【作者】 杨天宝；

【导师】 康明强；

【作者基本信息】 福建医科大学 ， 外科学， 2009， 硕士

【摘要】 肺移植已成为肺纤维化、原发性肺动脉高压、肺气肿、支气管扩张等慢性终末期肺病的最有效治疗措施,在肺移植中缺血再灌注所致肺损伤是移植失败和早期肺移植患者死亡的主要原因,因而如何减轻缺血再灌注损伤已成为现阶段研究的重要课题。缺血再灌注的病理生理过程中,氧自由基的快速大量产生及细胞间Ca2+积聚,这两种情况均可导致细胞凋亡的发生。肝细胞生长因子(hepatocyte growth factor HGF)具有很强的促进肺上皮细胞生长、抑制缺血-再灌注诱导的肺上皮细胞凋亡作用,故我们运用HGF的抗凋亡活性,起到减轻肺缺血再灌注的损伤的作用。课题分为以下两个部分:第一部分:改良三袖套法建立大鼠左肺原位移植动物模型目的探讨建立更稳定有效的大鼠原位左肺移植模型。方法将40只SD大鼠随机配对,采用三袖套法建立大鼠肺移植模型。结果进行大鼠左肺原位移植正式实验20例,手术平均时间(65±4)min。手术成功率85%。血气、病理学等检查证实成功复制了肺移植缺血再灌注模型。讨论该方法操作成功率高,经济实用,高度模仿临床肺移植,可用于肺移植缺血再灌注损伤的实验研究。第二部分:肝细胞生长因子对大鼠肺移植缺血再灌注损伤的保护作用目的通过建立大鼠肺移植缺血再灌注模型,探讨肝细胞生长因子对大鼠肺移植缺血再灌注损伤的影响及其作用机制。方法实验分3组,即假手术对照组(A组)、空白对照组(B组)和HGF组(C组),每组8例。B组用4℃LPDG液灌洗并保存供肺4h,C组用含HGF(10ug/100ml)的4℃LPDG液灌洗并保存供肺4h。移植肺再灌注2h后结束实验。测定肺组织湿-干重比、MPO活性,观察移植肺组织病理改变,ELISA测定TNF-α表达水平,免疫组化染色检测c-met表达,RT-PCR检测Bcl-2 mRNA的水平,TUNEL检测细胞凋亡。结果与空白对照组比较,HGF组肺组织损伤程度明显减轻,肺组织湿干重比、MPO活性、TNF-α均有下降(P < 0.01)。c-met、Bcl-2 mRNA的表达水平较空白对照组明显增加(P < 0.01)。讨论HGF能有效地减轻移植肺缺血再灌注损伤,明显改善移植肺功能,其保护作用可能与刺激c-met的表达,增加抗凋亡基因bcl-2的表达有关。更多还原

【Abstract】 Lung transplantation has become the most effective therapeutic measure for chronicity pulmonary tuberculosis at the final stage, such as pulmonary fibrosis, primary pulmonary hypertension, emphysema, bronchiectasis and so on. While lung transplantation ischemia reperfusion injury(IRI) is the main cause of the transportation failure and patients’death at the earlier period. Therefore, how to lessen ischemical reperfusion injury has been the major point at the very present study period. During the patho and physio process of ischemia reperfusion, rapidly increasing oxyradicals and accumulation of Ca2 + among cells may lead to the apoptosis.Hepatocyte growth factor HGF has extreme effection on enhancement of pulmonary epithelial cells and suppression on IRI which leads to apoptosis pulmonary epithelial cells. Hence, we make full use of antiapoptosis of HGF which can alleviate the pulmonary I/R injury. This whole study consists of two parts as follows:1. To establish rat orthotopic left pulmonary allograft transplantation by improving the“cuff-like”vessel anastomosis technique.Objective To establish a more stable and effective rat orthotopic left pulmonary allograft transplantation models.Methods 40 rats were randomly divided into two groups: donors group and recipients group. Pulmonary arteries and veins were anastomosed by traditional cuff technique.Bronchi was reconstructed by stent techniqueResults 20 cases of rat orthotopic left pulmonary allograft transplantation was finished successfully. The total time of surgical procedure was (65±4)min. The operation successful rate was 85%. By blood gas and pathologic study we successfully found the typical lung ischemia reperfusion injury model.Conclusion It is an economical and successful model in highly imitating clinic lung transplantation, which can be used in lung ischemia reperfusion injury, 2. Protection of Hepatocyte Growth Factor in Rat’s Lungs PreservationObjective To establish a more stable and effective rat orthotopic left pulmonary allograft transplantation models. To discuss the protection and mechanisms of hepatocyte growth factor in transplantated rat’s lungs from ischemia reperfusion injury.Methods This study was divided into three groups: sham operation control group(A group),empty control group(B group) and HGF group(C group). Every group had 8 cases.The harvested lung blocks were flushed and stored in 4℃low-potassium-dextran and glucose (LPDG) solution for 4 hours in B group. 4℃LPDG solution which contains HGF(10ug/100ml)was used in C group. The experiment was over after flushing the transplanted lung 2h. The transplanted lung tissue wet-to-dry (W/D) ratio and activity of myeloperoxidase (MPO) were measured. TNF-αwas detected by ELISA. C-met were detected by immunohistochemical staining. Bcl-2 mRNA was detected by RT-PCR. Apoptosis is detected by TUNEL.Results Comparing with the empty control group, obviously, the degree of lung tissue injury in the HGF group reduces, and the lung tissue wet-dry weight ratio, MPO activity, TNF-αalso decrease (P <0.01). Moreover, the expression levels of c-met, Bcl-2 mRNA increased significantly (P <0.01), too.Conclusion HGF can effectively reduce IRI and further improve transplanted lung functions.The protection mechanism is probably associated with the stimulation of c-met and bcl-2 expression.更多还原