Objective To identify the sperm membrane proteins that are associated with antisperm antibody Methods Using antisperm antibody positive serum through unidimensional polyacrylamide gel electrophoresis and 2-dimensi...Objective To identify the sperm membrane proteins that are associated with antisperm antibody Methods Using antisperm antibody positive serum through unidimensional polyacrylamide gel electrophoresis and 2-dimensional gel electrophoresis followed by Western blot analysis to determine the molecular weights (MW) and isoelectric points (pI) of sperm membrane proteins that are associated with antisperm antibody. Results Eight kinds of MW with more than ten sperm membrane proteins can be recognized by antisperm antibody positive serum, of which the MWs and pI were 23 kD, 31 kD, 32 kD, 34 kD, 41 kD, 51 kD, 60 kD, 78 kD and 5.3, 5.5,5.7, 5.0, 5.3, 5.8, 6.0, 5.5～6.2, 4.6,5.1,5.5～5.8 respectively. The identification ratios of the sperm membrane proteins on 78 kD (60.7%), 60 kD (71.4%), 51 kD (14.9%) and 23 kD (14.29%) were higher. Conclusion The sperm membrane proteins with MW of 78 kD, 60 kD, 51 kD and 23 kD were associated with antisperm antibody and immunological infertility. Two- dimensional gel electrophoresis and Western blotting can precisely identify the sperm membrane proteins that are associated with antisperm antibody.展开更多
Globozoospermia is a severe form of teratozoospermia characterized by round-headed spermatozoa with an absent acrosome, an aberrant nuclear membrane and midpiece defects. Globozoospermia is diagnosed by the presence o...Globozoospermia is a severe form of teratozoospermia characterized by round-headed spermatozoa with an absent acrosome, an aberrant nuclear membrane and midpiece defects. Globozoospermia is diagnosed by the presence of 100% round-headed spermatozoa on semen analysis, and patients with this condition are absolutely infertile. The objective of this study was to investigate the differences in protein expression between human round- headed and normal spermatozoa. Two-dimensional （2-D） fluorescence difference gel electrophoresis （DIGE） coupled with mass spectrometry （MS） was used in this study. Over 61 protein spots were analysed in each paired normal/round-headed comparison, using DIGE technology along with an internal standard. In total, 35 protein spots identified by tandem mass spectrometry （MS/MS） exhibited significant changes （paired t-test, P 〈 0.05） in the expression level between normal and round-headed spermatozoa. A total of nine proteins were found to be upregulated and 26 proteins were found to be downregulated in round-headed spermatozoa compared with normal spermatozoa. The differentially expressed proteins that we identified may have important roles in a variety of cellular processes and structures, including spermatogenesis, cell skeleton, metabolism and spermatozoa motility.展开更多
In this paper,a novel method to automatically detect protein spots on a two-dimensional (2-D) electrophoresis gel image is proposed to implement proteomics analysis of complex analyte.On the basis of the identifying s...In this paper,a novel method to automatically detect protein spots on a two-dimensional (2-D) electrophoresis gel image is proposed to implement proteomics analysis of complex analyte.On the basis of the identifying spots results based on color variation and spot size features,morphological feature is introduced as a new criterion to distinguish protein spots from non-protein spots.Image-sharpening,edge-detecting and morphological feature extraction methods were consequently combined to detect protein spots on a 2-D electrophoresis gel image subject to strong disturbance.The proposed method was applied to detect the protein spots of proteomic gel images from E.coli cell,human kidney tissue and human serum.The results demonstrated that this method is more accurate and reliable than previous methods such as PDQuest 7.2 and ImageMaster 5.0 software for detecting protein spots on gel images with strong interferences.展开更多
目的探讨2型糖尿病发生、发展相关的蛋白质,及其胰岛素信号传递系统组成的差异。方法将糖尿病地鼠作为实验组,昆明种封闭群正常小白鼠作为对照组,在注射胰岛素0、5、30、60及120 m in五个不同时间点分别测定每只小鼠的对应时间的血糖值...目的探讨2型糖尿病发生、发展相关的蛋白质,及其胰岛素信号传递系统组成的差异。方法将糖尿病地鼠作为实验组,昆明种封闭群正常小白鼠作为对照组,在注射胰岛素0、5、30、60及120 m in五个不同时间点分别测定每只小鼠的对应时间的血糖值。采用蛋白质组学技术筛选差异蛋白。结果不同时点血糖值实验组显著高于对照组(P<0.01),且实验组血糖下降速度缓慢;胰岛素作用后,糖尿病小鼠肝组织蛋白质发生明显改变,主要表现为2-DE图谱中蛋白质斑点的增减以及颜色的深浅上,初步筛选出6个与胰岛素细胞信号转导相关的蛋白质,包括RAF1、BTF3、SEC1、PAR2、MK10及TRAF2。结论2型糖尿病地鼠存在胰岛素抵抗,与胰岛素细胞信号转导相关的差异蛋白可能是糖尿病病理生理机制的分子基础。展开更多
文摘Objective To identify the sperm membrane proteins that are associated with antisperm antibody Methods Using antisperm antibody positive serum through unidimensional polyacrylamide gel electrophoresis and 2-dimensional gel electrophoresis followed by Western blot analysis to determine the molecular weights (MW) and isoelectric points (pI) of sperm membrane proteins that are associated with antisperm antibody. Results Eight kinds of MW with more than ten sperm membrane proteins can be recognized by antisperm antibody positive serum, of which the MWs and pI were 23 kD, 31 kD, 32 kD, 34 kD, 41 kD, 51 kD, 60 kD, 78 kD and 5.3, 5.5,5.7, 5.0, 5.3, 5.8, 6.0, 5.5～6.2, 4.6,5.1,5.5～5.8 respectively. The identification ratios of the sperm membrane proteins on 78 kD (60.7%), 60 kD (71.4%), 51 kD (14.9%) and 23 kD (14.29%) were higher. Conclusion The sperm membrane proteins with MW of 78 kD, 60 kD, 51 kD and 23 kD were associated with antisperm antibody and immunological infertility. Two- dimensional gel electrophoresis and Western blotting can precisely identify the sperm membrane proteins that are associated with antisperm antibody.
