目的:筛选人重组S100A8蛋白特异性的适配体。方法:采用配体指数级富集系统进化(systematic evolu-tion of ligand by exponential enrichment,SELEX)技术,以人重组S100A8蛋白为靶蛋白,以微孔板为筛选介质,从体外合成的随机单链DNA文库...目的:筛选人重组S100A8蛋白特异性的适配体。方法:采用配体指数级富集系统进化(systematic evolu-tion of ligand by exponential enrichment,SELEX)技术,以人重组S100A8蛋白为靶蛋白,以微孔板为筛选介质,从体外合成的随机单链DNA文库中筛选其适配体。利用生物素-链霉亲和素-辣根过氧化物酶系统检测每轮单链DNA文库与靶蛋白的亲和力直至亲和力的上升趋于饱和,将最后一轮筛选产物克隆测序,并进行生物信息学分析。结果:经过11轮筛选,单链DNA文库与人重组S100A8蛋白的亲和力趋向稳定,将第11轮筛选产物克隆测序,对获得的30个适配体进行分析。一级结构分析显示30个适配体并无共同的保守序列,但有3对适配体序列完全一致。二级结构预测分析表明,茎环和口袋结构为主要的结构形式,提示其可能是适配体与人重组S100A8蛋白特异性结合的基础。根据茎环和口袋的相对比例可将30个适配体分为4组,其中第1组中35号适配体与人重组S100A8蛋白亲和力最高。结论:获得了与人重组S100A8蛋白特异性结合的适配体群,为后续适配体的应用研究以及S100A8蛋白功能的研究奠定了基础。展开更多
Stone formation is induced by an increased level of urine crystallization promoters and reduced levels of its inhibitors.Crystallization inhibitors include citrate,magnesium,zinc,and organic compounds such as glycosam...Stone formation is induced by an increased level of urine crystallization promoters and reduced levels of its inhibitors.Crystallization inhibitors include citrate,magnesium,zinc,and organic compounds such as glycosaminoglycans.In the urine,there are various proteins,such as uromodulin(Tamm-Horsfall protein),calgranulin,osteopontin,bikunin,and nephrocalcin,that are present in the stone matrix.The presence of several carboxyl groups in these macromolecules reduces calcium oxalate monohydrate crystal adhesion to the urinary epithelium and could potentially protect against lithiasis.Proteins are the most abundant component of kidney stone matrix,and their presence may reflect the process of stone formation.Many recent studies have explored the proteomics of urinary stones.Among the stone matrix proteins,the most frequently identified were uromodulin,S100 proteins(calgranulins A and B),osteopontin,and several other proteins typically engaged in inflammation and immune response.The normal level and structure of these macromolecules may constitute protection against calcium salt formation.Paradoxically,most of them may act as both promoters and inhibitors depending on circumstances.Many of these proteins have other functions in modulating oxidative stress,immune function,and inflammation that could also influence stone formation.Yet,the role of these kidney stone matrix proteins needs to be established through more studies comparing urinary stone proteomics between stone formers and non-stone formers.展开更多
OBJECTIVE: To study the effects of a decoction of Fuzhengzengxiao formula on lung adenocarcinoma regarding the inflammatory protein S100A9 known to enhance cancer cell sensitivity.METHODS: A nude mouse model of human ...OBJECTIVE: To study the effects of a decoction of Fuzhengzengxiao formula on lung adenocarcinoma regarding the inflammatory protein S100A9 known to enhance cancer cell sensitivity.METHODS: A nude mouse model of human lung adenocarcinoma was established. The mice were randomly divided into four groups using the random number table method: Group Ⅰ, control;Group Ⅱ, treatment with a decoction of the Fuzhengzengxiao formula alone; Group Ⅲ, treatment with radiotherapy alone; and Group Ⅳ, treatment with radiotherapy plus a decoction of Fuzhengzengxiao formula. When the tumor body was 1 cm^3 in diameter, the tumor bearing mice in GroupsⅢ and Ⅳ were irradiated at a single dose of 10 Gy and the tumor inhibition rate was evaluated.The expression of S100A9 was determined using Western blotting and q-PCR(Real-time Quantitative PCR Detecting System). The sensitivity of cells containing RNAi S100A9 to radiotherapy was evaluated using the Click multiple target model,and the cell cycle was analyzed using flow cytometry.RESULTS: Relative to the control group,the expression of S100A9 in the tumors in each treatment group was decreased,especially in Group Ⅳ. The sensitizing enhancement ratio(SER) Dq was >1 after RNAi S100A9; it decreased the surviving fraction after a 2 Gy dose exposure,and also the D_0 and Dq of the tumor cells; in addition, the radiosensitivity of G_2/M cells was significantly increased.CONCLUSION: The decoction of the Fuzhengzengxiao formula downregulated the expression of S100A9 in lung adenocarcinoma cells.展开更多
