| Objective1. To establish the rat kidney transplantation model and observe the effect of the preservation of the kidney in ECs containing decoy ODN/Liposome complexes; 2. To evaluate the transfection efficiency of decoy ODN using liposome method into donor kidney under hypothermic and hypoxia condition and the affection of decoy ODN in the early inflammatory injury of the renal grafts. Methods1. To establish the rat kidney transplantation:25 adult male SD rats and Wistar rats. To harvest donor organs include arteria abdominalis, left kidney, left kidney vessels, left ureter, vesica urinarya and a part of bladder. Continuous overhand suture were applied in anastomosis for kidney transplantation, and titanium clips technique was used in anastomosis for ureter and bladder.2.Experimental Group:The rats were divided into 4 groups. Control group: the Wistar rats underwent the sham operation and the kidney was harvested after 6h. ECs roup: The donor kidney was stored for 6h under of hypothermic and hypoxia condition in ECs without ODN/Liposome complexes, then the donor kidney was transplanted into a SD recipient. The allograft was harvested at 6h posttransplantation. Decoy ODN group: The donor kidney was stored for 6h in ECs containing 1 μ M Liposome/decoy ODN complexes under hypothermic and hypoxia condition, then the donor kidney was transplanted into a SD recipient. The allograft was harvested at 6h posttransplantation. Scrambled ODN group: The donor kidney was stored for 6h in ECs containing 1 μM Liposome/scrambled ODN complexes under hypothermal and hypoxia condition, then the donor kidney was transplanted into a SD recipient, and at 6h posttransplantation the allograft was harvested. The distribution of FITC-labeled ODN in renal tissue were observed by fluorescence microscope and TCS SP2 Confocal microscope .The effect of NF-κ B decoy ODN on theactivity of NF- k B in renal allografts were measured by EMSA(electrophoretic mobility shift assay,EMSA); The gene expression of ICAM-1 > VCAM-1 > CINC in renal allografts were detected by RT-PCR; Renal allograft structure was assessed by light microscopy; The count and distribution of EDl-postive cells in renal allograft were evaluated by immunohistological study. ResultThe graft morphology was not affected by decoy ODN treatment.FITC-labeled ODN was mainly distributed in glomeruli of the transfected renal allograft at 15min posttransplantation, and mainly in glomeruli and the renal tubules at 6h posttransplantation. The most of ODN was transduced into nulei at 6h posttransplantation. The fluroscence caruft be seen in the ECs group. The RDU was 0.84+0.50 in the control group; and it was 57.65+3.58 in the ECs group; Compared to ECs group, the RDU was 1.16+0.28(p<0.05)in the decoy ODN group, however, the RDU was 57.57+4.12(p>0.05)in the scram.ODN group. Compared to the control group, the gene expression of ICAM-1, VCAM-1, CINC was observably increased in ECs group(p>0.05); it was significantly inhibited in the decoy ODN group(p<0.05) compared to the ECs group and the scram.ODN group. There was no significant difference between the ECs group and the scram.ODN group(p>0.05).Immunohistological study showed the infiltrating monocytes was increased and infiltration in tubulointerstitial region in the ECs group. In contrast, a minor infiltrate was seen in the tubulointerstitium in decoy ODN-treated grafts. But there were more infiltrate in the scram.ODN-treated grafts.Conclusionl.The graft morphology was well preserved by decoy ODN treatment. 2.Decoy ODN can be efficiently transfected into kidney using in vivo liposome method in ECs under hypothermic and hypoxic condition;3. The renal allograft treated with decoy ODN can markedly suppress the DNA binding ability of NF-kB,4. Decoy ODN can inhibit the gene expression of ICAM-1, VCAM and CINC.5.The infiltration of M/M was significantly reduced. Meanwhile, the graft morphology can be well pretected.
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