经供体气管转染腺病毒介导的人白介素10基因对大鼠同种异体肺移植物缺血再灌注损伤的影响

The effect of transbronchial gene transfer of human interleukin-10 gene to donor lung ischemia-reperfusion injury in the rat’s lung transplantation

目的:观察局部转染腺病毒介导的人白细胞介素(hIL)-10基因对移植肺缺血再灌注损伤(IRI)和肺功能的影响。方法:36只雄性SD大鼠随机分为空白对照组空载体对照组、基因转染组。在移植前经供体气管分别灌入生理盐水0.5 ml、Ad-5腺病毒5×109PFU和Ad-5-hIL-10 5×109PFU;24 h后正中开胸取肺,在4℃低钾右旋糖酐液(LPD液)中保存18 h。采用改良的三袖套法建立大鼠左肺原位移植模型,移植肺再灌注后4 h测定动脉血气、移植肺湿-干重比率(W/D)、丙二醛(MDA)含量、超氧化物歧化酶(SOD)含量和髓过氧化物酶(MPO)含量;观察移植肺组织病理改变及人hIL-10免疫组化染色;并使用酶联免疫吸附法(ELISA)测定移植组织中肿瘤坏死因子α(TNF-α)、干扰素γ(IFN-γ)的表达水平。结果:与空载体组和空白对照组相比,基因转染组肺泡间质水肿减轻,炎细胞浸润减少,hIL-10基因的表达明显增加,PaO2明显提高(P<0.01),W/D比率降低(P<0.05),MDA含量和MPO活性下降(P<0.01),SOD活性升高(P<0.01),TNF-α和IFN-γ的表达下调(P<0.01)。结论:移植前经供肺气管灌注途径转染外源性人IFN-10基因能有效地减轻移植后肺脏组织IRI,改善早期移植肺功能。

Objectlve：To investigate the effect of transtracheal transfering the adenovirus-mediated human interleukin 10 （hIL-10） gene on lung isehemia-reperfusion injury （IRI）,and posttransplant lung function in SD rat lungs. Methods：Thirty-six male SD rats were divided into 3 groups randomly： blank control group,empty vector-control group and gene transfer group. The donors of three groups received 0.5 ml of normal saline,5×10^9 PFU of Ad-5 or 5×10^9 PFU of Ad-5-hIL-10,respectively. After 24 h of in vivo transfection,lungs were harvested and stored in low-potassium dextran solution （LPDG） at 4℃ for 18 h. Then the left lung orthotopic transplantation was performed. After 4 h of reperfusion,arterial blood gas analysis and wet-to-dry （W/D） weight ratios were determined,superoxide dismutase （SOD） activity,malondialdehyde （MDA） contents and myeloperoxidase （MPO） activity were detected. Tumor necrosis factor-α （TNF-oL） and interferon-3＇ （IFN-3＇） were measured in graft tissue by ELISA. Besides,hIL-lO was detected by immunohistochemistry. Graft pathohistological changes were examined under a light microscope. Results：Empty vector and blank control groups showed a significant lung ischemia-reperfusion injury （IRI）,while gene transfer group＇s lung IRI reduced significantly. The lung tissue edema, interstitial inflammation and exudation in gene transfer group were obviously reduced. Expression of hIL-10 gene was obviously incrdased when detected by immunohistochemistry （IHC）. Partial pressure of oxygen （PaO2） levels in the gene transfer group were higher than in empty vector and blank vector groups （P 〈 0.01）, respectively. W/D ratios were reduced in hIL-10 lungs （P 〈 0.05） compared with the other groups. MDA contents and MPO activity were reduced in tissue of the gene transfer group（P 〈 0.01 ）. But the activity of SOD was increased significantly （P 〈 0.01）. TNF-α and IFN-γ were reduced in tissue of the gene transfer group （P 〈 0.01） measured by ELISA. Conclusion：Transtracheal adenovirus mediated gene transfection of hIL-10can obviously lighten IRI after lung transplantation and improve early lung function effectively after lung transplantation.