate the molecular mechanisms underlying the developmental pluripotency of pES cells and to compare the protein expression of pES cells generated from either the in vivo-matured ovulated(IVO)oocytes or from the in vitro-matured(IVM)oocytes with that of fertilized embryonic stem(fES)cells derived from fertilized embryos.A total of 76 proteins were upregulated and 16 proteins were downregulated in the IVM pES cells,whereas 91 proteins were upregulated and 9 were downregulated in the IVO pES cells based on a minimal 1.5-fold change as the cutoff value.No distinct pathways were found in the differentially expressed proteins except for those involved in metabolism and physiological processes.Notably,no differences were found in the protein expression of imprinted genes between the pES and fES cells,suggesting that genomic imprinting can be corrected in the pES cells at least at the early passages.The germline competent IVM pES cells may be applicable for germ cell renewal in aging ovaries if oocytes are retrieved at a younger age.
