Objective： To construct the recombinant adenovirus expressing small RNA of rats caspase-3 and observe the down-regulation effect of caspase-3 in neurons induced by lipopolysaccharide（LPS） in vitro. Methods： pShutt...Objective： To construct the recombinant adenovirus expressing small RNA of rats caspase-3 and observe the down-regulation effect of caspase-3 in neurons induced by lipopolysaccharide（LPS） in vitro. Methods： pShuttleHl-siCas3 containing Oligo DNA of the targeting sequences and pEGFPC1-Cas3 containing caspase-3 and EGFP sequences were constructed respectively, pShuttleH 1-siCas3 and pEGFPC 1-Cas3 were co-transfected to the 293 cells by liposomes to determine interfering efficacy by flow cytometry, pShuttleHl-siCas3 was linearized and transformed into E. coli B J5183 cells containing backbone plasmid pAdEasy-1. The recombinant plasmid was transfected into 293 cells to package the adenovirus Ad-siCas3. The titers of adenovirus were determined by the specific 50% tissue culture infection dosage method. After virus infected the cultured hippocampus neurons, LPS-induced apoptosis and caspase-3 mRNA expression were observed. Results： It was identified that the sequence of target gene was correctly inserted into the genome of virus. The expression of green fluorescence protein was reduced by pShuttleHl-siCas3 in 293 cells. The titer of recombinant adenovirus was 1.06×10^10pfu/ml. After virus infection, caspase-3 mRNA was greatly reduced and neurons apoptosis was suppressed. Conclusion： The recombinant adenovirus expressing rats caspase-3 siRNA were successfully constructed, which may probably be further used in pain therapy by its anti-apoptosis effect.展开更多
In the last few years,the overall incidence of malignant tumors has been increasing in the country.In order to find effective anti-tumor drugs,the development and modification of natural drugs have become the focus of...In the last few years,the overall incidence of malignant tumors has been increasing in the country.In order to find effective anti-tumor drugs,the development and modification of natural drugs have become the focus of domestic and international researchers.Thallus japonica,composed of polysaccharides,minerals,amino acids,natural proteins,and lipids,is a traditional Chinese medicine with the functions of eliminating phlegm and disinhibiting water.At the same time,it has the edible and medicinal efficacy.There are a lot of active components that have been demonstrated to possess diverse biological activities including anti-tumor,anti-viral,anti-coagulant,and anti-bacterial,and so on in thallus japonica.Laminarin,composed of glucose,galactose,fucose and so on,is a common and high molecular weight heteropolysaccharide in brown algae(such as thallus japonica,kelp),and it is a potential anti-tumor drug.However,the lack of biological activity of natural polysaccharides limits the application of polysaccharides,so it is necessary to modify the structure or physical and chemical properties of it to enhance the anti-tumor activity of polysaccharides.In recent years,in order to improve the biological activity of laminarin,domestic and international researchers have introduced some groups on the surface of it.Moreover,use the function of modified groups to cause the changes in physicochemical property,spatial structure,substituent group and molecular weight to improve the anti-tumor activity in vitro and in vivo.Therefore,the structural modification has become the main and powerful means to improve the activity of polysaccharides.展开更多
文摘Objective： To construct the recombinant adenovirus expressing small RNA of rats caspase-3 and observe the down-regulation effect of caspase-3 in neurons induced by lipopolysaccharide（LPS） in vitro. Methods： pShuttleHl-siCas3 containing Oligo DNA of the targeting sequences and pEGFPC1-Cas3 containing caspase-3 and EGFP sequences were constructed respectively, pShuttleH 1-siCas3 and pEGFPC 1-Cas3 were co-transfected to the 293 cells by liposomes to determine interfering efficacy by flow cytometry, pShuttleHl-siCas3 was linearized and transformed into E. coli B J5183 cells containing backbone plasmid pAdEasy-1. The recombinant plasmid was transfected into 293 cells to package the adenovirus Ad-siCas3. The titers of adenovirus were determined by the specific 50% tissue culture infection dosage method. After virus infected the cultured hippocampus neurons, LPS-induced apoptosis and caspase-3 mRNA expression were observed. Results： It was identified that the sequence of target gene was correctly inserted into the genome of virus. The expression of green fluorescence protein was reduced by pShuttleHl-siCas3 in 293 cells. The titer of recombinant adenovirus was 1.06×10^10pfu/ml. After virus infection, caspase-3 mRNA was greatly reduced and neurons apoptosis was suppressed. Conclusion： The recombinant adenovirus expressing rats caspase-3 siRNA were successfully constructed, which may probably be further used in pain therapy by its anti-apoptosis effect.
文摘In the last few years,the overall incidence of malignant tumors has been increasing in the country.In order to find effective anti-tumor drugs,the development and modification of natural drugs have become the focus of domestic and international researchers.Thallus japonica,composed of polysaccharides,minerals,amino acids,natural proteins,and lipids,is a traditional Chinese medicine with the functions of eliminating phlegm and disinhibiting water.At the same time,it has the edible and medicinal efficacy.There are a lot of active components that have been demonstrated to possess diverse biological activities including anti-tumor,anti-viral,anti-coagulant,and anti-bacterial,and so on in thallus japonica.Laminarin,composed of glucose,galactose,fucose and so on,is a common and high molecular weight heteropolysaccharide in brown algae(such as thallus japonica,kelp),and it is a potential anti-tumor drug.However,the lack of biological activity of natural polysaccharides limits the application of polysaccharides,so it is necessary to modify the structure or physical and chemical properties of it to enhance the anti-tumor activity of polysaccharides.In recent years,in order to improve the biological activity of laminarin,domestic and international researchers have introduced some groups on the surface of it.Moreover,use the function of modified groups to cause the changes in physicochemical property,spatial structure,substituent group and molecular weight to improve the anti-tumor activity in vitro and in vivo.Therefore,the structural modification has become the main and powerful means to improve the activity of polysaccharides.
