基于TIPE2抑制的PCNP上调改善感染后膈肌无力的分子机制

The key causes for respiratory failure(RF) were generally from lung-related diseases. Recent studies demonstrated that diaphragmatic weakness(DW) was also involved in RF.The sepsis was the main reason of diaphragmatic weakness. Nevertheless the mechanism of DW by sepsis was elusive. . Our preliminary data showed that TIPE2 upregulation and then PCNP down-regulation were found in JNK1 inhibition by dominant negative JNK1 expression in C2C12 cells with LPS exposure. Along with our previous results found in C2C12，rats and mice models exposed with LPS, we proposed a hypothesis that PCNP plays a key role in DW in sepsis. . PCNP-over expression and silencing were separately done in C2C12 treated with LPS.PCNP protein was used in rats model of sepsis exposed to LPS and force generation of diaphragm was measured.PCNP KO mice by our group were ready for use in the similar model to the above-mentioned rats. . We will confirm that sepsis result in PCNP decrease mediated by TIPE2 increase. PCNP interacted with JNK1 will cause caspase activation and the following myosin degradation in diaphragm, the true etiology of DW in sepsis. Thus PCNP will be benificial to improve the ventilation in RF.

近来证实膈肌无力（DW）参与呼吸衰竭，且感染是DW的主因，感染后DW与JNK1等信号通路有关。前期发现LPS处理后骨骼肌C2C12细胞系PCNP降低,JNK1和免疫负调控因子TIPE2上调。为探讨PCNP在感染后DW中的作用，运用基因沉默和过表达技术，了解PCNP对肌细胞生长、凋亡、收缩蛋白、细胞周期等的影响；分别用大鼠LPS感染和盲肠结扎穿孔感染模型，探讨补充PCNP对DW的影响；用新培育的条件性基因敲除PCNP小鼠，验证感染后PCNP缺失对膈肌肌力的影响。预期确定感染抑制膈肌PCNP表达，通过JNK1通路激活Caspase 8，促进肌球蛋白降解（而非肌细胞凋亡），进而降低膈肌肌力；且PCNP的抑制作用受到上游TIPE2的调控。本项目率先培育出PCNP基因敲除小鼠，确认PCNP与感染后膈肌肌力降低有关。有助于探明ICU病人呼衰的分子机制;使用PCNP及其靶点药物有望改善感染致DW。

近年来证实膈肌无力（diaphragmatic weakness,DW）参与呼吸衰竭的发生与发展，且感染是DW的主因，感染后DW与JNK1等信号通路有关。前期发现细菌内毒素LPS处理后骨骼肌C2C12细胞系中新型核蛋白PCNP（PEST proteolytic signal-containing nuclear protein）降低,JNK1和免疫负调控因子TIPE2则上调。本研究探讨了PCNP在感染后DW中的作用，通过运用基因沉默和过表达技术，了解PCNP对肌细胞生长、凋亡、收缩蛋白、细胞周期等的影响；分别用大鼠LPS感染和盲肠结扎穿孔感染模型，探讨补充PCNP对DW的影响；用新培育的条件性基因敲除PCNP小鼠，验证感染后PCNP缺失对膈肌肌力的影响。结果初步确定感染可以抑制膈肌细胞PCNP表达，通过JNK1通路激活Caspase 8，促进肌球蛋白降解（而非肌细胞凋亡），进而降低膈肌肌力；且PCNP的抑制作用受到上游TIPE2的调控。本项目的意义在于率先培育出PCNP基因敲除小鼠，为确认PCNP与感染后膈肌肌力降低有关奠定了基础。未来经过深入研究，将有助于探明基于PCNP的ICU病人呼衰的分子机制;并有望使用PCNP及其靶点药物去改善感染致DW。本项目部分结果已经发表在 Oxidative Medicine and Cellular Longevity(SCI II, IF4.59)，申报了相关的国家发明专利，培养了四届硕士研究生，招收了3届外籍留学生，1届外籍博士后，于2017年获批了河南省核蛋白基因调控国际联合实验室。