详细解读多篇高分SCI文献中2023国自然热点PANoptosis/泛 ...

福奇福齐

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<hr/>通过昨天PANoptosis/泛凋亡的科普文紧跟2023年国自然热点之PANoptosis/泛凋亡，一种集焦亡+凋亡+坏死的炎症性细胞死亡方式，相比大家已经了解了什么是泛凋亡，在此基础上，很多小伙伴就会问了，在具体的实验设计中，应该怎么样去研究/检测PANoptosis？
不要着急，今天我们就来聊一聊这个问题，我们主要通过几篇高分文献的相关结果展示和结果描述来看
示例1：Karki R et al. ADAR1 restricts ZBP1-mediated immune response and PANoptosis to promote tumorigenesis. Cell Rep. 2021 Oct 19;37(3):109858. doi: 10.1016/j.celrep.2021.109858. IF=9.995


目的：核输出抑制剂(NEIs)联合干扰素(IFNs)激活PANoptosis
处理：两种核输出抑制剂(NEIs)：leptomycin B (LMB) or Selinexor (KPT-330)
      两种INF因子：IFN-β or IFN-γ
主要结果描述：

• Treatment with either KPT-330 or LMB for 24 h induced a low level of cell death in BMDMs；Treatment with IFN-β or IFN-γ in combination with KPT-330 or LMB increased the incidence of cell death compared with treatment with KPT-330 or LMB alone (Figures 1A-C) 【KPT-330或 LMB处理24导致BMDM细胞低水平的细胞死亡，而加入IFN-β或IFN-γ增加了细胞死亡的发生；A，B，C三组分别是不同方法检测的细胞死亡，包括PI染色，活细胞自动化系统，LDH释放】
  
• In line with the incidence of cell death, treatment with IFN-β or IFN-γ potentiated KPT-330- or LMB-induced cleavage of caspase-1 and GSDME (pyroptosis); cleavage of caspase-8, −3, and −7 (apoptosis); and phosphorylation of MLKL (necroptosis) (Figures 1D–1F). Collectively, these data suggest that combining IFNs with NEIs sensitizes the cells to undergo inflammasome activation and cell death involving the components of pyroptosis, apoptosis, and necroptosis, indicating that PANoptosis is occurring. 【根据细胞死亡的发生率，NEIs联合IFNs处理，诱导caspase-1和GSDME的裂解(焦亡)；caspase-8、−3和−7的裂解(凋亡);MLKL磷酸化(坏死)，这些数据表明IFNs与NEIs结合可致敏细胞发生炎性小体激活和细胞死亡，涉及焦亡、凋亡和坏死等成分，表明PANoptosis正在发生。】
  

详细的结果描述参考下图：


示例2：Karki R et al. Interferon regulatory factor 1 regulates PANoptosis to prevent colorectal cancer. JCI Insight. 2020 Jun 18;5(12):e136720. doi: 10.1172/jci.insight.136720. IF=9.484


目的：IRF1是否在暴露于AOM/DSS后调节PANoptosis
主要结果：

• We observed a reduced number of TUNEL+ cells in the colons of Irf1–/– mice 14 and 80 days after AOM injection compared with those of WT mice, whereas no difference was observed in untreated (day 0) WT and Irf1–/– mice (Figure 5A).【注射AOM后14天和80天，Irf1–/–小鼠结肠中TUNEL+细胞数量与WT小鼠相比有所减少，而未处理(第0天)的WT和Irf1–/–小鼠结肠中TUNEL+细胞数量没有差异】
  
• In line with reduced cell death in the colons of Irf1–/– mice, we observed defective CASP3 and CASP7 activation in the colons of Irf1–/– mice 14 and 80 days after AOM injection with respect to WT mice (Figure 5B-C), suggesting that IRF1 plays a role in apoptosis under these conditions. 【与Irf1–/–小鼠结肠细胞死亡减少一致，AOM注射后14天和80天，Irf1–/–小鼠结肠中CASP3和CASP7激活缺陷，提示Irf1在细胞凋亡中起作用】
  
