以在新疆昌吉某猪场分离的一株猪肺炎支原体(CJ)为试验材料,采用SDS-PAGE和Western-Blot印迹对该菌的膜蛋白进行了分析。该猪肺炎支原体(CJ)经过Friis改良液体培养基培养,以12 000 r/min离心收集菌体,低温反复冻融获得膜蛋白后,应用12%...以在新疆昌吉某猪场分离的一株猪肺炎支原体(CJ)为试验材料,采用SDS-PAGE和Western-Blot印迹对该菌的膜蛋白进行了分析。该猪肺炎支原体(CJ)经过Friis改良液体培养基培养,以12 000 r/min离心收集菌体,低温反复冻融获得膜蛋白后,应用12%分离胶,4%浓缩胶进行SDS-PAGE分离,在电泳图谱上呈现大概26条蛋白条带,分子量在10 k D^200 k D。再以膜蛋白做免疫印迹检测,其Western-Blot膜上有14种反应条带,其中p36、p38、p40、p52、p65,p70、p114、p200条带颜色较深。展开更多
Since preparation and solubilization of protein samples are crucial factors in proteome research,the authors established a sequential extraction technique to prepare protein samples from the body wall of the 5th insta...Since preparation and solubilization of protein samples are crucial factors in proteome research,the authors established a sequential extraction technique to prepare protein samples from the body wall of the 5th instar larvae of silkworm.Bombyx mori.Two kinds of protein samples were obtained from the body wall using the method.Between the two types of samples only about 15% proteins were identical;the majority were different,indicating that more species of proteins could be obtained with the sequential extraction method;which will be useful for preparation of protein samples for proteome study.展开更多
Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was applied in the detection of the end point temperature (EPT) of thermal denatured protein in fish and meat in this study. It was also used in stu...Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was applied in the detection of the end point temperature (EPT) of thermal denatured protein in fish and meat in this study. It was also used in studying the thermal denatured temperature range of proteins in salmon and chicken meat. The results show that the temperature ranges of denatured proteins were from 65 ℃ to 75 ℃ , and these temperature ranges were influenced by the processing methods. Through SDS-PAGE, the features of repeated heating thermal denatured proteins under the same temperature and processing time were studied. The electrophoresis patterns of thermal denatured proteins determined through repeated heating at the same temperature did not exhibit any change. For the detection of cooked fish and meat samples, they were subjected to applying the SDS-PAGE method, which revealed an EPT ranging from 60 ℃ to 80 ℃ .展开更多
Idiopathic pulmonary arterial hypertension(IPAH) is a rare disease of unknown etiology.The exact pathogenesis of pulmonary arterial hypertension is still not well known.In the past decades,many protein molecules have ...Idiopathic pulmonary arterial hypertension(IPAH) is a rare disease of unknown etiology.The exact pathogenesis of pulmonary arterial hypertension is still not well known.In the past decades,many protein molecules have been found to be in-volved in the development of IPAH.With proteomic techniques,profiling of human plasma proteome becomes more feasible in searching for disease-related markers.In present study,we showed the protein expression profiles of the serum of IPAH and healthy controls after depleting a few high-abundant proteins in serum.Thirteen spots had changed significantly in IPAH com-pared with healthy controls and were identified by LC-MS/MS.Alpha-1-antitrypsin and vitronectin were down-regulated in IPAH and may be valuable candidates for further explorations of their roles in the development of IPAH.展开更多
Early detection of pregnancy is advantageous to breeders. However, tools that were used in the early pregnancy detection are expensive, laboratory-based and not efficient and applicable enough in the field to farmers....Early detection of pregnancy is advantageous to breeders. However, tools that were used in the early pregnancy detection are expensive, laboratory-based and not efficient and applicable enough in the field to farmers. Thus, there is a need to find a biomarker which not only can detect pregnancy but also could be applied to the livestock on the field. Proteomic approach was used in this study as to search for early pregnancy biomarker. When goat sera at day 14 of gestation were resolved by using two dimensional gel electrophoresis, five differentially expressed proteins were detected. Four of the proteins were identified as albumin precursors, immunoglobulin lambda light chain and zinc-alpha-2-glycoprotein. Pregnancy associated glycoprotein-1 was detected by using liquid chromatography mass spectrometry and the protein was validated by immunoblotting. These proteins have potential to be used as a biomarker for early pregnancy detection, however, more extensive analysis are needed to validate its possibility.展开更多
