【摘要】 目的探讨肿瘤坏死因子相关凋亡诱导配体(TRAIL)及其受体(TRAILR)信号转导通路在煤工尘肺(CWP)患者肺泡巨噬细胞(AM)凋亡中的作用机制。方法以接触煤尘及煤矽尘的24例观察对象和CWP壹期、贰期患者各22例为研究对象,收集大容量肺灌洗回收液,分离、纯化AM后,分为对照组、超氧化物歧化酶(SOD)组、TRAIL组、抗-TRAIL抗体组、天冬氨酸特异性半胱氨酸蛋白酶(Caspase)-8抑制组、核转录因子(NF)-κB抑制剂组,给予相应处理后进行培养。收获细胞后,检测细胞凋亡情况,并提取蛋白采用免疫印迹法检测TRAIL/TRAILR信号转导通路Caspase-3、Caspase-8和死亡受体5(DR5)蛋白相对表达水平。结果 5个处理组AM凋亡指数均低于对照组(P<0.05),TRAIL组AM凋亡指数分别高于SOD组、抗-TRAIL抗体组、Caspase-8抑制剂组和NF-κB抑制剂组(P<0.05);CWP壹期和贰期患者AM凋亡指数均高于观察对象(P<0.05),CWP贰期患者AM凋亡指数高于CWP壹期患者(P<0.05);不同处理和不同程度患者AM凋亡指数的交互作用有统计学意义(P<0.01)。5个处理组Caspase-3、Caspase-8和DR5的蛋白相对表达水平分别低于对照组(P<0.05),TRAIL组Caspase-3和Caspase-8的蛋白相对表达水平分别高于SOD组、抗-TRAIL抗体组、Caspase-8抑制剂组和NF-κB抑制剂组(P<0.05),SOD组DR5蛋白相对表达水平分别低于TRAIL组、抗-TRAIL抗体组、Caspase-8抑制剂组和NF-κB抑制剂组(P<0.05);CWP壹期和贰期患者Caspase-3、Caspase-8和DR5的蛋白相对表达水平均高于观察对象(P<0.05);不同处理和不同程度患者Caspase-3的蛋白相对表达水平的交互作用有统计学意义(P<0.01),不同处理和不同程度患者Caspase-8和DR5的蛋白相对表达水平的交互作用均无统计学意义(P>0.05)。结论 TRAIL/TRAILR信号转导通路介导的AM凋亡可能在CWP发病中发挥一定作用;在不同环节对TRAIL/TRAILR信号转导通路进行抑制或阻断,可有效抑制CWP患者AM凋亡的发生。更多还原

【Abstract】 Objective To study the mechanism of tumor necrosis factor-related apoptosis-inducing ligand( TRAIL) and its receptors( TRAILR) signal transduction pathways in the apoptosis of alveolar macrophage( AM),which were collected from coal workers’ pneumoconiosis( CWP). Methods Twenty-four observed subjects exposed to coal dust and coal silica dust,22 cases of CWP at stage Ⅰ,and 22 cases of CWP at stage Ⅱ were chosen as the subjects. The bronchoalveolar lavage fluids of whole-lung lavage of all subjects were collected. AM were separated and purified. Then they were divided into 6 groups: a control group,a superoxide dismutase( SOD) group,a TRAIL group,an anti-TRAIL antibody group,a Caspase-8 suppression group and a nuclear factor-κB( NF-κB) suppression group. The AM of 6 groups with corresponding treatment were cultivated. After the cells were harvested,apoptosis were detected,proteins were extracted by Western blotting and relative expression levels of Caspase-3,Caspase-8 and death receptor 5( DR5) protein in the TRAIL / TRAILR signal transduction pathway were detected. Results The apoptotic indexes of AM in 5 treatment groups were lower than that of the control group( P < 0. 05). The apoptotic index of AM in TRAIL group was higher than those of other groups except the control group( P < 0. 05). The apoptotic indexes of AM in CWP patients at stage Ⅰ and stage Ⅱ were significantly higher than those of the observed subjects respectively( P < 0. 05). The apoptotic indexe of AM in CWP patients at stageⅡ was significantly higher than that of CWP patients at stageⅠ( P < 0. 05). The interaction of different treatment and different degree patients had significant difference in apoptotic indexe of AM( P < 0. 01). The protein relative expression levels of Caspase-3,Caspase-8 and DR5 in 5 treatment groups were significantly lower than those of the control group respectively( P < 0. 05). The protein relative expression levels of Caspase-3 and Caspase-8 in TRAIL group were significantly higher than those of other groups except the control group( P < 0. 05). The protein relative expression levels of DR5 in SOD group were significantly lower than those of other groups except the control group( P < 0. 05). The protein relative expression levels of Caspase-3,Caspase-8 and DR5 in CWP patients at stage Ⅰ and stage Ⅱ were significantly higher than those of the observed subjects( P < 0. 05); the relative expression levels of protein Caspase-3 in different treatments and different degree patients had statistical significant difference in interaction( P < 0. 01),while the protein relative expression levels of Caspase-8 and DR5 in different treatments and different degree patients had no statistical significant difference in interaction( P > 0. 05). Conclusion AM apoptosis mediated by TRAIL / TRAILR signal transduction pathway may play a certain role in the occurrence and development of CWP. The TRAIL / TRAILR signal transduction pathways inhibited or blocked at different stages can effectively inhibit the occurrence of AM apoptosis.更多还原