LYVE1 ectodomain shedding blunts lymphatic transmigration and clearance of macrophages during kidney injury
Jing Liu, Yuqing Liu, Wenqian Zhou, Saiya Zhu, Jianyong Zhong, Haichun Yang, Annet Kirabo, Valentina Kon, Chen Yu
Jing Liu, Yuqing Liu, Wenqian Zhou, Saiya Zhu, Jianyong Zhong, Haichun Yang, Annet Kirabo, Valentina Kon, Chen Yu
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Research Article Inflammation Nephrology Vascular biology

LYVE1 ectodomain shedding blunts lymphatic transmigration and clearance of macrophages during kidney injury

Abstract

Although renal fibrosis is predominantly driven by the accumulated inflammatory cells that secrete proinflammatory factors within the kidney, the key mechanisms underlying macrophage clearance from the kidney are not well understood. The interaction of hyaluronan with lymphatic endothelial hyaluronan receptor 1 (LYVE1) constitutes a critical initial step in macrophage adhesion and removal by lymphatic vessels. This study investigates alterations in LYVE1 during kidney disease and elucidates its role in macrophage trafficking. Three renal fibrosis models demonstrated a reduction in full-length LYVE1 and an increase in the soluble LYVE1 fragment. Immunostaining of fibrotic kidneys showed significantly reduced expression of soluble LYVE1 compared with the intracellular fragment (Cyto-LYVE1), demonstrating ectodomain shedding of LYVE1 in vivo and in vitro. Functionally, human lymphatic endothelial cells exposed to TGF-β1 exhibited a significant decrease in macrophage adhesion and transendothelial migration compared with controls. Mechanistic analyses identified increased matrix metalloproteinase 9 (MMP9) in renal injury as a key upstream regulator of LYVE1 shedding. MMP9 inhibitors reduced LYVE1 shedding, enhanced macrophage adhesion and trafficking, and mitigated macrophage accumulation and disease progression. In conclusion, MMP9-induced LYVE1 shedding is linked to progressive kidney fibrosis and macrophage accumulation. LYVE1 shedding inhibitors offer potential as therapeutic agents for mitigating immune overload and kidney fibrosis.

Authors

Jing Liu, Yuqing Liu, Wenqian Zhou, Saiya Zhu, Jianyong Zhong, Haichun Yang, Annet Kirabo, Valentina Kon, Chen Yu

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Figure 3

MMP9 mediates LYVE1 ectodomain shedding.

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MMP9 mediates LYVE1 ectodomain shedding. (A) Fold change of RNA expressi...
(A) Fold change of RNA expression of MMPs on LECs after TGF-β1 stimulation. (B) Time course for full-length LYVE1 and Cyto-LYVE1 in TGF-β1–treated hLECs assessed by Western blot. (C) MMP9 in supernatant of TGF-β1–treated hLECs measured by gelatin zymography. (D) Full-length and Cyto-LYVE1 protein expression of MMP9-exposed hLECs assessed by Western blot. (E) Full-length and Cyto-LYVE1 protein expression of MMP9-treated hLECs assessed by ICC (original magnification, ×1,000; scale bars: 20 μm). (F) Soluble LYVE1 in supernatant of MMP9-exposed hLECs assessed by ELISA. (G) MMP9 expression in hLECs after siRNA-mediated knockdown. (H) MMP9 in the supernatant of hLECs treated with siMMP9 assessed by gelatin zymography. (I) Full-length and Cyto-LYVE1 protein expression of hLECs treated with siMMP9 inhibitors assessed by Western blot. (J) Full-length and Cyto-LYVE1 protein expression of hLECs treated with siMMP9 assessed by ICC (original magnification, ×1,000; scale bars: 20 μm). (K) Soluble LYVE1 in the supernatant of hLECs treated with siMMP9 measured by ELISA. (L and M) Western blot of MMP9, LYVE1, and Cyto-LYVE1 in UUO at the end of the experiment (14 days) (L) and over the course of the study (M). n = 3 per group for cell experiments; statistics used included a 2-tailed t test (2 groups, in D, F, G, and L) or 1-way ANOVA (multiple groups, in I and K).*P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.