Objective:The aim of the study was to screen differentially expressed proteins between left-and right-sided colon cancers by proteomics techniques and provide molecular genetic basis for oncobiological difference betw...Objective:The aim of the study was to screen differentially expressed proteins between left-and right-sided colon cancers by proteomics techniques and provide molecular genetic basis for oncobiological difference between left-and rightsided colon cancers.Methods:Tissue samples including left-and right-sided colon cancers were collected and preserved in the-80 ℃ refrigeratory.In the first part of our experiment,protein separating was performed by using two-dimensional gel electrophoresis(2-DE) and the images of the gels were acquired by the scanner and then analyzed to find the differentially expression protein-spots in different groups.The peptide mass fingerprintings(PMF) was acquired by matrix assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF-MS) and the proteins were identified by data searching in the Mascot-database.Differentially expression proteins were assayed by RT-PCR,Western blot,and immunohistochemical methods.Results:The 55 differentially expressed protein spots were screened and 23 spots of them were identified.Compared to right-sided colon cancer,15 proteins up-regulated and 8 proteins down-regulated including HSP27 in left-sided colon cancer.HSP27 expressed higher in right-sided than in left-sided colon cancers by RT-PCR,Western blot and immunohistochemical methods.Conclusion:There were differentially expressed proteins between left-and right-sided colon cancers,especially differences in HSP27 expression in mRNA and protein level,which were molecular genetic basis for oncobiological difference between left-and right-sided colon cancers.展开更多
Mostly fed with grass in fresh or conserved form, cattle and other livestock have to cope with silicate defence bodies from plants （phytoliths） and environmental silicates （grit）, which abrade tooth enamel and cou...Mostly fed with grass in fresh or conserved form, cattle and other livestock have to cope with silicate defence bodies from plants （phytoliths） and environmental silicates （grit）, which abrade tooth enamel and could additionally interact with various salivary proteins. To detect potential candidates for silicate-binding proteins, bovine whole saliva was incubated with grass-derived phytoliths and silicates. Interactions of salivary proteins with pulverized bovine dental enamel and dentine were additionally analysed. After intense washing, the powder fractions were loaded onto 1D-polyacrylamide gels, most prominent adhesive protein bands were cut out and proteins were identified by mass spectrometry within three independent replicates. All materials were mainly botmd by bovine odorant-binding protein, bovine salivary protein 30× 10^3 and carbonic anhydrase VI. The phytolith/silicate fraction showed additional stronger interaction with haemoglobin β and lactoperoxidase. Conceivably, the binding of these proteins to the surfaces may contribute to biological processes occurring on them.展开更多
Background： The disclosure of the poplar genome strengthens its position as well-established model organism. Populus has been subject of several proteome studies, but up to date no comparative study was performed on ...Background： The disclosure of the poplar genome strengthens its position as well-established model organism. Populus has been subject of several proteome studies, but up to date no comparative study was performed on the extraction method of soluble proteins for this species. The extraction is the most critical step in two-dimensional gel electrophoresis and each extraction method has its advantages, disadvantages and limitations. Therefore protein extraction methods should be optimized for each tissue before starting an experimental setup. In prospect of future DIGE （Differential Gel electrophoresis） experiments for the investigation of the effects of cadmium and inoculation with plant growth promoting bacteria at the proteome level, the aim of this study was to optimize an extraction method for soluble proteins of poplar leaves and roots. Results： The acetone-phenol extraction method was found to be the most suited, rendering a high spot number and low background interference. During further optimization, several critical steps in the extraction method were revealed. Conclusion： Aiming to optimize the extraction of soluble leaf and root proteins of Populus deltoides × （trichocarpa× deltoides） compatible with DIGE analysis, a protocol rendering high reproducibility, low background interference and a high spot number was established, however no novel insights were acquired.展开更多
