CD-1, a genetically-engineered CHO cell line, was cultivated with a Biosilon microcarrier culture system.We successfully cultivated CD-1 cells to a very high density (over1×107cells/ml). Prourokinase was stably s...CD-1, a genetically-engineered CHO cell line, was cultivated with a Biosilon microcarrier culture system.We successfully cultivated CD-1 cells to a very high density (over1×107cells/ml). Prourokinase was stably secreted at about 180 IU/ 1e6 cells/24 h. Experiments showed that CD-1 cells growing on Biosilon microcarriers were able to spontaneously release from the microcarriers, then reatthch and proliferate on fresh microcarriers. This makes it very easy to scale up preduction. The microcarriers could be reused several times without affecting adhesion. proliferation and prourokinase secretion. With CMPECC membrane radial flow chromatography and MPG chromatography, the prourokinase in conditioned medium could be purified to a specific activity of 1×105 IU/mg of protein. The purification factor was about 600 fold, and approxiamately 90 % of the biological activity was recovered.展开更多
There are rising concerns about the hazardous effects of heavy metals on the environment. In this study, comet assay and DNA alkaline unwinding assay were conducted on the tissues (gills, hepatopancreas, and hemocytes...There are rising concerns about the hazardous effects of heavy metals on the environment. In this study, comet assay and DNA alkaline unwinding assay were conducted on the tissues (gills, hepatopancreas, and hemocytes) of Charybdis japonica in order to illustrate genotoxicity of three heavy metal ions (Cu2+, Pb2+, and Cd2+) on the marine crabs C. japonica. The crabs were exposed to Cu2+ (10, 50, and 100 ?g.L?1), Pb2+ (50, 250, and 500 ?g L?1) and Cd2+ (5, 25, and 50 ?g L?1), and the tissues were sampled at days 0.5, 1, 3, 6, 9, and 15. DNA alkaline unwinding assay was used for testing the DNA single strand break in gills and hepatopancreas and comet assay was employed for testing the DNA damage in hemocytes. The results showed that the DNA damage (F-value) of gills in the crabs exposed to the three heavy metals was decreased gradually during the exposure periods and there was a dose-time response relationship in certain time, suggesting that the levels of DNA single strand break in all the experimental groups increased significantly compared to the controls. Changes of F-value in hepatopancreas of the crabs exposed to the three heavy metals were similar to those in gills except that the peak values were found in the 500 ?g L?1 Pb2+ treatment group at day 3 and the 50 ?g L?1 Cd2+ treatment group at day 9. The ranks of DNA damage in gills and hepatopancreas induced by the three heavy metal ions (50 ?g L?1, day 15) were Cd2+ >Pb2+ >Cu2+ and Pb2+ >Cu2+ >Cd2+. The levels of DNA damage in gills were higher than those in hepatopancreas in the same experimental group. It can be concluded that indices of DNA damage can be used as the potential biomarkers of heavy metal pollution in marine environment.展开更多
TGF-β and Foxp3 expressions are crucial for the induction and functional activity of CD4＋Foxp3＋ regulatory T （iTreg） cells. Here, we demonstrate that although TGF-β-primed CD8＋ cells display much lower Foxp3 ex...TGF-β and Foxp3 expressions are crucial for the induction and functional activity of CD4＋Foxp3＋ regulatory T （iTreg） cells. Here, we demonstrate that although TGF-β-primed CD8＋ cells display much lower Foxp3 expression, their suppressive capacity is equivalent to that of CD4＋ iTreg cells, and both Foxp3- and Foxp3＋ CD8＋ subsets have suppressive activities in vitro and in vivo. CD8＋Foxp3- iTreg cells produce little IFN-γ but almost no IL-2, and display a typical anergic phenotype. Among phenotypic markers expressed in CD8＋Foxp3- cells, we identify CD103 expression particularly crucial for the generation and function of this subset. Moreover, IL-IO and TGF-β signals rather than cytotoxicity mediate the suppressive effect of this novel Treg population. Therefore, TGF-β can induce both CD8＋Foxp3- and CD8＋Foxp3＋ iTreg subsets, which may represent the unique immunoregulatory means to treat autoimmune and inflammatory diseases.展开更多
To investigate the expression of cluster of differentiation 44 variant 6 (CD 44V6 ) and proliferating cell nuclear antigen (PCNA) in ocular squamous cell carcinomas Methods Streptavidin biotin complex (SABC) i...To investigate the expression of cluster of differentiation 44 variant 6 (CD 44V6 ) and proliferating cell nuclear antigen (PCNA) in ocular squamous cell carcinomas Methods Streptavidin biotin complex (SABC) immunohistochemistry was used to explore the expression of CD 44V6 and PCNA in 35 cases of ocular squamous cell carcinomas, 20 cases of papillomas, and 11 cases of normal eyelid tissue Results The CD 44V6 positive rate was 62 9% (22/35) in ocular squamous cell carcinomas, 15 0% (3/20) in papillomas, but not detectable in the 11 cases of normal eyelid tissue The positive expression rates of CD 44V6 in ocular squamous cell carcinomas were significantly higher than in benign tumors (χ 2=11 57, P 【0 01)or control tissue ( P =0 001), and the positive expression rates of CD 44V6 in metastasis were significantly higher than without metastasis ( P =0 049) PCNA labeling indexes (PI) in tumors with CD 44V6 expression were significantly higher than those without ( t =20 21, P 【0 01) Conclusions Overexpression of CD 44V6 is correlated with the progress and metastasis of ocular squamous cell carcinomas CD 44V6 protein positive staining is associated with high PI CD 44V6 and PCNA are useful for evaluating prognosis展开更多
文摘CD-1, a genetically-engineered CHO cell line, was cultivated with a Biosilon microcarrier culture system.We successfully cultivated CD-1 cells to a very high density (over1×107cells/ml). Prourokinase was stably secreted at about 180 IU/ 1e6 cells/24 h. Experiments showed that CD-1 cells growing on Biosilon microcarriers were able to spontaneously release from the microcarriers, then reatthch and proliferate on fresh microcarriers. This makes it very easy to scale up preduction. The microcarriers could be reused several times without affecting adhesion. proliferation and prourokinase secretion. With CMPECC membrane radial flow chromatography and MPG chromatography, the prourokinase in conditioned medium could be purified to a specific activity of 1×105 IU/mg of protein. The purification factor was about 600 fold, and approxiamately 90 % of the biological activity was recovered.
