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Oroxylin A-induced Trained Immunity Promotes LC3-associated Phagocytosis in Macrophage in Protecting Mice Against Sepsis

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Abstract

Sepsis is defined as a dysregulated host response to infection that leads to multiorgan failure. Innate immune memory, i.e., “trained immunity”, can result in stronger immune responses and provide protection against various infections. Many biological agents, including β-glucan, can induce trained immunity, but these stimuli may cause uncontrolled inflammation. Oroxylin A (OA) is an active flavonoid compound that is derived from Scutellaria baicalensis. OA is an agonist for inducing trained immunity in vivo and in vitro, and β-glucan was used as a positive control. The protective effects of OA-induced trained immunity were evaluated in mouse models that were established by either lipopolysaccharide (LPS) administration or caecal ligation and puncture (CLP). The expression of inflammatory factors and signaling pathway components involved in trained immunity was evaluated in vitro using qRT‒PCR, western blotting (WB) and enzyme-linked immunosorbent assay (ELISA). Flow cytometry and confocal microscopy were used to examine reactive oxygen species (ROS) levels and phagocytosis in trained macrophages. A PCR array was used to screen genes that were differentially expressed in trained macrophages. Here, we revealed that OA alleviated sepsis via trained immunity. OA-treated macrophages displayed increased glycolysis and mTOR phosphorylation, and mTOR inhibitors suppressed OA-induced trained immunity by effectively reprogramming macrophages. The PCR array revealed key genes in the mTOR signaling pathway in OA-treated macrophages. Furthermore, OA targeted the Dectin-1-syk axis to promote LC3-associated phagocytosis (LAP) by trained macrophages, thereby enhancing the ability of these macrophages to protect against infection. This ability could be transferred to a new host via the adoptive transfer of peritoneal macrophages. This study is the first to provide new insights into the potential of OA-induced trained immunity to be used as a strategy to protect mice against sepsis by promoting LAP by macrophages.

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Data Availability

No datasets were generated or analysed during the current study.

Abbreviations

AST:

Aspartate aminotransferase

ALT:

Alanine aminotransferase

CLL:

Clodronate liposomes

CLP:

Cecal ligation and puncture

CMC-Na:

Carboxymethyl cellulose-Na

DMSO:

Dimethyl sulfoxide

ELISA:

Enzyme-linked immunosorbent assay

H&E:

Hematoxylin and eosin staining

IHC:

Immunohistochemistry

LPS:

Lipopolysaccharid

LAP:

LC3-associated phagocytosis

OA:

Oroxylin A

PFA:

Paraformaldehyde

ROS:

Reactive oxygen species

WB:

Western blotting

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Funding

This work was supported by the National Key R&D Program of China (2023YFC2308200) and National Natural Science Foundation of China Special Project [T2341015].

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Authors and Affiliations

  1. The State Key Laboratory of Pharmaceutical Biotechnology, Division of Immunology, Medical School, Nanjing University, Nanjing, 210093, China

    Lijie Yin, Ziqian Bing, Yaojun Zheng, Yue Dong, Jiali Wang, Renjie Luo, Yue Zhao, Huan Dou & Yayi Hou

  2. Jiangsu International Laboratory of Immunity and Metabolism, The Department of Pathogenic Biology and Immunology, Xuzhou Medical University, Xuzhou, 221004, China

    Yuchen Pan

  3. Jiangsu Key Laboratory of Molecular Medicine, Nanjing, 210093, China

    Yue Zhao, Huan Dou & Yayi Hou

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  1. Lijie Yin
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  2. Ziqian Bing
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  4. Yuchen Pan
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  5. Yue Dong
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Contributions

LY and YH conceived this idea. LY and ZB performed the experiments in vitro. JZ, YP, YD, JW and RL performed the experiments in vivo. YL participated in the collection, analysis, and interpretation of the data. YL wrote the manuscript. YZ and HD revised it critically for important intellectual content. YH approved the version to be published and agreed to be accountable for all aspects of the work in ensuring that questions related to the accuracy or integrity of any part of the work are appropriately investigated and resolved. All authors read and approved the final manuscript.

Corresponding authors

Correspondence to Yue Zhao, Huan Dou or Yayi Hou.

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The animal study was reviewed and approved by the Animal Protection and Ethics Committee of Nanjing University (IACUC-D2202077).

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Yin, L., Bing, Z., Zheng, Y. et al. Oroxylin A-induced Trained Immunity Promotes LC3-associated Phagocytosis in Macrophage in Protecting Mice Against Sepsis. Inflammation (2024). https://doi.org/10.1007/s10753-024-02033-2

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