Objective： To investigate the proteornic differences between the high-sensitivity （HS） group and low-sensitivity （LS） group of cervical cancer treated by radiotherapy and confirm the radiotherapy sensitivity asso...Objective： To investigate the proteornic differences between the high-sensitivity （HS） group and low-sensitivity （LS） group of cervical cancer treated by radiotherapy and confirm the radiotherapy sensitivity associated proteins in early cervical cancer. Methods： The fresh carcinoma tissues were collected from 10 untreated cervical cancer patients and preserved in the -80 ℃ refrigeratory. The tissues were classified into two groups： high sensitivity group （HS） and low sensitivity group （LS）, according to their response to radiotherapy. In the first part of our experiment, protein separating was performed by using two-dimensional gel electrophoresis （2-DE） with Amersham 18 cm linear pH 3-10 immobilized pH gradient （IPG） strips. The images of the gels were acquired by the scanner and then analyzed by using PD-quest7.3 software to find the differentially expression protein-spots in each group. Then the differentially expressed protein-spots was incised from the gels and digested by trypsin. The peptide mass fingerprintings （PMF） was acquired by matrix assisted laser desorption/ionization time-of-flight mass spectrometry （MALDI-TOF-MS） and the proteins were identified by data searching in the Mascot-database. Part of differentially expression proteins were assayed by Western Blot. Results： Most of the gels were clear and successfully analyzed by PD-quest7.3 software. Most of the protein-spots concentrated on the area of 20-100Kda （Mw） and pH4-8. The average number of the protein-spots was 754 ± 64 in HS group and 777 ±48个 in LS group. The match rate was 87.6% between two groups. Five high expression proteins were found in HS group which were low expression in LS group, 3 high expression protein were found in LS group which were low expression in HS group. Reselts of Western Blot were in coincidence to proteomic result. Conclusion： The 2-DE gels image of HS group and LS group with early cervical cancer tissues treated by radiotherapy are successfully acquired. Some differentially expression proteins between the two groups are further confirmed by immunohistochemical assay.展开更多
Compound SLXM-2, a derivative of cyclophosphamide （CTX）, has shown potent growth-inhibitory effect on tumor cells with low toxicity in previous studies. However, the mechanism of its anti-tumor effect, especially on...Compound SLXM-2, a derivative of cyclophosphamide （CTX）, has shown potent growth-inhibitory effect on tumor cells with low toxicity in previous studies. However, the mechanism of its anti-tumor effect, especially on DNA damage, remains largely unclear. This study investigated the effect of SLXM-2 on the survival time of mice transplanted with the ascitie fluid-type hepatocarcinoma 22 （H22）. We also evaluated the correlation between DNA damaging effect of SLXM-2 and its anti-tumor effect, and to probe the possible molecular mechanism for its effect on H22 cells. The results suggested that SLXM-2 significantly （P〈0.05） prolonged the survival time of mice bearing the ascitic fluid-type H22. Furthermore, SLXM-2 induced DNA damage in a dose-dependent manner in H22 cells. Further investigation revealed that SLXM-2 significantly （P〈0.05） up-regulated the expression levels of a series of DNA damage-related proteins, such as γH2AX （Ser139）, p-Chkl （Ser296）, p-Chk2 （Thr68）, p-p53 （Ser15）, p-p53 （Ser20） and p21, and down-regulated the expression of p-ATR （Ser428） and p-ATM （Ser1981）. In conclusion, SLXM-2 showed a remarkable anti-tumor activity on ascitic fluid-type H22 cells, and its molecular mechanism is related to its DNA damaging effect.展开更多
Proteomic analysis of upland cotton was performed to profile the global detectable proteomes of ovules and fibers using two-dimensional electrophoresis(2DE).A total of 1,203 independent protein spots were collected fr...Proteomic analysis of upland cotton was performed to profile the global detectable proteomes of ovules and fibers using two-dimensional electrophoresis(2DE).A total of 1,203 independent protein spots were collected from representative 2DE gels,which were digested with trypsin and identified by matrix-assisted laser desorption and ionization-time-offlight/time-of-flight(MALDI-TOF/TOF)mass spectrometry.The mass spectrometry or tandem mass spectrometry(MS or MS/MS)data were then