To study the effect on regulation of cell cycle of osteosarcoma cell line MG63 tranceduced with exogenous p16ink4a and hRbl genes, pIRES-p16ink4a-hRb1, pIRES-p16ink4a and pIREShRbl plasmids were constructed by gene re...To study the effect on regulation of cell cycle of osteosarcoma cell line MG63 tranceduced with exogenous p16ink4a and hRbl genes, pIRES-p16ink4a-hRb1, pIRES-p16ink4a and pIREShRbl plasmids were constructed by gene recombination technology. The recombinant plasmid was transferred into osteosarcoma cell line MG63 by metafectene, and the resistant clones were selected by G418 selective medium, mRNA and protein expression of osteosarcoma cell line were assayed by RT-PCR and Western-Blot respectively. Cell cycle and apoptosis were analyzed by subG1 flow cytometric. Cell proliferation was tested by MTT. In the genome of these transfected target cells, the expression of p16ink4a and hRb1 mRNA and protein were detected respectively in vitro. It was demonstrated with subG1 flow cytometric analysis and MTT method that p16ink4a and hRbl genes cooperation more significantly inhibited cell growth and induced a more marked G1 arrest and apoptosis than p16ink4a/hRb1 alone （P〈0.01）. Coexpression of exogenous p16ink4a with hRbl broke the regulatory feedback loop of p16ink4a-cyclinD1/CDK-hRbl and played a more significant role in inhibiting cell growth as well as inducing cell apoptosis than p16ink4a or hRbl did alone in vitro.展开更多
Observation of stilbene dropping pill and yiqi drug-containing serum influence mechanism of vascular smooth muscle proliferation， cell cycle and Cyclin D1 and CDK4Choose male SD rats were randomly divided into 2 gr...Observation of stilbene dropping pill and yiqi drug-containing serum influence mechanism of vascular smooth muscle proliferation， cell cycle and Cyclin D1 and CDK4Choose male SD rats were randomly divided into 2 groups， lavage qishen yiqi pill and the gastric saline group，extract the drug-containing serum and normal serum；To set the two groups of serum respectively different concentrations，concentration in different time by CCK8 detection effects on vascular smooth muscle cell proliferation， select best concentration and action time.Flow cytometry instrument and high-throughput screening detect serum medicated effect on vascular smooth muscle cell cycle；Western blot detect the drug-containing serum of cell cycle protein Cyclin D1 and CDK4 expression.Result is 5%， 10% medicated serum inhibits cell proliferation significantly higher than the normal serum concentrations of same within 24 h， 48 h.G1 phase cells 5% medicated serum group was obviously higher than that of 5% in normal group (P<005)， serum and cell proliferation index significantly less than 5% normal serum group (P<005)，At the same time， Cyclin D1 and CDK4 expression significantly less than 5% normal serum group (P<005).Conclusion serum of qishen yiqi pill can inhibit vascular smooth muscle cell proliferation， may be through inhibiting cell cycle protein Cyclin D1 and CDK4 expression， block the cell cycle G1 process is closely related to the role.展开更多
The work herein employed a rotating packed bed(RPB)to intensify the sulfonation process of 1,4-diaminoanthraquinone leuco(DL)in an attempt to improve the yield of the product 1,4-diaminoanthra quinone-2-sulfonic acid(...The work herein employed a rotating packed bed(RPB)to intensify the sulfonation process of 1,4-diaminoanthraquinone leuco(DL)in an attempt to improve the yield of the product 1,4-diaminoanthra quinone-2-sulfonic acid(DSA).First,the effects of operating conditions in a stirred tank reactor(STR),including stirring speed,chlorosulfonic acid/DL molar ratio(η),solvent/DL mass ratio(ζ),reaction temperature and dropping speed of chlorosulfonic acid,on the yield of DSA were investigated.The yield of DSA can reach 87.34%under the optimal operating conditions:stirring speed of 500 r·min^(-1),ηof 4.5,ζof 7,reaction temperature of 150℃,dropping speed of 0.61 ml·min^(-1).In addition,the kinetics of the sulfonation process via the shrinking core model revealed that the reaction is controlled by diffusion via a product layer under the reaction temperature of 140℃.Furthermore,the RPB was employed to intensify the mass transfer between liquid and solid phases during the sulfonation reaction process.The results showed that the DSA yield of 92.69%obtained by RPB was 5.35%higher than that by STR,indicating that RPB can significantly intensify the mass transfer in the liquid-solid phase sulfonation reaction process.展开更多
