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Background: Placement of an external support has been reported to prevent intimal hyperplasia of vein grafts. However, it’s application limited by potential complications. Peri-adventitial gene delivery is a promising alternative therapy to reduce intimal hyperplasia, but it is limited by low and transient levels of gene transfection. To get more effective inhibition of intimal hyperplasia and to avoid the limitations associated with these two approaches, a study was undertaken to investigate whether mixing adenovirus with fibrin glue may increase the level and prolong the time period of gene expression. Methods: Right jugular vein to common carotid artery interposition grafting was performed in 36 male New Zealand white rabbits (2.5–3.0 kg) and the animals were divided into four groups: control group (n = 6); fibrin glue group (n = 6); Ad-GAL group (n = 12); fibrin glue/Ad-GAL group (n = 12). Commercially available fibrin glue and adenovirus expressing the gene for β-galactosidase (Ad-GAL) was applied separately or in mixing around vein grafts. At 7th day and 14th day after implantation, the grafts were harvested to evaluate transfection rate. At 28th day the grafts were harvested for morphometric analysis. Results: Compared with weak staining in 2.1 ± 0.5% in Ad-GAL alone grafts, a high level of β-Galactosidase staining was evident in 13.2 ± 4.6% in fibrin glue/Ad-GAL grafts at 7th day (P < 0.001). At 14th day, almost no staining (0%) was detected in Ad-GAL alone grafts. However, there was still a relative high level staining (6.3 ± 3.8%) in fibrin glue/Ad-GAL grafts (P < 0.001 versus Ad-GAL alone group). At 28th day, a statistically significantly decrease in neointimal area (0.68 ± 0.06 mm2 versus 1.00 ± 0.08 mm2, P < 0.05) was shown in fibrin glue grafts compared with unsupported vein grafts (control group). The same statistically significantly difference was also existed in fibrin glue/Ad-GAL group and unsupported group in neointimal area (0.66 ± 0.07 mm2, P < 0.05). Conclusions: A novel method of adventitial gene delivery using fibrin glue as external support is proposed. Fibrin glue may be an ideal candidate for controlled release delivery that would facilitate adventitial gene transfer. [Copyright &y& Elsevier]

Background: To construct a primary rat colonic epithelial cell model for treatment with specific methylated oligonucleotides (MOs) and to determine whether the transcriptional inactivation of ERβ mRNA is mediated by the induction of hypermethylation of the ERβ gene promoter. Methods: Suckling rat colonic epithelial cells were cultured in DMEM. Two methylated oligonucleotides complementary to the promoter regions of ERβ were synthesized and applied to the cultured cells to induce promoter hypermethylation of the ERβ gene. Methylation-specific PCR (MSP) was used to determine the methylation status of the ERβ promoter in the cultured cells. Reverse transcription–polymerase chain reaction (RT-PCR) was used to quantify the expression of ERβ mRNA after treatment with MOs. Results: Suckling rat colonic epithelial cells were successfully cultured in vitro. The MOs and unmethylated oligonucleotides (UMOs) we designed, synthesized, successfully transfected into the colonic epithelial cells, and assembled in the nuclei of the cells, which had extremely elevated proliferative activity. RT-PCR demonstrated that the expression of ERβ mRNA was significantly suppressed in the cells treated with MOs, whereas its expression in the control cells treated with UMOs was not. MSP analysis showed that the promoter of ERβ in the cells treated with MOs was hypermethylated compared with that of the control cells. Conclusion: The transcriptional inactivation of ERβ mRNA in rat colonic epithelial cells may be mediated by the hypermethylation of the ERβ gene promoter. Our model markedly simulates the epigenetic modification of the ERβ gene in colonic cancer cells. [Copyright &y& Elsevier]