转化生长因子-β1转基因联合FTY720对大鼠心脏移植物缺血-再灌注损伤的影响

目的 探讨经冠脉灌注转化生长因子(TGF)-β1基因联合FTY720对移植心脏缺血-再灌注损伤(IRI)的影响及其机制.方法 实验分为空白对照组、空载体组、FTY720组、转基因组和转基因+FTY720组.心脏移植8 h后切取移植心脏,免疫组织化学检测mTGF-β1、ICAM-1、核转录因子(NF)-κB表达;逆转录-聚合酶链反应(RT-PCR)检测mTGF-β1 mRNA转录强度;测定超氧化物歧化酶(SOD)活性、丙二醛(MDA)含量、髓过氧化物酶(MPO)活性.结果 mTGF-β1成功转到心肌细胞,转基因组和转基因+FIY720组mTGF-β1的表达强度分别是(1.08±0.24)和(1.16±0.22),明显高于其他3组(P<0.01);而两组ICAM-1的表达积分分别是(2.43±0.46)和(1.90±0.20),NF-κB的表达积分分别是(9.80±1.85)和(10.10±2.27),较其他3组显著下调(P<0.01);FRY720组、转基因组和转基因+FTY720组心肌细胞凋亡指数分别是(9.20±1.12)、(5.90±1.09)和(5.40±0.77),显著减少(P<0.01);FTY720组、转基因组和转基因+FTY720组SOD活性分别是(51.03±5.54)、(55.91±6.66)和(73.42±6.42)U/mg,明显升高(P<0.01),MDA含量(10.90±1.93)、(11.02±2.45)和(9.28±1.64)U/g,明显下降(P<0.01);而MPO活性分别是(4.38±1.43)、(4.63±1.04)和(3.16±0.64)U/g,亦明显下降(P<0.01);转基因和FTY720两因素对减轻心肌细胞凋亡以及对SOD、MDA和MPO的影响存在交互作用(P<0.05).结论 TGF-β1和FTY720均具有减轻移植心脏缺血-再灌注损伤的作用,机制可能与抑制心肌细胞凋亡、下调ICAM-1、NF-κB表达、增高SOD活性等因素有关;两者联合应用在减轻缺血-再灌注损伤方面有协同作用.

Effect of Ad. mTGF-1-gene transfection in vitro combined with FTY720 administration on ischemia-reperfusion injury in rat cardiac allografts

Objective To explore the role of Ad. mTGF-β1 gene transfection in vitro combined with FTY720 administration on ischemia-reperfusion injury after heart transplantation in rats, to determine if the combination of them had a synergistic effect, and to discuss the mechanisms. Methods All animals were divided into 5 groups:control group,control vector only-group, FTY720-treated group, Ad. mTGF-β1-transfected group,and Ad. mTGF-β1 FTY720-treated group. All recipients were sacrificed, and the donor hearts were removed at the 8th h after transplantation. The expression of mTGF-β1,ICAM-1,and NF-κB in the heart allografts was detected by using immunohistochemieal staining. Gene product expression in the tissues was quantified by one step RT-PCR. SOD,MDA,and MPO in the grafts were measured. Results The expression of mTGF-β1 in Ad. mTGF-β1-transfected and Ad. mTGF-β1 FTY720-treated groups was (1.08±0.24) and (1.16±0.22), higher than in other three groups (P<0.01). The expression of ICAM and NF-κB in Ad. mTGF-β1-transfected group was (2.43±0.46) and (9.80±1.85), and (1.90±0.20) and (10.1±2.27) in Ad. mTGF-β1 FTY720-treated group respectively,which were lower than in other three groups (P<0.01). The apoptotic index in FTY720 treated, Ad. mTGF-β1-transfected,and Ad. mTGF-β1 FTY720-treated groups was (9.2±1.12), (5.9±1.09) and (5.4±0.77) respec-tively, which was significantly lower than in other groups (P<0.01). SOD activity in FTY720, Ad. mTGF-β1-transfected, and Ad. mTGF-β1 FTY7 20-treated groups was (51.0 3±5.54), (55.91±6.66), and (73.42±6.42) U/mg respectively, higher than other groups (P<0.01). MDA content in FTY720, Ad.mTGF-β1-transfected, and Ad. mTGF-β1 FTY720-treated groups was (10.90±1.93), (11.02±2.45),and (9.28±1.64) U/g respectively, lower than in other groups (P<0.01). MPO activity in FTY720,Ad. mTGF-β1-transfected, and Ad. mTGF-β1 FTY720-treated groups was (4.38±1.43), (4.63±1.04),and (3.16±0.64) U/g respectively, lower than in other groups (P<0.01). Interactions were proven above in gene transfer and FTY720. Conclusion Combined Ad. mTGF-β1 gene transfection in vitro with FTY720 ameliorates the myocardial ischemia-reperfusion injury in rat heart transplantation, in-creases the SOD activity and decreases the MDA content and MPO activity, and reduce the expressions of ICAM-1 and NF-κB. As a result,the combination of mTGF-β1 gene transfection in vitro and FTY720 ad-ministration showed an additive effect.