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目的观察氯化钆(GdCl3)诱导急性坏死性胰腺炎(ANP)肺泡巨噬细胞(AM)凋亡时核因子-κB(NF-κB)表达情况,探讨NF-κB在其中的作用及机制。方法36只SD大鼠随机分为正常对照组,ANP组,GdCl3治疗组。逆行性胰胆管注射5%牛磺胆酸钠建立ANP大鼠模型,正常对照组大鼠以同法注入生理盐水,GdCl3治疗组制模后即刻自阴茎背静脉注射GdCl3。各组大鼠于成模后6 h经支气管肺泡灌洗获取AM,检测支气管肺泡灌洗液(BALF)中TNF-α和IL-1β含量及肺组织髓过氧化物酶(MPO)水平。用琼脂糖凝胶电泳、流式细胞仪检测AM的凋亡情况;Western-blot检测AM中NF-κB活性。同时对肺组织行病理学检查。结果ANP组TNF-α和IL-1β含量高于对照组(P<0.05),而治疗组显著低于ANP组(P<0.05)。仅治疗组DNA电泳见典型的细胞凋亡的梯状条带。对照组,ANP组,治疗组AM凋亡率分别为(10.81±0.86)%,(6.47±1.52)%,(17.41±3.36)%,3组间差异均有统计学意义(P<0.05);Western-blot检测3组AM中NF-κB表达的相对灰度值分别为(0.80±0.05),(1.96±0.15),(1.42±0.10),3组间差异亦有统计学意义(P<0.05)。AM的凋亡率与NF-κB活性呈负相关(r=-0.554,P<0.01)。结论GdCl3可能通过抑制NF-κB的活化诱导大鼠ANP肺泡巨噬细胞凋亡,从而减轻肺损伤。
Objective To observe the expression of nuclear factor-κB (NF-κB) in GdCl3-induced apoptosis of alveolar macrophages (AM) in acute necrotizing pancreatitis (ANP) and to explore the role and mechanism of NF-κB in it. Methods Thirty - six SD rats were randomly divided into normal control group, ANP group and GdCl3 treatment group. The rat model of ANP was established by injecting 5% sodium taurocholate into the pancreaticobiliary duct. Rats in the normal control group were injected normal saline with the same method. GdCl3 was injected into the dorsal venous of the penis immediately after the model was established. Rats in each group were given AM by bronchoalveolar lavage 6 h after injection, and the content of TNF-α and IL-1β and the level of myeloperoxidase (MPO) in bronchoalveolar lavage fluid (BALF) were measured. The apoptosis of AM was detected by agarose gel electrophoresis and flow cytometry. The NF-κB activity in AM was detected by Western-blot. At the same time, pathological examination of lung tissue. Results The levels of TNF-α and IL-1β in ANP group were significantly higher than those in control group (P <0.05), while those in treatment group were significantly lower than those in ANP group (P <0.05). Electrophoresis of the DNA alone in the treatment group shows a typical ladder of apoptotic bands. The apoptotic rates of AM in the control group, ANP group and treatment group were (10.81 ± 0.86)%, (6.47 ± 1.52)% and (17.41 ± 3.36)%, respectively. There were significant differences among the three groups (P <0.05) The relative gray values of NF-κB in three groups were (0.80 ± 0.05) and (1.96 ± 0.15) and (1.42 ± 0.10) respectively by Western-blot. There was also significant difference between the three groups (P <0.05 ). The apoptotic rate of AM was negatively correlated with the activity of NF-κB (r = -0.554, P <0.01). Conclusion GdCl3 may induce apoptosis of ANP-alveolar macrophages and inhibit lung injury by inhibiting the activation of NF-κB.