基金supported by the Science Foundation of the Key Laboratory of Mariculture, Ministry of Education, Ocean University of China
文摘There are rising concerns about the hazardous effects of heavy metals on the environment. In this study, comet assay and DNA alkaline unwinding assay were conducted on the tissues (gills, hepatopancreas, and hemocytes) of Charybdis japonica in order to illustrate genotoxicity of three heavy metal ions (Cu2+, Pb2+, and Cd2+) on the marine crabs C. japonica. The crabs were exposed to Cu2+ (10, 50, and 100 ?g.L?1), Pb2+ (50, 250, and 500 ?g L?1) and Cd2+ (5, 25, and 50 ?g L?1), and the tissues were sampled at days 0.5, 1, 3, 6, 9, and 15. DNA alkaline unwinding assay was used for testing the DNA single strand break in gills and hepatopancreas and comet assay was employed for testing the DNA damage in hemocytes. The results showed that the DNA damage (F-value) of gills in the crabs exposed to the three heavy metals was decreased gradually during the exposure periods and there was a dose-time response relationship in certain time, suggesting that the levels of DNA single strand break in all the experimental groups increased significantly compared to the controls. Changes of F-value in hepatopancreas of the crabs exposed to the three heavy metals were similar to those in gills except that the peak values were found in the 500 ?g L?1 Pb2+ treatment group at day 3 and the 50 ?g L?1 Cd2+ treatment group at day 9. The ranks of DNA damage in gills and hepatopancreas induced by the three heavy metal ions (50 ?g L?1, day 15) were Cd2+ >Pb2+ >Cu2+ and Pb2+ >Cu2+ >Cd2+. The levels of DNA damage in gills were higher than those in hepatopancreas in the same experimental group. It can be concluded that indices of DNA damage can be used as the potential biomarkers of heavy metal pollution in marine environment.
文摘TGF-β and Foxp3 expressions are crucial for the induction and functional activity of CD4＋Foxp3＋ regulatory T （iTreg） cells. Here, we demonstrate that although TGF-β-primed CD8＋ cells display much lower Foxp3 expression, their suppressive capacity is equivalent to that of CD4＋ iTreg cells, and both Foxp3- and Foxp3＋ CD8＋ subsets have suppressive activities in vitro and in vivo. CD8＋Foxp3- iTreg cells produce little IFN-γ but almost no IL-2, and display a typical anergic phenotype. Among phenotypic markers expressed in CD8＋Foxp3- cells, we identify CD103 expression particularly crucial for the generation and function of this subset. Moreover, IL-IO and TGF-β signals rather than cytotoxicity mediate the suppressive effect of this novel Treg population. Therefore, TGF-β can induce both CD8＋Foxp3- and CD8＋Foxp3＋ iTreg subsets, which may represent the unique immunoregulatory means to treat autoimmune and inflammatory diseases.
文摘To investigate the expression of cluster of differentiation 44 variant 6 (CD 44V6 ) and proliferating cell nuclear antigen (PCNA) in ocular squamous cell carcinomas Methods Streptavidin biotin complex (SABC) immunohistochemistry was used to explore the expression of CD 44V6 and PCNA in 35 cases of ocular squamous cell carcinomas, 20 cases of papillomas, and 11 cases of normal eyelid tissue Results The CD 44V6 positive rate was 62 9% (22/35) in ocular squamous cell carcinomas, 15 0% (3/20) in papillomas, but not detectable in the 11 cases of normal eyelid tissue The positive expression rates of CD 44V6 in ocular squamous cell carcinomas were significantly higher than in benign tumors (χ 2=11 57, P 【0 01)or control tissue ( P =0 001), and the positive expression rates of CD 44V6 in metastasis were significantly higher than without metastasis ( P =0 049) PCNA labeling indexes (PI) in tumors with CD 44V6 expression were significantly higher than those without ( t =20 21, P 【0 01) Conclusions Overexpression of CD 44V6 is correlated with the progress and metastasis of ocular squamous cell carcinomas CD 44V6 protein positive staining is associated with high PI CD 44V6 and PCNA are useful for evaluating prognosis
