内毒素／脂多糖通过PIM3对人中性粒细胞早期凋亡的作用  

Effects of endotoxin/lipopolysaccharide on early apoptosis of human neutrophil through PIM3

作　　者：宋明明 李平[1] 金新源[1] 苏建东[1] 孙炳伟 Song Mingming;Li Ping;Jin Xinyuan;Su Jiandong;Sun Bingwei(Department of Burns and Plastic Surgery,Suzhou Hospital Affiliated to Nanjing Medical University,Suzhou 215002,China)

机构地区：[1]南京医科大学附属苏州医院烧伤整形科,215002

出　　处：《中华烧伤杂志》2018年第11期809-814,共6页Chinese Journal of Burns

基　　金：国家自然科学基金（81471903）;江苏省社会发展重点项目（BE2017695）;苏州市医学重点学科基金（SZXK201511）;苏州科技发展计划（SYS2018086）

摘　　要：目的探讨内毒素／脂多糖（LPS）通过PIM3对人中性粒细胞早期凋亡的影响。方法采集1名健康成年志愿者静脉血，分离出中性粒细胞，采用随机数字表法分为对照组、LPS组、LPS＋PIM447组。对照组不行任何处理，LPS组采用1μLμg／mL的LPS刺激，LPS＋PIM447组在采用上述LPS刺激的前30min，用1μL终物质的量浓度为5μmol／L的PIM447预干预30min。常规培养1h后，采用流式细胞仪检测细胞早期凋亡率，二氢罗丹明123荧光探针法检测细胞的活性氧生成水平，蛋白质印迹法检测细胞内PIM3的表达水平。常规培养2h后，琼脂糖细胞趋化模型检测细胞趋化距离。以上4个实验每组样本数均为5。对数据行单因素方差分析、SNK检验。结果（1）LPS组细胞早期凋亡率[（0．891±0．012）％]与对照组[（1．351±0．183）％]相近（P〉0．05），LPS＋PIM447组细胞早期凋亡率[（82．057±0．121）％]明显高于LPS组（P〈0．01）。（2）LPS组细胞趋化距离[（984±5）μm]明显小于对照组[（2241±7）μm，P〈0．01]，LPS＋PIM447组细胞趋化距离[（1785±11）μm]明显大于LPS组（P〈0．05）。（3）LPS组细胞活性氧生成水平较对照组明显增加（P〈0．05），LPS＋PIM447组细胞活性氧生成水平较LPS组明显降低（P〈0．05）。（4）LPS组细胞PIM3表达水平（1．297±0．015）较对照组（0．789±0．021）明显增加（P〈0．05），LPS＋PIM447组细胞PIM3表达水平（0．731±0．011）较LPS组明显降低（P〈0．05）。结论LPS刺激可以降低人中性粒细胞的早期凋亡，使用PIM447预干预可以明显增加LPS刺激后中性粒细胞的早期凋亡，恢复趋化功能，且很好地抑制活性氧的产生，其机制可能与LPS促进PIM3的表达有关。Objective To explore the effects of endotoxin/lipopolysaccharide （LPS） on early apop-tosis of human neutrophil through PIM3. Methods Venous blood samples were collected from a healthy adult volunteer to isolate neutrophils. The neutrophils were divided into control group, LPS group, and LPS ＋ PIM447 group according to the random number table. No treatment was given to the ceils in control group. The cells in LPS group underwent LPS stimulation （ 1 μL, 1 μg/mL）. The cells in LPS ＋ PIM447 group un-derwent PIM447 （1 μL, final amount-of-substance concentration of 5 μmol/L） intervention 30 min before having the same LPS stimulation as that in LPS group. After conventional culture for 1 h, the early cell ap-optosis rate was determined with flow cytometer; the generation level of reactive oxygen species （ROS） was assessed with dihydrogenrhodamine 123 fluorescent probe staining method; and the level of PIM3 was detec-ted by Western blotting. After conventional culture for 2 h, the cell chemotaxis distance was measured by agarose chemotaxis cell model. The sample numbers of each group in the 4 experiments were all 5. Data were processed with one-way analysis of variance and Student-Newman-Keuls test. Results （1） The ear-ly apoptosis rate of cells in LPS group [ （0. 891 ± 0. 012）% ] was close to that in control group [ （1. 351 ± 0.183）%, P 〉0.05）]. The early apoptosis rate of cells in LPS＋PIM447 group [（82.057 ±0.121）%] was higher than that in LPS group （ P 〈 0.01）. （2） The cell chemotaxis distance of cells in LPS group [ （984 ± 5） μm] was significantly shorter than that in control group [ （2 241 ± 7） μm, P 〈 0. 01 ]. The cell chemotaxis distance of cells in LPS ＋ PIM447 group [ （ 1 785 ± 11 ） μm ] was significantly longer than that in LPS group （ P 〈 0.05）. （3） The generation level of ROS in cells of LPS group was significantly higher than that in control group （ P 〈 0.05）. The generation level of ROS in cells of LPS ＋ PIM44

关 键 词：脓毒症 脂多糖类 细胞凋亡 中性粒细胞 PIM3 

分 类 号：R392[医药卫生—免疫学]