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Galectin-3 Mediates Endotoxin Internalization and Caspase-4/11 Activation in Tubular Epithelials and Macrophages During Sepsis and Sepsis-Associated Acute Kidney Injury

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Abstract

Besides being recognized by membrane receptor TLR4, lipopolysaccharide (LPS) can also be internalized into the cytosol and activate Caspase-4/11 pyroptotic pathways to further amplify inflammation in sepsis. The objective of this study was to investigate whether Galectin-3 (Gal3) could promote the uptake of LPS by governing RAGE or administering endocytosis, consequently activating Caspase 4/11 and mediating pyroptosis in sepsis-associated acute kidney injury (SA-AKI). By pinpointing Gal3, LPS, and EEA1 (endosome-marker) or LAMP1 (lysosome-marker) respectively, immunofluorescence discovered that Gal3 and LPS were mainly aggregated in early endosomes initially and translocated into lysosomes afterwards. In cells and animal models, Gal3 and the Caspase-4/11 pathways were simultaneously activated, and the overexpression of Gal3 could exacerbate pyroptosis, whereas inhibition of Gal3 or the knockdown of its expression could ameliorate pyroptosis, reduce the pathological changes of SA-AKI and improve the survival of the animals with SA-AKI. Silencing RAGE reduced pyroptosis in primary tubular epithelial cells (PTCs) activated by Gal3 and LPS but not in cells activated by Gal3 and outer membrane vesicles (with LPS inside), whereas pyroptosis in both was reduced by blockade of Gal3, indicating Gal3 promoted pyroptosis through both RAGE-dependent and RAGE-independent pathways. Our investigation further revealed a positive correlation between serum Gal3 and pyroptotic biomarkers IL-1 beta and IL-18 in patients with sepsis, and that serum Gal3 was an independent risk factor for mortality. Through our collective exploration, we unraveled the significant role of Gal3 in the internalization of LPS and the provocation of more intense pyroptosis, thus making it a vital pathogenic factor in SA-AKI and a possible therapeutic target. Gal3 enabled the internalization of endotoxin into endosomes and lysosomes via both RAGE-dependent (A) and RAGE-independent (B) pathways, leading to pyroptosis. The suppression of Gal3 curbed Caspase4/11 noncanonical inflammasomes and diminished sepsis and SA-AKI.

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Abbreviations

Gal3:

Galectin-3

MCP:

Modified citrus pectin

AKI:

Acute kidney injury

RAGE:

Receptor for advanced glycation end products

LPS:

Lipopolysaccharide

OMVs:

Outer membrane vesicles

TEM:

Transmission electron microscopy

CRE:

Creatinine

ELISA:

Enzyme-linked immunosorbent assay

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Acknowledgements

We sincerely thank Pro. Isaac Eliaz of Amitabha Medical Center (Santa Rosa, CA, USA) and Pro. Glenn Chertow of the Department of Medicine, Stanford University School of Medicine (Stanford, CA, USA) for their academic guidance.

Funding

This work was supported by the National Natural Science Foundation of China (Grant 82241039, 82272208 and 81971816 to Pro. Zhiyong Peng and 82102273 to Dr. Yiming Li).

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Authors and Affiliations

  1. Department of Critical Care Medicine, Zhongnan Hospital, Wuhan University, Wuhan, Hubei, China

    Fengyun Wang, Junwei Ye, Weiwei Zhu, Ruiqi Ge, Chang Hu, Yaoyao Qian & Zhiyong Peng

  2. Clinical Research Center of Hubei Critical Care Medicine, Zhongnan Hospital of Wuhan University, Wuhan, Hubei, China

    Fengyun Wang, Junwei Ye, Weiwei Zhu, Ruiqi Ge, Chang Hu, Yaoyao Qian, Yiming Li & Zhiyong Peng

  3. Department of Critical Care Medicine, Huangshi Central Hospital, Affiliated Hospital of Hubei Polytechnic University, Huangshi, China

    Junwei Ye

  4. Department of Critical Care Medicine, Center of Critical Care Nephrology, University of Pittsburgh School of Medicine, Pittsburgh, PA, USA

    Zhiyong Peng

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Contributions

FW and JY contributed equally to this work; they conceived the idea, did the experiments, and analyzed the data together. The manuscript was written by FW. WZ, RG, CH, and YQ validated the data of this work. ZP and YL were involved in revising the manuscript critically for important intellectual content. All authors read and approved the final manuscript.

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Correspondence to Yiming Li or Zhiyong Peng.

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Wang, F., Ye, J., Zhu, W. et al. Galectin-3 Mediates Endotoxin Internalization and Caspase-4/11 Activation in Tubular Epithelials and Macrophages During Sepsis and Sepsis-Associated Acute Kidney Injury. Inflammation 47, 454–468 (2024). https://doi.org/10.1007/s10753-023-01928-w

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