searched against a local database constructed from Gossypium hirsutum genome sequences,resulting in successful identification of 975 protein spots(411 for ovules and 564 for fibers).Functional annotation analysis of the 975identified proteins revealed that ovule-specific proteins were mainly enriched in functions related to fatty acid elongation,sulfur amino acid metabolism and post-replication repair,while fiber-specific proteins were enriched in functions related to root hair elongation,galactose metabolism and D-xylose metabolic processes.Further annotation analysis of the most abundant protein spots showed that 28.96%of the total proteins in the ovule were mainly located in the Golgi apparatus,endoplasmic reticulum,mitochondrion and ribosome,whereas in fibers,27.02%of the total proteins were located in the cytoskeleton,nuclear envelope and cell wall.Quantitative real-time polymerase chain reaction(q RT-PCR)analyses of the ovule-specific protein spots P61,P93 and P198 and fiber-specific protein spots 230,477 and 511 were performed to validate the proteomics data.Protein-protein interaction network analyses revealed very different network cluster patterns between ovules and fibers.This work provides the largest protein identification dataset of 2DE-detectable proteins in cotton ovules and fibers and indicates potentially important roles of tissue-specific proteins,thus providing insights into the cotton ovule and fiber proteomes on a global scale.展开更多
Objective： Recently, a high frequency of mutations in mitochondrial DNA （mtDNA） has been detected in ovarian cancer. To explore the alterations of proteins in mitochondria in ovarian cancer, a pair of human ovarian...Objective： Recently, a high frequency of mutations in mitochondrial DNA （mtDNA） has been detected in ovarian cancer. To explore the alterations of proteins in mitochondria in ovarian cancer, a pair of human ovarian carcinoma cell lines （SKOV3/SKOV3.ip1） with different metastatic potentials was examined. Methods： Cancer cells SKOV3.ipl were derived from the ascitic tumor cells of nude mice bearing a tumor of ovarian cancer cells SKOV3. SKOV3.ipl exhibited a higher degree of migration potential than its paired cell line SKOV3. The proteins in the mi- tochondria of these two cells were isolated and separated by 2-D gel electrophoresis. The differently expressed proteins were extracted and identified using matrix assisted laser desorption ionisation/time-of-flight/time-of-flight （MALDITOF/TOF）, and finally a selected protein candidate was further investigated by immunohistochemistry （IHC） method in nude mice bearing tumor tissues of these two cells. Results： A total of 35 spots with different expressions were identified between the two cells using 2D-polyacrylamide gel electrophoresis （PAGE） approach. Among them, 17 spots were detected only in either SKOV3 or SKOV3.ipl cells. Eighteen spots expressed different levels, with as much as a three-fold difference between the two cells. Twenty spots were analyzed using MALDI-TOF/TOF, and 11 of them were identified successfully; four were known to be located in mitochondria, including superoxide dismutase 2 （SOD2）, fumarate hydratase （FH）, mitochondrial ribosomal protein L38 （MRPL38）, and mRNA turnover 4 homolog （MRTO4）. An increased staining of SOD2 was observed in SKOV3.ipl over that of SKOV3 in IHC analysis. Conclusions： Our results indicate that the enhanced antioxidation and metabolic potentials of ovarian cancer cells might contribute to their aggressive and metastatic behaviors. The underlying mechanism warrants further study.展开更多
文摘以在新疆昌吉某猪场分离的一株猪肺炎支原体(CJ)为试验材料,采用SDS-PAGE和Western-Blot印迹对该菌的膜蛋白进行了分析。该猪肺炎支原体(CJ)经过Friis改良液体培养基培养,以12 000 r/min离心收集菌体,低温反复冻融获得膜蛋白后,应用12%分离胶,4%浓缩胶进行SDS-PAGE分离,在电泳图谱上呈现大概26条蛋白条带,分子量在10 k D^200 k D。再以膜蛋白做免疫印迹检测,其Western-Blot膜上有14种反应条带,其中p36、p38、p40、p52、p65,p70、p114、p200条带颜色较深。
文摘Since preparation and solubilization of protein samples are crucial factors in proteome research,the authors established a sequential extraction technique to prepare protein samples from the body wall of the 5th instar larvae of silkworm.Bombyx mori.Two kinds of protein samples were obtained from the body wall using the method.Between the two types of samples only about 15% proteins were identical;the majority were different,indicating that more species of proteins could be obtained with the sequential extraction method;which will be useful for preparation of protein samples for proteome study.
基金supported by a research project (No. 2006IK012) of the General Administration of Quality Supervision, Inspection and Quarantine of P. R. China.