Objective To investigate the roles of the cyclin D1/CDK4 and E2F-1/4 pathways and compare their work patterns in cell cycle changes induced by different doses of B[a]E Methods Human embryo lung fibroblasts （HELFs） w...Objective To investigate the roles of the cyclin D1/CDK4 and E2F-1/4 pathways and compare their work patterns in cell cycle changes induced by different doses of B[a]E Methods Human embryo lung fibroblasts （HELFs） were treated with 2 μmol/L or 100 μmol/L B[a]P which were provided with some characteristics of transformed cells （T-HELFs）. Cyclin D l, CDK4 and E2F-1/4 expressions were determined by Western blotting. Flow cytometry was used to detect the distribution of cell cycle. Results After B[a]P treatment, the proportion of the first gap （G 1） phase cells decreased. CDK4 and E2F-4 expression did not change significantly. In 2 μmol/L treated cells, a marked overexpression of cyclin D1 and E2F-1 was observed. However, in T-HELFs overexpression was limited to cyclin D1 only, and no overexpression of E2F-1 was observed. The decreases of G1 phase in response to B[a]P treatment were blocked in antisense cyclin D1 and antisense CDK4 transfected HELFs （A-D1 and A-K4） and T-HELFs （T-A-D1 and T-A-K4）. After 2 μmol/L B[a]P treatment, overexpression of E2F-1 was attenuated in A-D1, and E2F-4 expression was decreased significantly in A-K4. In T-A-D1 and T-A-K4, E2F-4 expression was increased significantly, compared with T-HELFs. The E2F-1 expression remained unchanged in T-A-D1 and T-A-K4. Condusions Cyclin DI/CDK4-E2F-1/4 pathways work in different patterns in response to low dose and high dose B[a]P treatment. In HELFs treated with 2 μmol/L B[a]P, cyclin D1 positively regulates the E2F-1 expression while CDK4 negatively regulates the E2F-4 expression; however, in HELFs treated with 100 μmol/L B[a]P, both cyclin D1 and CDK4 negatively regulate the E2F-4 expression.展开更多
Latent membrane protein 1 （LMP1）, an important protein encoded by Epstein Barr virus （EBV）, has been implied to link with the pathogenesis of nasopharyngeal carcinoma （NPC）. Its dual effects of increasing cell p...Latent membrane protein 1 （LMP1）, an important protein encoded by Epstein Barr virus （EBV）, has been implied to link with the pathogenesis of nasopharyngeal carcinoma （NPC）. Its dual effects of increasing cell proliferation and inhibiting cell apoptosis have been confirmed. In this study, we showed that the expression of Survivin and CDK4 protein in CNE-LMP1, a LMP1 positive NPC epithelial cell line, is higher than in LMP1 negative NPC epithelial cell line- CNE1, and the expression is LMP1 dosage-dependent. Although it was reported that Survivin specifically expressed in cell cycle G2/M phase, our studies suggested that LMP1 could promote the expression of Survivin in G0/G1, S and G2/ M phase. It also showed that Survivin and CDK4 could be accumulated more in the nuclei triggered by LMP1. More interestingly, Survivin and CDK4 could form a protein complex in the nuclei of CNE-LMP1 rather than in that of CNE1, which demonstrated that the interaction between these two proteins could be promoted by LMPI. These results strongly suggested that the role of LMP1 in the regulation of Survivin and CDK4 may also shed some light on the mechanism research of LMP1 in NPC.展开更多
文摘To study the effect on regulation of cell cycle of osteosarcoma cell line MG63 tranceduced with exogenous p16ink4a and hRbl genes, pIRES-p16ink4a-hRb1, pIRES-p16ink4a and pIREShRbl plasmids were constructed by gene recombination technology. The recombinant plasmid was transferred into osteosarcoma cell line MG63 by metafectene, and the resistant clones were selected by G418 selective medium, mRNA and protein expression of osteosarcoma cell line were assayed by RT-PCR and Western-Blot respectively. Cell cycle and apoptosis were analyzed by subG1 flow cytometric. Cell proliferation was tested by MTT. In the genome of these transfected target cells, the expression of p16ink4a and hRb1 mRNA and protein were detected respectively in vitro. It was demonstrated with subG1 flow cytometric analysis and MTT method that p16ink4a and hRbl genes cooperation more significantly inhibited cell growth and induced a more marked G1 arrest and apoptosis than p16ink4a/hRb1 alone （P〈0.01）. Coexpression of exogenous p16ink4a with hRbl broke the regulatory feedback loop of p16ink4a-cyclinD1/CDK-hRbl and played a more significant role in inhibiting cell growth as well as inducing cell apoptosis than p16ink4a or hRbl did alone in vitro.
文摘Observation of stilbene dropping pill and yiqi drug-containing serum influence mechanism of vascular smooth muscle proliferation， cell cycle and Cyclin D1 and CDK4Choose male SD rats were randomly divided into 2 groups， lavage qishen yiqi pill and the gastric saline group，extract the drug-containing serum and normal serum；To set the two groups of serum respectively different concentrations，concentration in different time by CCK8 detection effects on vascular smooth muscle cell proliferation， select best concentration and action time.Flow cytometry instrument and high-throughput screening detect serum medicated effect on vascular smooth muscle cell cycle；Western blot detect the drug-containing serum of cell cycle protein Cyclin D1 and CDK4 expression.Result is 5%， 10% medicated serum inhibits cell proliferation significantly higher than the normal serum concentrations of same within 24 h， 48 h.G1 phase cells 5% medicated serum group was obviously higher than that of 5% in normal group (P<005)， serum and cell proliferation index significantly less than 5% normal serum group (P<005)，At the same time， Cyclin D1 and CDK4 expression significantly less than 5% normal serum group (P<005).Conclusion serum of qishen yiqi pill can inhibit vascular smooth muscle cell proliferation， may be through inhibiting cell cycle protein Cyclin D1 and CDK4 expression， block the cell cycle G1 process is closely related to the role.