基金Acknowledgment We thank Beijing Proteome Research Center, （Beijing, China） for its enthusiastic technological support and for the theory of 2-D DIGE. We also thank（Changsha, China） College of Life Sciences at Hunan Normal University for supporting the MS technology. Finally, we are very grateful to our collaborators for their help, as well as their valuable discussions and suggestions during the course of this work. This work was supported by two grants from the National Natural Science Foundation of China （NO. 30170480 and NO. 30470884）.
文摘Globozoospermia is a severe form of teratozoospermia characterized by round-headed spermatozoa with an absent acrosome, an aberrant nuclear membrane and midpiece defects. Globozoospermia is diagnosed by the presence of 100% round-headed spermatozoa on semen analysis, and patients with this condition are absolutely infertile. The objective of this study was to investigate the differences in protein expression between human round- headed and normal spermatozoa. Two-dimensional （2-D） fluorescence difference gel electrophoresis （DIGE） coupled with mass spectrometry （MS） was used in this study. Over 61 protein spots were analysed in each paired normal/round-headed comparison, using DIGE technology along with an internal standard. In total, 35 protein spots identified by tandem mass spectrometry （MS/MS） exhibited significant changes （paired t-test, P 〈 0.05） in the expression level between normal and round-headed spermatozoa. A total of nine proteins were found to be upregulated and 26 proteins were found to be downregulated in round-headed spermatozoa compared with normal spermatozoa. The differentially expressed proteins that we identified may have important roles in a variety of cellular processes and structures, including spermatogenesis, cell skeleton, metabolism and spermatozoa motility.
基金the National Natural Science Foundat ion of China(Grant No.90209005) Science and Technology Project of Zhejiang Province(Grant No.2004C33026).
文摘In this paper,a novel method to automatically detect protein spots on a two-dimensional (2-D) electrophoresis gel image is proposed to implement proteomics analysis of complex analyte.On the basis of the identifying spots results based on color variation and spot size features,morphological feature is introduced as a new criterion to distinguish protein spots from non-protein spots.Image-sharpening,edge-detecting and morphological feature extraction methods were consequently combined to detect protein spots on a 2-D electrophoresis gel image subject to strong disturbance.The proposed method was applied to detect the protein spots of proteomic gel images from E.coli cell,human kidney tissue and human serum.The results demonstrated that this method is more accurate and reliable than previous methods such as PDQuest 7.2 and ImageMaster 5.0 software for detecting protein spots on gel images with strong interferences.
文摘目的探讨2型糖尿病发生、发展相关的蛋白质,及其胰岛素信号传递系统组成的差异。方法将糖尿病地鼠作为实验组,昆明种封闭群正常小白鼠作为对照组,在注射胰岛素0、5、30、60及120 m in五个不同时间点分别测定每只小鼠的对应时间的血糖值。采用蛋白质组学技术筛选差异蛋白。结果不同时点血糖值实验组显著高于对照组(P<0.01),且实验组血糖下降速度缓慢;胰岛素作用后,糖尿病小鼠肝组织蛋白质发生明显改变,主要表现为2-DE图谱中蛋白质斑点的增减以及颜色的深浅上,初步筛选出6个与胰岛素细胞信号转导相关的蛋白质,包括RAF1、BTF3、SEC1、PAR2、MK10及TRAF2。结论2型糖尿病地鼠存在胰岛素抵抗,与胰岛素细胞信号转导相关的差异蛋白可能是糖尿病病理生理机制的分子基础。