文摘Sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE) was applied in the detection of the end point temperature (EPT) of thermal denatured protein in fish and meat in this study. It was also used in studying the thermal denatured temperature range of proteins in salmon and chicken meat. The results show that the temperature ranges of denatured proteins were from 65 ℃ to 75 ℃ , and these temperature ranges were influenced by the processing methods. Through SDS-PAGE, the features of repeated heating thermal denatured proteins under the same temperature and processing time were studied. The electrophoresis patterns of thermal denatured proteins determined through repeated heating at the same temperature did not exhibit any change. For the detection of cooked fish and meat samples, they were subjected to applying the SDS-PAGE method, which revealed an EPT ranging from 60 ℃ to 80 ℃ .
基金Project (No. A-007) supported by the Key Medicine Foundation of Zhejiang Province, China
文摘Idiopathic pulmonary arterial hypertension(IPAH) is a rare disease of unknown etiology.The exact pathogenesis of pulmonary arterial hypertension is still not well known.In the past decades,many protein molecules have been found to be in-volved in the development of IPAH.With proteomic techniques,profiling of human plasma proteome becomes more feasible in searching for disease-related markers.In present study,we showed the protein expression profiles of the serum of IPAH and healthy controls after depleting a few high-abundant proteins in serum.Thirteen spots had changed significantly in IPAH com-pared with healthy controls and were identified by LC-MS/MS.Alpha-1-antitrypsin and vitronectin were down-regulated in IPAH and may be valuable candidates for further explorations of their roles in the development of IPAH.
文摘Early detection of pregnancy is advantageous to breeders. However, tools that were used in the early pregnancy detection are expensive, laboratory-based and not efficient and applicable enough in the field to farmers. Thus, there is a need to find a biomarker which not only can detect pregnancy but also could be applied to the livestock on the field. Proteomic approach was used in this study as to search for early pregnancy biomarker. When goat sera at day 14 of gestation were resolved by using two dimensional gel electrophoresis, five differentially expressed proteins were detected. Four of the proteins were identified as albumin precursors, immunoglobulin lambda light chain and zinc-alpha-2-glycoprotein. Pregnancy associated glycoprotein-1 was detected by using liquid chromatography mass spectrometry and the protein was validated by immunoblotting. These proteins have potential to be used as a biomarker for early pregnancy detection, however, more extensive analysis are needed to validate its possibility.
基金Supported by a grant of Foundation Project from Bureau of Health in Hunan Province (No. C2009-010)
文摘Objective:The aim of the study was to screen differentially expressed proteins between left-and right-sided colon cancers by proteomics techniques and provide molecular genetic basis for oncobiological difference between left-and rightsided colon cancers.Methods:Tissue samples including left-and right-sided colon cancers were collected and preserved in the-80 ℃ refrigeratory.In the first part of our experiment,protein separating was performed by using two-dimensional gel electrophoresis(2-DE) and the images of the gels were acquired by the scanner and then analyzed to find the differentially expression protein-spots in different groups.The peptide mass fingerprintings(PMF) was acquired by matrix assisted laser desorption/ionization time-of-flight mass spectrometry(MALDI-TOF-MS) and the proteins were identified by data searching in the Mascot-database.Differentially expression proteins were assayed by RT-PCR,Western blot,and immunohistochemical methods.Results:The 55 differentially expressed protein spots were screened and 23 spots of them were identified.Compared to right-sided colon cancer,15 proteins up-regulated and 8 proteins down-regulated including HSP27 in left-sided colon cancer.HSP27 expressed higher in right-sided than in left-sided colon cancers by RT-PCR,Western blot and immunohistochemical methods.Conclusion:There were differentially expressed proteins between left-and right-sided colon cancers,especially differences in HSP27 expression in mRNA and protein level,which were molecular genetic basis for oncobiological difference between left-and right-sided colon cancers.
基金supported by the German Research Foundation (DFG, SU 124/15-1)
文摘Mostly fed with grass in fresh or conserved form, cattle and other livestock have to cope with silicate defence bodies from plants （phytoliths） and environmental silicates （grit）, which abrade tooth enamel and could additionally interact with various salivary proteins. To detect potential candidates for silicate-binding proteins, bovine whole saliva was incubated with grass-derived phytoliths and silicates. Interactions of salivary proteins with pulverized bovine dental enamel and dentine were additionally analysed. After intense washing, the powder fractions were loaded onto 1D-polyacrylamide gels, most prominent adhesive protein bands were cut out and proteins were identified by mass spectrometry within three independent replicates. All materials were mainly botmd by bovine odorant-binding protein, bovine salivary protein 30× 10^3 and carbonic anhydrase VI. The phytolith/silicate fraction showed additional stronger interaction with haemoglobin β and lactoperoxidase. Conceivably, the binding of these proteins to the surfaces may contribute to biological processes occurring on them.