基金financially supported by the National Key Research and Development Program of China(2016YFB0301500)the National Natural Science Foundation of China(21878009)。
文摘The work herein employed a rotating packed bed(RPB)to intensify the sulfonation process of 1,4-diaminoanthraquinone leuco(DL)in an attempt to improve the yield of the product 1,4-diaminoanthra quinone-2-sulfonic acid(DSA).First,the effects of operating conditions in a stirred tank reactor(STR),including stirring speed,chlorosulfonic acid/DL molar ratio(η),solvent/DL mass ratio(ζ),reaction temperature and dropping speed of chlorosulfonic acid,on the yield of DSA were investigated.The yield of DSA can reach 87.34%under the optimal operating conditions:stirring speed of 500 r·min^(-1),ηof 4.5,ζof 7,reaction temperature of 150℃,dropping speed of 0.61 ml·min^(-1).In addition,the kinetics of the sulfonation process via the shrinking core model revealed that the reaction is controlled by diffusion via a product layer under the reaction temperature of 140℃.Furthermore,the RPB was employed to intensify the mass transfer between liquid and solid phases during the sulfonation reaction process.The results showed that the DSA yield of 92.69%obtained by RPB was 5.35%higher than that by STR,indicating that RPB can significantly intensify the mass transfer in the liquid-solid phase sulfonation reaction process.
基金Grants of National Natural Science Foundation of China (30371206, 30028019)973 National Key Basic Research and Development Program (2002 CB 512905)
文摘Objective To investigate the roles of the cyclin D1/CDK4 and E2F-1/4 pathways and compare their work patterns in cell cycle changes induced by different doses of B[a]E Methods Human embryo lung fibroblasts （HELFs） were treated with 2 μmol/L or 100 μmol/L B[a]P which were provided with some characteristics of transformed cells （T-HELFs）. Cyclin D l, CDK4 and E2F-1/4 expressions were determined by Western blotting. Flow cytometry was used to detect the distribution of cell cycle. Results After B[a]P treatment, the proportion of the first gap （G 1） phase cells decreased. CDK4 and E2F-4 expression did not change significantly. In 2 μmol/L treated cells, a marked overexpression of cyclin D1 and E2F-1 was observed. However, in T-HELFs overexpression was limited to cyclin D1 only, and no overexpression of E2F-1 was observed. The decreases of G1 phase in response to B[a]P treatment were blocked in antisense cyclin D1 and antisense CDK4 transfected HELFs （A-D1 and A-K4） and T-HELFs （T-A-D1 and T-A-K4）. After 2 μmol/L B[a]P treatment, overexpression of E2F-1 was attenuated in A-D1, and E2F-4 expression was decreased significantly in A-K4. In T-A-D1 and T-A-K4, E2F-4 expression was increased significantly, compared with T-HELFs. The E2F-1 expression remained unchanged in T-A-D1 and T-A-K4. Condusions Cyclin DI/CDK4-E2F-1/4 pathways work in different patterns in response to low dose and high dose B[a]P treatment. In HELFs treated with 2 μmol/L B[a]P, cyclin D1 positively regulates the E2F-1 expression while CDK4 negatively regulates the E2F-4 expression; however, in HELFs treated with 100 μmol/L B[a]P, both cyclin D1 and CDK4 negatively regulate the E2F-4 expression.
基金National Nature Science Foundation for Distinguished Young Scholar of China (No.39525022)National Basic Research Program(No.2004CB518703) National Nature Science Foundation of China (No.30570085).
文摘Latent membrane protein 1 （LMP1）, an important protein encoded by Epstein Barr virus （EBV）, has been implied to link with the pathogenesis of nasopharyngeal carcinoma （NPC）. Its dual effects of increasing cell proliferation and inhibiting cell apoptosis have been confirmed. In this study, we showed that the expression of Survivin and CDK4 protein in CNE-LMP1, a LMP1 positive NPC epithelial cell line, is higher than in LMP1 negative NPC epithelial cell line- CNE1, and the expression is LMP1 dosage-dependent. Although it was reported that Survivin specifically expressed in cell cycle G2/M phase, our studies suggested that LMP1 could promote the expression of Survivin in G0/G1, S and G2/ M phase. It also showed that Survivin and CDK4 could be accumulated more in the nuclei triggered by LMP1. More interestingly, Survivin and CDK4 could form a protein complex in the nuclei of CNE-LMP1 rather than in that of CNE1, which demonstrated that the interaction between these two proteins could be promoted by LMPI. These results strongly suggested that the role of LMP1 in the regulation of Survivin and CDK4 may also shed some light on the mechanism research of LMP1 in NPC.
