Lymphatic dysfunction and ZFP36 deficiency contribute to myxomatous valve degeneration in Marfan syndrome mice
Can Tan, Ziyou Ren, Shreya Kurup, Xianpeng Liu, Zhi-Dong Ge, Shodai Suzuki, Pritika Jakka, Cheryl Tang, M. Luisa Iruela-Arispe, Tsutomu Kume
Can Tan, Ziyou Ren, Shreya Kurup, Xianpeng Liu, Zhi-Dong Ge, Shodai Suzuki, Pritika Jakka, Cheryl Tang, M. Luisa Iruela-Arispe, Tsutomu Kume
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Research Article Cardiology Development Vascular biology

Lymphatic dysfunction and ZFP36 deficiency contribute to myxomatous valve degeneration in Marfan syndrome mice

Abstract

Enhanced TGF-β signaling caused by mutations in Fibrillin-1 (FBN1) in patients with Marfan syndrome (MFS) leads to myxomatous degeneration of the mitral valve (MDMV). MDMV can result in mitral valve prolapse, severe regurgitation, and sudden cardiac death. However, it remains unknown whether lymphatic vessel (LV) dysfunction contributes to MDMV development in MFS. Here, we show that lymphangiogenesis in murine mitral valves (MVs) begins postnatally. However, this process is inhibited in a mouse MFS model, Fbn1 mutant (Fbn1C1039G/+) mice, accompanied by disrupted lymphatic cell-cell junctions, impaired lymphatic drainage, and an abnormally widespread distribution of MHCII+ infiltrating macrophages. Treatment of Fbn1 mutant mice with VEGF-C156S, a selective VEGFR3 agonist, stimulates the ERK and Akt pathways, increases LV density in MVs, and ameliorates MDMV. Fbn1 mutant MVs display disorganized valvular endothelial cells (VECs) and decreased expression of the antiinflammatory modulator Zfp36 (zinc finger protein 36) in VECs and immune cells. Treatment with FTY720 (fingolimod), a ZFP36 activator and S1P antagonist, rescues MDMV phenotypes in Fbn1 mutant mice by reducing immune cell infiltration and restoring lymphatic cell junctions and drainage. These findings suggest that the Fbn1 mutation causes LV hypoplasia and defective lymphatic drainage in MVs, driven in part by proinflammatory VECs, leading to MFS-related MDMV.

Authors

Can Tan, Ziyou Ren, Shreya Kurup, Xianpeng Liu, Zhi-Dong Ge, Shodai Suzuki, Pritika Jakka, Cheryl Tang, M. Luisa Iruela-Arispe, Tsutomu Kume

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Figure 7

FTY720 promotes lymphatic function and the VEC barrier in MVs of Fbn1 mutant mice.

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FTY720 promotes lymphatic function and the VEC barrier in MVs of Fbn1 mu...
(A) Representative confocal images show LVs (arrows) in aLs at P60. Scale bars: 200 μm. (B) Quantification of LV density in both leaflets. Data are mean ± SEM, ordinary 1-way ANOVA test, each symbol represents 1 mouse. N = 5-7, male/female = 4:3, 3:4, 4:2, and 3:2 in the 4 groups, respectively. **P < 0.01. (C) Representative confocal images show LEC junctions in lymphatic capillaries of MVs. Arrows indicate disrupted junctions. Scale bars: 10 μm. (D) Representative confocal images show VEC junctions at EC zone 5. Arrows indicate reticular adherens junctions. Scale bars: 20 μm. (E) Timeline of treatments for F and G. (F) Evans blue permeability assay in MVs. White dashed lines outline the regions of MV leaflets. EC-Z1, EC zone 1. Scale bars: 200 μm. (G) Quantification of fluorescence intensity (FI) of Evans blue in the interstitium beneath EC zone 1. Data are mean ± SEM, unpaired 2-tailed Student’s t test, each symbol represents 1 mouse. N = 4-5, male/female = 2-3:2. (H) Schematic summary of VEGF-C156S and FTY720 rescuing the MDMV phenotypes in Fbn1 mutant mice. In LECs of the Fbn1 mutant MVs, enhanced TGF-β signaling suppresses Prox1 expression and downregulates the VEGFR3 pathway. VEGF-C156S promotes lymphangiogenesis and lymphatic drainage, facilitating immune cell egress and interstitial fluid clearance, thereby inhibiting inflammation and preventing MDMV. Additionally, the antiinflammatory modulator ZFP36 is downregulated in VECs, macrophages, and DCs. The FDA-approved compound FTY720 activates PP2A, which dephosphorylates and thereby activates ZFP36, suppressing inflammation and preventing MDMV. FTY720 also facilitates zippering of LEC junctions via VEGF-A upregulation, thereby promoting efficient lymphatic drainage.