• We found reduced expression and activation of the pyroptosis executioner, GSDMD, in the colons of mice lacking IRF1 (Figure 5D), suggesting that IRF1 regulates pyroptosis.【在敲除IRF1的小鼠结肠中，焦亡驱动因子GSDMD的表达和激活减少，表明IRF1调节焦亡】
  
• Colons of mice lacking IRF1 showed reduced expression of MLKL compared with the colons of WT mice (Figure 5D). In addition, we observed reduced cell death in organoids (Figure 5E) derived from Irf1–/– mice as compared with those of WT mice in response to the necroptotic trigger TNF + zVAD, further supporting the role of IRF1 in necroptosis.【与WT小鼠相比，敲除IRF1的小鼠结肠中MLKL的表达减少。此外，与WT小鼠相比，来源于响应坏死触发因子TNF+zVAD的Irf1–/–小鼠的类器官细胞死亡减少，进一步支持了IRF1在坏死中的作用】
  

示例3：Bi Y et al. FUNDC1 protects against doxorubicin-induced cardiomyocyte PANoptosis through stabilizing mtDNA via interaction with TUFM. Cell Death Dis. 2022 Dec 5;13(12):1020. IF=9.685


目的：FUNDC1缺失促进dox诱导的心肌细胞PANoptosis
分组：WT-Vehicle; WT-DOX; FUNDC1–/– -Vehicle; FUNDC1–/– -DOX
主要结果：

• PANoptosis is driven through a multiprotein complex activated by the DNA sensers AIM2, ZBP1, and Pyrin to form a flexible skeleton to recruit Caspase1, Caspase3, Caspase8, RIPK1, and RIPK3, ultimately resulting in the execution of concurrent pyroptosis, apoptosis and necroptosis (Fig. 5a) 【PANoptosis是由DNA传感器AIM2、ZBP1和Pyrin激活的多蛋白复合物驱动，形成一个灵活的骨架，招募Caspase1、Caspase3、Caspase8、RIPK1和RIPK3，最终导致同时发生pyroptosis、凋亡和necroptosis；作者首先画了一个简单的示意图展示PANoptosis的机制】
  
• Next, we examined PANoptosis in DOX challenged heart tissues in mice. DOX boosted PANoptosis in mouse hearts as evidenced by increased levels of AIM2, ZBP1, Pyrin (members of PANoptosome) (Fig. 5b–d), active forms of Caspase1 and GSDMD (pyroptosis markers) (Fig. 5e, f), active forms of Caspase3 and Caspase8 (apoptosis markers) (Fig. 5g, h), and phosphorylation of MLKL, RIPK1, RIPK3 (necroptosis markers) (Fig. 5i–k), the effects of which were aggravated by FUNDC1 ablation with little effect from FUNDC1 deficiency itself (Fig. 5b–k).【DOX增强了小鼠心脏的泛凋亡，表现为AIM2、ZBP1、Pyrin (PANoptosome成员)表达、Caspase1和GSDMD (pyroptosis标记物)的活性形式、Caspase3和Caspase8(凋亡标记物)的活性形式以及MLKL、RIPK1、RIPK3 (necroptosis标记物)的磷酸化增加， FUNDC1缺失加重了这些作用，而FUNDC1缺失本身影响不大。】
  

根据这3篇文章结果展示，可以总结出PANoptosis相关的检测包括：

• 检测细胞死亡
  
• 检测泛凋亡复合体PANoptosome成员的表达水平，例如AIM2, ZBP1, Pyrin
  
• 检测泛凋亡标记物，包括焦亡markers(Caspase1, Caspase11, GSDMD, GSDME)，凋亡markers (Caspase3, Caspase7, Caspase8)，坏死markers (MLKL、RIPK1、RIPK3)
  

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