文摘目的:筛选人重组S100A8蛋白特异性的适配体。方法:采用配体指数级富集系统进化(systematic evolu-tion of ligand by exponential enrichment,SELEX)技术,以人重组S100A8蛋白为靶蛋白,以微孔板为筛选介质,从体外合成的随机单链DNA文库中筛选其适配体。利用生物素-链霉亲和素-辣根过氧化物酶系统检测每轮单链DNA文库与靶蛋白的亲和力直至亲和力的上升趋于饱和,将最后一轮筛选产物克隆测序,并进行生物信息学分析。结果:经过11轮筛选,单链DNA文库与人重组S100A8蛋白的亲和力趋向稳定,将第11轮筛选产物克隆测序,对获得的30个适配体进行分析。一级结构分析显示30个适配体并无共同的保守序列,但有3对适配体序列完全一致。二级结构预测分析表明,茎环和口袋结构为主要的结构形式,提示其可能是适配体与人重组S100A8蛋白特异性结合的基础。根据茎环和口袋的相对比例可将30个适配体分为4组,其中第1组中35号适配体与人重组S100A8蛋白亲和力最高。结论:获得了与人重组S100A8蛋白特异性结合的适配体群,为后续适配体的应用研究以及S100A8蛋白功能的研究奠定了基础。
文摘Stone formation is induced by an increased level of urine crystallization promoters and reduced levels of its inhibitors.Crystallization inhibitors include citrate,magnesium,zinc,and organic compounds such as glycosaminoglycans.In the urine,there are various proteins,such as uromodulin(Tamm-Horsfall protein),calgranulin,osteopontin,bikunin,and nephrocalcin,that are present in the stone matrix.The presence of several carboxyl groups in these macromolecules reduces calcium oxalate monohydrate crystal adhesion to the urinary epithelium and could potentially protect against lithiasis.Proteins are the most abundant component of kidney stone matrix,and their presence may reflect the process of stone formation.Many recent studies have explored the proteomics of urinary stones.Among the stone matrix proteins,the most frequently identified were uromodulin,S100 proteins(calgranulins A and B),osteopontin,and several other proteins typically engaged in inflammation and immune response.The normal level and structure of these macromolecules may constitute protection against calcium salt formation.Paradoxically,most of them may act as both promoters and inhibitors depending on circumstances.Many of these proteins have other functions in modulating oxidative stress,immune function,and inflammation that could also influence stone formation.Yet,the role of these kidney stone matrix proteins needs to be established through more studies comparing urinary stone proteomics between stone formers and non-stone formers.
基金Supported by a Grant from the National Natural Science Foundation of China:the Effect of TCM Combined Radiotherapy and RNAi Suppression on the Protein Expression Changes of S100A9 & Cyclophilin A and Radiosensitivity in Lung Adenocarcinoma(No.81072925)
文摘OBJECTIVE: To study the effects of a decoction of Fuzhengzengxiao formula on lung adenocarcinoma regarding the inflammatory protein S100A9 known to enhance cancer cell sensitivity.METHODS: A nude mouse model of human lung adenocarcinoma was established. The mice were randomly divided into four groups using the random number table method: Group Ⅰ, control;Group Ⅱ, treatment with a decoction of the Fuzhengzengxiao formula alone; Group Ⅲ, treatment with radiotherapy alone; and Group Ⅳ, treatment with radiotherapy plus a decoction of Fuzhengzengxiao formula. When the tumor body was 1 cm^3 in diameter, the tumor bearing mice in GroupsⅢ and Ⅳ were irradiated at a single dose of 10 Gy and the tumor inhibition rate was evaluated.The expression of S100A9 was determined using Western blotting and q-PCR(Real-time Quantitative PCR Detecting System). The sensitivity of cells containing RNAi S100A9 to radiotherapy was evaluated using the Click multiple target model,and the cell cycle was analyzed using flow cytometry.RESULTS: Relative to the control group,the expression of S100A9 in the tumors in each treatment group was decreased,especially in Group Ⅳ. The sensitizing enhancement ratio(SER) Dq was >1 after RNAi S100A9; it decreased the surviving fraction after a 2 Gy dose exposure,and also the D_0 and Dq of the tumor cells; in addition, the radiosensitivity of G_2/M cells was significantly increased.CONCLUSION: The decoction of the Fuzhengzengxiao formula downregulated the expression of S100A9 in lung adenocarcinoma cells.
