Ameliorative effect of Xuebijing injection on acute lung injury in sepsis by interfering with cGAS/STING pathway
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摘要:
探讨血必净注射液（Xuebijing injection，XBJ）干预cGAS/STING通路减轻脓毒症诱导的急性肺损伤（acute lung injury，ALI）的作用。采用盲肠结扎与穿刺（CLP）建立小鼠脓毒症ALI模型，LPS刺激RAW264.7细胞构建细胞炎症模型。利用HE染色、ELISA法、Western blot等方法，考察XBJ对脓毒症小鼠肺组织损伤与cGAS/STING通路相关蛋白表达的影响。结果发现，在体内，XBJ干预能够缓解肺组织损伤，降低血清IL-6、TNF-α、IFN-β、IL-1β水平，肺组织cGAS、STING、p-TBK1和p-IRF3蛋白的表达。在体外，XBJ干预后能够降低RAW264.7细胞相关炎症因子mRNA水平，以及cGAS/STING通路蛋白的表达。结果表明，XBJ能够通过抑制cGAS/STING通路活化，抑制炎症反应而发挥防治脓毒症ALI的作用。本研究为临床XBJ防治脓毒症ALI提供新的分子机制。
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关键词:
- 脓毒症 /
- 急性肺损伤 /
- 血必净注射液 /
- cGAS/STING通路
Abstract:To investigate the effect of Xuebijing injection (XBJ) on cGAS/STING pathway in alleviating sepsis-induced acute lung injury (ALI), the mouse sepsis-induced ALI model was established by cecal ligation and puncture (CLP), and the cell inflammation model was constructed by LPS stimulating RAW264.7 cells. The effects of XBJ on lung tissue injury and cGAS/STING pathway-related protein expression in septic mice were investigated by HE staining, ELISA, and Western blot. The results showed that XBJ intervention could alleviate lung tissue injury, reduce serum IL-6, TNF-α, IFN-β, IL-1-β levels, and the expression of cGAS, STING, p-TBK1, and p-IRF3 proteins in lung tissue in vivo, and reduce the mRNA level of related inflammatory factors in RAW264.7 cells and the expression of cGAS/STING pathway proteins in vitro. The results showed that XBJ could play a role in the prevention and treatment of sepsis-induced ALI by inhibiting the inflammatory response via inhibition of the activation of cGAS/STING pathway. This study provides a new molecular mechanism for the clinical prevention and treatment of sepsis-induced acute lung injury with XBJ.
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Keywords:
- sepsis /
- acute lung injury /
- Xuebijing injection /
- cGAS/STING pathway
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Figure 1. Xuebijing injection (XBJ) alleviates lung tissue injury caused by sepsis-induced acute lung injury（ALI） mice
A: Histopathological changes in the lung measured by HE staining (200 ×); B: Comparison of lung injury scores ($ \overline{\text{x}} \text{ ± } \text{s} $, n = 3); C: Lung wet/dry (W/D) ratio was determined at 24 h after cecal ligation and puncture (CLP) ($ \overline{\text{x}} \text{ ± } \text{s} $, n = 8); D: Myeloperoxidase (MPO) activity in lung tissue were determined at 24 h after CLP ($ \overline{\text{x}} \text{ ± } \text{s} $, n = 8) Dex：Dexamethasone. ###P < 0.001 vs sham group； *P < 0.05, **P < 0.01, ***P < 0.001 vs model group
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Figure 2. XBJ reduced the level of inflammatory factors in serum of sepsis-induced ALI mice ($ \overline{\text{x}} \text{ ± } \text{s} $, n = 6)
A-D: Levels of IL-1β (A), IL-6 (B), TNF-α (C) and IFN-β (D) of serum in CLP-induced mice were determined by ELISA ###P < 0.001 vs sham group；*P < 0.05, **P < 0.01, ***P < 0.001 vs model group
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Figure 3. XBJ (4 mL/kg) inhibits cGAS/STING pathway activation in lung tissue of sepsis ALI mice
A-B: Western blot of cGAS (A), STING (B)($ \overline{\text{x}} \text{ ± } \text{s} $, n = 6); C: cGAS and STING level in immunohistochemical staining (200 ×); D-E: Percentage contribution of positive of cGAS (D) and STING (E) was quantified by Image J ($ \overline{\text{x}} \text{ ± } \text{s} $, n = 3); F-G: Western blot of p-TBK1 (F) and p-IRF3 (G) ($ \overline{\text{x}} \text{ ± } \text{s} $, n = 6) ###P < 0.001 vs sham group；*P < 0.05, **P < 0.01, ***P < 0.001 vs model group
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Figure 4. XBJ inhibits LPS-induced production of inflammatory cytokines in RAW264.7 cells ($ \overline{\text{x}} \text{ ± } \text{s} $, n = 6)
A-D: Level of IL-1β (A), IL-6 (B), TNF-α (C) and IFN-β (D) were determined by RT-qPCR ###P < 0.001 vs control group；**P < 0.01, ***P < 0.001 vs model group
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