文摘Background： The disclosure of the poplar genome strengthens its position as well-established model organism. Populus has been subject of several proteome studies, but up to date no comparative study was performed on the extraction method of soluble proteins for this species. The extraction is the most critical step in two-dimensional gel electrophoresis and each extraction method has its advantages, disadvantages and limitations. Therefore protein extraction methods should be optimized for each tissue before starting an experimental setup. In prospect of future DIGE （Differential Gel electrophoresis） experiments for the investigation of the effects of cadmium and inoculation with plant growth promoting bacteria at the proteome level, the aim of this study was to optimize an extraction method for soluble proteins of poplar leaves and roots. Results： The acetone-phenol extraction method was found to be the most suited, rendering a high spot number and low background interference. During further optimization, several critical steps in the extraction method were revealed. Conclusion： Aiming to optimize the extraction of soluble leaf and root proteins of Populus deltoides × （trichocarpa× deltoides） compatible with DIGE analysis, a protocol rendering high reproducibility, low background interference and a high spot number was established, however no novel insights were acquired.
基金Supported by grants from the Hunan Natural Science foundation (No.06JJ4199)the Hunan Science Technology Foundation (No.2007SK3010)
文摘Objective： To investigate the proteornic differences between the high-sensitivity （HS） group and low-sensitivity （LS） group of cervical cancer treated by radiotherapy and confirm the radiotherapy sensitivity associated proteins in early cervical cancer. Methods： The fresh carcinoma tissues were collected from 10 untreated cervical cancer patients and preserved in the -80 ℃ refrigeratory. The tissues were classified into two groups： high sensitivity group （HS） and low sensitivity group （LS）, according to their response to radiotherapy. In the first part of our experiment, protein separating was performed by using two-dimensional gel electrophoresis （2-DE） with Amersham 18 cm linear pH 3-10 immobilized pH gradient （IPG） strips. The images of the gels were acquired by the scanner and then analyzed by using PD-quest7.3 software to find the differentially expression protein-spots in each group. Then the differentially expressed protein-spots was incised from the gels and digested by trypsin. The peptide mass fingerprintings （PMF） was acquired by matrix assisted laser desorption/ionization time-of-flight mass spectrometry （MALDI-TOF-MS） and the proteins were identified by data searching in the Mascot-database. Part of differentially expression proteins were assayed by Western Blot. Results： Most of the gels were clear and successfully analyzed by PD-quest7.3 software. Most of the protein-spots concentrated on the area of 20-100Kda （Mw） and pH4-8. The average number of the protein-spots was 754 ± 64 in HS group and 777 ±48个 in LS group. The match rate was 87.6% between two groups. Five high expression proteins were found in HS group which were low expression in LS group, 3 high expression protein were found in LS group which were low expression in HS group. Reselts of Western Blot were in coincidence to proteomic result. Conclusion： The 2-DE gels image of HS group and LS group with early cervical cancer tissues treated by radiotherapy are successfully acquired. Some differentially expression proteins between the two groups are further confirmed by immunohistochemical assay.
基金Eleventh Five Year Plan for National Science and Technology Major Project(Grant No.2009ZX0930010)
文摘Compound SLXM-2, a derivative of cyclophosphamide （CTX）, has shown potent growth-inhibitory effect on tumor cells with low toxicity in previous studies. However, the mechanism of its anti-tumor effect, especially on DNA damage, remains largely unclear. This study investigated the effect of SLXM-2 on the survival time of mice transplanted with the ascitie fluid-type hepatocarcinoma 22 （H22）. We also evaluated the correlation between DNA damaging effect of SLXM-2 and its anti-tumor effect, and to probe the possible molecular mechanism for its effect on H22 cells. The results suggested that SLXM-2 significantly （P〈0.05） prolonged the survival time of mice bearing the ascitic fluid-type H22. Furthermore, SLXM-2 induced DNA damage in a dose-dependent manner in H22 cells. Further investigation revealed that SLXM-2 significantly （P〈0.05） up-regulated the expression levels of a series of DNA damage-related proteins, such as γH2AX （Ser139）, p-Chkl （Ser296）, p-Chk2 （Thr68）, p-p53 （Ser15）, p-p53 （Ser20） and p21, and down-regulated the expression of p-ATR （Ser428） and p-ATM （Ser1981）. In conclusion, SLXM-2 showed a remarkable anti-tumor activity on ascitic fluid-type H22 cells, and its molecular mechanism is related to its DNA damaging effect.
