Objective

Programmed death 1 (PD-1) and macrophages are the most intriguing candidates in sepsis-induced inflammatory disorders. We aimed to investigate the association between monocyte PD-1 and sepsis severity and the mechanism by which blocking macrophage-associated PD-1 causes inflammatory disorders in sepsis.

Materials and methods

We first measured whether the expression of PD-1 on the monocyte subset is clinically associated with sepsis severity in an observational study. This study included 42 septic patients and 16 healthy controls (HCs) whose serum inflammatory factors were examined by Luminex MagPix. Then, we investigated the effect of PD-1 blockade on macrophages from septic mice (C57BL/6 mice) constructed by caecal ligation and puncture (CLP) via RNA sequencing. The positive genes screened by RNA-seq were verified in LPS-stimulated RAW264.7 cells by Western blot.

Results

The results showed that the expression of PD-1 on CD14⁺CD16⁺ monocytes (intermediate monocytes, IM Mo) was significantly higher in both septic and septic shock patients than in HCs. Further analysis of serum cytokines in septic patients showed that the levels of IL-6 and TNF-α were significantly higher than those in HCs, while serum PD-1 levels were decreased in septic patients. More interestingly, blockade of PD-1 on macrophages from septic mice suppressed the gene expression levels of NLRP3/Caspase-4/AKT2/STAT3. The protein levels associated with pyroptosis including NLRP3, Caspase4, GSDMD and NT-GSDMD were significantly decreased in LPS-stimulated RAW264.7 cells treated with PD-1 antibody.

Conclusion

Our results suggested that intermediate monocytes with high expression of PD-1 may be involved in the progression of sepsis. PD-1 might play a critical role in regulating the pyroptosis signalling pathway in sepsis.

中文翻译：

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巨噬细胞相关程序性死亡1的阻断抑制脓毒症中的焦亡信号通路

客观的

程序性死亡 1 (PD-1) 和巨噬细胞是脓毒症引起的炎症性疾病中最有趣的候选者。我们旨在研究单核细胞 PD-1 与脓毒症严重程度之间的关联，以及阻断巨噬细胞相关 PD-1 导致脓毒症炎症性疾病的机制。

材料和方法

我们首先在一项观察性研究中测量了单核细胞亚群上 PD-1 的表达是否在临床上与脓毒症严重程度相关。这项研究包括 42 名脓毒症患者和 16 名健康对照 (HC)，他们的血清炎症因子通过 Luminex MagPix 进行了检测。然后，我们通过 RNA 测序研究了 PD-1 阻断对通过盲肠结扎和穿刺 (CLP) 构建的脓毒症小鼠 (C57BL/6 小鼠) 巨噬细胞的影响。通过蛋白质印迹在 LPS 刺激的 RAW264.7 细胞中验证 RNA-seq 筛选的阳性基因。

结果

结果表明，感染性休克和感染性休克患者的 CD14 ⁺ CD16 ⁺单核细胞（中间单核细胞，IM Mo）上 PD-1 的表达明显高于 HCs。对脓毒症患者血清细胞因子的进一步分析表明，脓毒症患者的 IL-6 和 TNF-α 水平显着高于 HCs，而血清 PD-1 水平降低。更有趣的是，阻断脓毒症小鼠巨噬细胞上的 PD-1 抑制了 NLRP3/Caspase-4/AKT2/STAT3 的基因表达水平。在用 PD-1 抗体处理的 LPS 刺激的 RAW264.7 细胞中，与细胞焦亡相关的蛋白质水平（包括 NLRP3、Caspase4、GSDMD 和 NT-GSDMD）显着降低。

结论

我们的研究结果表明，PD-1 高表达的中间单核细胞可能参与脓毒症的进展。PD-1 可能在脓毒症细胞焦亡信号通路的调节中发挥关键作用。