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Lymphangiogenesis and accumulation of reparative macrophages contribute to liver repair after hepatic ischemia–reperfusion injury

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Abstract

Hepatic tissue repair plays a critical role in determining the outcome of hepatic ischemia–reperfusion (I/R) injury. Hepatic lymphatics participate in the clearance of dead tissues and contribute to the reparative process after acute hepatic injury; however, it remains unknown whether lymphangiogenesis in response to hepatic inflammation is involved in liver repair. Herein, we determined if hepatic lymphangiogenesis improves liver repair after hepatic I/R injury. Using a mouse model of hepatic I/R injury, we investigated hepatic lymphatic structure, growth, and function in injured murine livers. Hepatic I/R injury enhanced lymphangiogenesis around the portal tract and this was associated with increased expression of pro-lymphangiogenic growth factors including vascular endothelial growth factor (VEGF)-C and VEGF-D. Recombinant VEGF-D treatment facilitated liver repair in association with the expansion of lymphatic vessels and increased expression of genes related to the reparative macrophage phenotype. Treatment with a VEGF receptor 3 (VEGFR3) inhibitor suppressed liver repair, lymphangiogenesis, drainage function, and accumulation of VEGFR3-expressing reparative macrophages. VEGF-C and VEGF-D upregulated expression of genes related to lymphangiogenic factors and the reparative macrophage phenotype in cultured macrophages. These results suggest that activation of VEGFR3 signaling increases lymphangiogenesis and the number of reparative macrophages, both of which play roles in liver repair. Expanded lymphatics and induction of reparative macrophage accumulation may be therapeutic targets to enhance liver repair after hepatic injury.

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Abbreviations

ALT:

Alanine aminotransferase

BM:

Bone marrow

BMM:

Bone marrow-derived macrophage

GAPDH:

Glyceraldehyde-3-phosphate dehydrogenase

H&E:

Hematoxylin and eosin

IL:

Interleukin

i.p.:

Intraperitoneal

I/R:

Ischemia–reperfusion

LVA:

Lymphatic vessel area

LVD:

Lymphatic vessel density

LYVE:

Lymphatic vessel endothelial hyaluronan receptor

MR:

Mannose receptor

MVA:

Microvessel area

MVD:

Microvessel density

PBS:

Phosphate-buffered saline

PCNA:

Proliferating cell nuclear antigen

SD:

Standard deviation

TNF:

Tumor necrosis factor

VEGF:

Vascular endothelial growth factor

VEGFR3:

Vascular endothelial growth factor receptor 3

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Acknowledgements

We thank Michiko Ogino and Kyoko Yoshikawa for their technical assistance. This research was supported by research Grants (18K16373, 18K15760 18H02605, and 19K09156) from the Japanese Ministry of Education, Culture, Sports, Science, and Technology and by a Project Research Grant from the Graduate School of Medical Sciences, Kitasato University.

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  1. Shuji Nakamoto and Yoshiya Ito have contributed equally to this work.

Authors and Affiliations

  1. Department of Molecular Pharmacology, Graduate School of Medical Sciences, Kitasato University, Sagamihara, Kanagawa, Japan

    Shuji Nakamoto, Yoshiya Ito, Takuya Goto & Masataka Majima

  2. Department of Pharmacology, Kitasato University School of Medicine, 1-15-1 Kitasato, Minami-ku, Kanagawa, Sagamihara, 252-0374, Japan

    Shuji Nakamoto, Yoshiya Ito, Takuya Goto & Masataka Majima

  3. Department of Surgery, Kitasato University School of Medicine, Sagamihara, Kanagawa, Japan

    Shuji Nakamoto, Nobuyuki Nishizawa, Takuya Goto, Ken Kojo, Yusuke Kumamoto & Masahiko Watanabe

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  1. Shuji Nakamoto
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Correspondence to Masataka Majima.

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All experimental procedures were approved by the Animal Experimentation and Ethics Committee of the Kitasato University School of Medicine (2017-59) and were performed in accordance with the guidelines for animal experiments outlined by the Kitasato University School of Medicine, which are in accordance with the Guidelines for Proper Conduct of Animal Experiments published by the Science Council of Japan. This article does not contain any studies with human participants performed by any of the authors.

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Nakamoto, S., Ito, Y., Nishizawa, N. et al. Lymphangiogenesis and accumulation of reparative macrophages contribute to liver repair after hepatic ischemia–reperfusion injury. Angiogenesis 23, 395–410 (2020). https://doi.org/10.1007/s10456-020-09718-w

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