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| 1 | Inhibition of mouse hepatocyte apoptosis via anti-Fas ribozyme显示文摘AIM: TO investigate the effects of anti-Fas ribozyme on Fas expression and apoptosis in primary cultured mouse hepatocytes.METHODS: Mouse hepatocytes were isolated by using collagenase irrigation. A hammerhead ribozyme targeting the Fas mRNA was constructed, and transfected into mouse hepatocytes via Effectene. Then Fas expression in mouse hepatocytes was detected by RT-PCR and western blotting.After being treated with anti-Fas antibody (JO2), hepatocytes viability was measured with MTI- assay. Caspase-3 proteolytic activity was detected, and cell apoptosis was measured according to Annexin V-FITC apoptosis detection kit.RESULTS: Fas expressed in primary mouse hepatocytes.Fas expression in hepatocytes transfected with anti-Fas ribozyme was decreased remarkably and correlated with the resistance to Fas-mediated apoptosis as determined by flow cytometry and caspase-3 proteolytic activity.CONCLUSION: AnU-Fas ribozyme can remarkably decrease the Fas expression in mouse hepatocytes, thus inhibit Fasmediated apoptosis in hepatocytes, It is suggested that anti-Fas ribozyme could significantly increase the resistance of transplanted hepatocytes to apoptosis and improve the survival of transplanted hepatocytes, | MinZhang WeiHe FangLiu PingZou JuanXiao Zhao-DongZhong Zhong-BoHu | 2004 | World Journal of Gastroenterology2004，10，17： | 8 |
| 2 | Repression of allo-cell transplant rejection through CⅡTA ribonuclease P^+hepatocyte显示文摘AIM: Allo-cell transplant rejection and autoimmune responses were associated with the presence of class Ⅱmajor histocompatibility complex (MHC Ⅱ) molecules on cells.This paper studied the effect of Ribonuclease P (RNase P)against CIITA, which was a major regulator of MHCII molecules, on repressing the expression of MHCII molecules on hepatocyte.METHODS: M1-RNA is the catalytic RNA subunit of RNase P from Escherichia coli. It were constructed that M1-RNA with guide sequences (GS) recognizing the 452, 3408 site of CIITA by PCR from pTK117 plasmid, then were cloned into the EcoRI/Bg/II or EcoR//SalIsite of vector psNAV (osNAV-M1-452-GS, psNAV-M1-3408-GS) respectively. The target mould plate (3176-3560) of CIITA was obtained from Raji cell by RT-PCR, and then inserted into the XhoI/EcoRIof pGEM-7zf(+) plasmid (pGEM-3176). These recombinant plasmids were screened out by sequence analysis. psNAV-M 1-452-GS, psNAV-M1-3408-GS and its target RNA pGEM-3176 were transcribed and then mixed up and incubated in vitro. It showed that M1-3408-GS could exclusively cleave target RNA that formed a base pair with the GS. Stable transfectants of hepatocyte cell line with psNAVl-M1-3408-GS were tested for expression of class Ⅱ MHC through FCM, for mRNA abundance of MHCII, Ii and CIITA by RTPCR., for the level of IL-2 mRNA on T cell by mixed lymphocyte reaction.RESULTS: When induced with recombinant human interferon-gamma (IFN-γ), the expression of HLA-DR, -DP,-DQ on psNAV-M1-3408-GS+ hepatocyte was reduced 83.27 %, 88.93 %, 58.82 % respectively, the mRNA contents of CIITA, HLA-DR, -DP, -DQ and Ii decreased significantly.While T cell expressed less IL-2 mRNA in the case of psNAV-M1-3408-GS+ hepatocyte.CONCLUSION: The Ribonuclease P against CIITA-M1-3408-GS could effectively induce antigen-specific tolerance through cleaving CIITA. These results provided insight into the future application of M1-3408-GS as a new nucleic acid drug against allo-transplantation rejection and autoimmune diseases. | RongGuo PingZou Hua-HuaFan FengGao Qing-XinShang Yi-LinCao Hua-ZhongLu | 2003 | World Journal of Gastroenterology2003，9，5： | 5 |
