实验性急性胰腺炎肺内细胞凋亡状况及其意义的初步探讨

A STUDY ON INTRAPULMONARY CELL APOPTOSIS AND ITS SIGNIFICANCE IN RATS WITH ACUTE PANCREATITIS

目的 :探讨重症急性胰腺炎时肺内细胞凋亡的状况及其在肺损伤发病机制中的意义。方法 :以不同浓度牛磺胆酸钠液逆行胰胆管注射造成大鼠急性水肿性胰腺炎 (AEP)与急性坏死性胰腺炎 (ANP)两种模型 ,测定血浆TNF -α与内毒素水平的动态变化 ,免疫组化检测肺内TNF -α的表达 ,并以TUNEL法结合激光扫描共聚焦显微镜检测肺组织切片内细胞凋亡的情况。结果 :正常时大鼠肺内偶见淋巴细胞及纤维母细胞等发生凋亡 ,诱导AEP或ANP后凋亡细胞数量无明显变化。随着肺损伤的出现 ,少许浸润的炎细胞、肺泡上皮细胞及血管内皮细胞等发生了凋亡。凋亡指数 (‰ )在AEP组呈一过性下降 ,在ANP组表现为持续下降 ,在 6h后各时点均显著低于AEP组相应值 (P <0 .0 5 )。分析表明 ,ANP组血中TNF -α、内毒素含量的增加与凋亡指数的变化存在负相关 (P <0 .0 5 )。结论 :ANP时肺内浸润的以中性粒细胞为代表的大量炎细胞出现延迟凋亡 ,这种现象可能是肺损伤发生的重要前提 ,同时内毒素血症及TNF -α的过度合成可能是中性粒细胞延迟凋亡的部分原因。

Objective:It is known that LPS/TNF-dependent mechanism is a major pathway in modulating cell apoptosis. As our previous study shown, plasma endotoxin and intrapulmonary production of TNF-α increase significantly in acute necrotizing pancreatitis (ANP). So we want to know the state of cell apoptosis in lung tissue and its correlation with lung injury during ANP.Methods:Ninety SD rats were divided into three groups randomly. Acute edematous pancreatitis (AEP) and ANP were induced in two groups of rats by retrograde infusion of sodium taurochoiate (0.5% and 5%) into biliopancreatic duct respectively. Animals in the last group only underwent sham-operation. All the animals were sacrificed at certain time, plasma level of amylase and TNF-α was assessed, while intrapulmonary expression of TNF-α protein was detected by immunohistochemistry technique. Moreover, cell apoptosis was studied in paraffin embedding puhnonary sections with TUNEL method accompanied with laser scanning confocal microscope.Results:Some lymphocytes in bronchial lymph nodes and fibroblasts in interstitial tissue presented as apoptosis in sham-operation group. It is noted that the number of apoptotic cell in certain field of section under microscope did not change markedly with prolonged time after acute pancreatitis was induced either in AEP group or ANP group, but apoptotic cells involved little inflammatory cell, alveolar epithelial Cell and vascular endothelial cell. Taking account in the number of apoptotic cell per 1?000 cells on pulmonary sections, so called apoptosis index (‰), it appeared to decrease tran siently in AEP group. However, it' decreased continuously in ANP group, less than the corresponding value of AEP group significantly since 6 hour after the model was induced ( P <0. 05). Statistic analysis indicate that the change of apoptosis index had a negative correlation with TNFα in plasma and histopathological score in the two groups respectively, as well as the elevated level of plasma endotoxin in ANP group ( P <0.05). Conclusion: We found that the number of alveolar epithelial cell and vascular endothelial cell undergoing apoptosis is small even in ANP group, so this phenomenon is almost impossible to cause pulmonary dysfunction seen in ANP. Apoptosis is rarely detected in a large number of inflammatory cells, in which neutrophils'are the main part, aggregating in lung tissue during ANP, this is the cause of marked reduction of apoptosis index. It is possible for many activated neutrophils in which apoptosis is delayed to play an important harmful role in pathogenesis of lung injury. Increased production of TNFα and endotoxin in blood may partially explain the significantly delayed apoptosis of neutrophils found in lung tissue after ANP is induced.