基金the Special Fund for Agro-scientific Research in the Public Interest of the People’s Republic of China (201403075)Major Technology Project of Hainan (ZDZX2013010-1)+1 种基金Program for Top Young Talents in the Chinese Academy of Tropical Agricultural Sciences (ITBB130102)China Postdoctoral Science Foundation (20110490003)
文摘Proteomic analysis of upland cotton was performed to profile the global detectable proteomes of ovules and fibers using two-dimensional electrophoresis(2DE).A total of 1,203 independent protein spots were collected from representative 2DE gels,which were digested with trypsin and identified by matrix-assisted laser desorption and ionization-time-offlight/time-of-flight(MALDI-TOF/TOF)mass spectrometry.The mass spectrometry or tandem mass spectrometry(MS or MS/MS)data were then searched against a local database constructed from Gossypium hirsutum genome sequences,resulting in successful identification of 975 protein spots(411 for ovules and 564 for fibers).Functional annotation analysis of the 975identified proteins revealed that ovule-specific proteins were mainly enriched in functions related to fatty acid elongation,sulfur amino acid metabolism and post-replication repair,while fiber-specific proteins were enriched in functions related to root hair elongation,galactose metabolism and D-xylose metabolic processes.Further annotation analysis of the most abundant protein spots showed that 28.96%of the total proteins in the ovule were mainly located in the Golgi apparatus,endoplasmic reticulum,mitochondrion and ribosome,whereas in fibers,27.02%of the total proteins were located in the cytoskeleton,nuclear envelope and cell wall.Quantitative real-time polymerase chain reaction(q RT-PCR)analyses of the ovule-specific protein spots P61,P93 and P198 and fiber-specific protein spots 230,477 and 511 were performed to validate the proteomics data.Protein-protein interaction network analyses revealed very different network cluster patterns between ovules and fibers.This work provides the largest protein identification dataset of 2DE-detectable proteins in cotton ovules and fibers and indicates potentially important roles of tissue-specific proteins,thus providing insights into the cotton ovule and fiber proteomes on a global scale.
文摘Objective： Recently, a high frequency of mutations in mitochondrial DNA （mtDNA） has been detected in ovarian cancer. To explore the alterations of proteins in mitochondria in ovarian cancer, a pair of human ovarian carcinoma cell lines （SKOV3/SKOV3.ip1） with different metastatic potentials was examined. Methods： Cancer cells SKOV3.ipl were derived from the ascitic tumor cells of nude mice bearing a tumor of ovarian cancer cells SKOV3. SKOV3.ipl exhibited a higher degree of migration potential than its paired cell line SKOV3. The proteins in the mi- tochondria of these two cells were isolated and separated by 2-D gel electrophoresis. The differently expressed proteins were extracted and identified using matrix assisted laser desorption ionisation/time-of-flight/time-of-flight （MALDITOF/TOF）, and finally a selected protein candidate was further investigated by immunohistochemistry （IHC） method in nude mice bearing tumor tissues of these two cells. Results： A total of 35 spots with different expressions were identified between the two cells using 2D-polyacrylamide gel electrophoresis （PAGE） approach. Among them, 17 spots were detected only in either SKOV3 or SKOV3.ipl cells. Eighteen spots expressed different levels, with as much as a three-fold difference between the two cells. Twenty spots were analyzed using MALDI-TOF/TOF, and 11 of them were identified successfully; four were known to be located in mitochondria, including superoxide dismutase 2 （SOD2）, fumarate hydratase （FH）, mitochondrial ribosomal protein L38 （MRPL38）, and mRNA turnover 4 homolog （MRTO4）. An increased staining of SOD2 was observed in SKOV3.ipl over that of SKOV3 in IHC analysis. Conclusions： Our results indicate that the enhanced antioxidation and metabolic potentials of ovarian cancer cells might contribute to their aggressive and metastatic behaviors. The underlying mechanism warrants further study.