| 3 | Inhibition of Fas-mediated Apoptosis in Yac-1 Cell via Anti-Fas Ribozyme显示文摘To detect a new and more effective way against apoptosis mouse lymphomatic cell line- Yac-1 in which fas gene was expressed highly was used as a model for studying the effects of anti-Fas ribozyme on Fas-mediated apoptosis. A hammerhead ribozyme gene targeting the fas mRNA was synthesized and its in vitro transcription vector was constructed, which was transfected into Yac-1 cells using electroporation. Rz596 expression was detected using RT-PCR, and Fas expression in Yac-1 cells was detected using RT-PCR, Western blot and flow cytometry. After treated with anti-Fas antibody (JO2), Yac-1 cell viability was measured with MTT assay, caspase-3 proteolytic activity was detected, and cell apoptosis was measured according to annexin V apoptosis detecting kit. Anti-Fas ribozyme could cleave fas mRNA efficiently in vivo and in vitro. Fas expression in Yac-1 cells transfected with anti-Fas ribozyme was decreased remarkably and correlated with resistance to Fas-mediated apoptosis as determined by flow cytometry and caspase-3 proteolytic activity. Anti-Fas ribozyme was detected in cells transfected with pU6-RZ596 and pU6-dRZ596 and could remarkably decrease the Fas expression in Yac-1 cells, which made Yac-1 cells get rid of Fas-mediated apoptosis. Because of wide expression of fas in organs and tissues, our research was very useful for studying the inhibition of apoptosis of many organs and tissues in the future. | MinZHANG FangLIU WeiHE YongYOU PingZOU Ling-BoLIU | 2004 | Acta Biochimica et Biophysica Sinica2004，36，3： | 3 |
| 4 | Inhibiting Apoptosis of CTLL-2 Cells to Enhance Their GVL Effects via Anti-Fas Ribozyme显示文摘To investigate the inhibition role of anti-Fas hammerhead ribozyme on fas expression and Fas-mediated apoptosis of CTL cell line CTLL-2 cells, the cDNA of an anti-Fas hammerhead ribozyme was synthesized, its expression plasmid was constructed and transfected into CTLL-2 cells by electroporation. fas expression of CTLL-2 cells was detected by RT-PCR and Western blot. CTLL-2 cell viability was measured using MTT assay when co-cultured with mouse T cell leukemia cell line EL4 cells that highly expressed Fas ligand (FasL). Meanwhile, caspase-3 proteolytic activity was detected, and cell apoptosis was measured by flow cytometry and Hochest-PI double staining. Killing activity of CTLL-2 cells was detected by lactate dehydrogenase (LDH) releasing assay in vitro. Results showed that the expression of both Fas mRNA and protein in CTLL-2 cells were decreased after transfection of anti-Fas ribozyme. Compared with mock- transfected group and mutant ribozyme-transfected group, viability of CTLL-2 cells co-cultured with EL4 cells was increased significantly and cells killing activity was enhanced after transfected with anti-Fas ribozyme, while the caspase-3 activity and apoptosis rate was significantly decreased. The results demonstrated anti-Fas ribozyme could efficiently cleave Fas and inhibit Fas-mediated apoptosis of CTLL-2 cells to improve their viability. Our study made a basis for enhancing CTLL-2 cells anti-leukemia effect in DLI. | MinZHANG FangLIU Lin-BoLIU YongYOU Zhi-ChaoCHEN PingZOU | 2004 | Acta Biochimica et Biophysica Sinica2004，36，8： | 0 |
| 5 | Novel Fluorene-based Conjugated Copolymer Containing Cyclobutenedione Unit for Light Emitting Diodes显示文摘A novel fluorene-based conjugated copolymer containing cyclobutenedione unit was synthesized by Suzuki reaction.Its structure and properties were characterized by FTIR,^1HNMR,elemental analysis,PL spectroscopy,DSC,TGA and cyclic voltammetry.The resulting polymer shows strong yellow PL emission(561nm)and good solubility in polar aprotic solvents,i.e.THF,DMF,DMAC,DMSO,etc.DSC and TGA studies reveal that the novel polymer possesses excellent thermal stability with high glass transition temperature of 127℃ and onset decomposition temperature of 411℃.Cyclic voltammetry measurement demonstrated that the polymer has both hole and electron-transporting property. | QiangPENG YanHUANG ZhiYunLU PingZOU MingGuiXIE | 2004 | Chinese Chemical Letters2004，15，4： | 0 |
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