Macrophages regulate salt-dependent volume and blood pressure by a vascular endothelial growth factor-C–dependent buffering mechanism

Abstract

In salt-sensitive hypertension, the accumulation of Na+ in tissue has been presumed to be accompanied by a commensurate retention of water to maintain the isotonicity of body fluids. We show here that a high-salt diet (HSD) in rats leads to interstitial hypertonic Na+ accumulation in skin, resulting in increased density and hyperplasia of the lymphcapillary network. The mechanisms underlying these effects on lymphatics involve activation of tonicity-responsive enhancer binding protein (TonEBP) in mononuclear phagocyte system (MPS) cells infiltrating the interstitium of the skin. TonEBP binds the promoter of the gene encoding vascular endothelial growth factor-C (VEGF-C, encoded by Vegfc) and causes VEGF-C secretion by macrophages. MPS cell depletion or VEGF-C trapping by soluble VEGF receptor-3 blocks VEGF-C signaling, augments interstitial hypertonic volume retention, decreases endothelial nitric oxide synthase expression and elevates blood pressure in response to HSD. Our data show that TonEBP–VEGF-C signaling in MPS cells is a major determinant of extracellular volume and blood pressure homeostasis and identify VEGFC as an osmosensitive, hypertonicity-driven gene intimately involved in salt-induced hypertension.

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Figure 1: Lymph vessel hyperplasia in response to dietary salt loading.
Figure 2: MPS cells are necessary for the hyperplastic response of the lymphcapillary network to HSD.
Figure 3: MPS depletion leads to augmented volume retention and further blood pressure increase in response to HSD.
Figure 4: MPS cells and VEGF-C modulate eNOS protein expression in response to HSD.
Figure 5: High salt increases TonEBP and VEGF-C mRNA and protein expression in macrophages.

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Acknowledgements

This work was supported in part by grants from the Interdisziplinäres Zentrum für klinische Forschung Erlangen (TP B13), from the Bundesministerium für Bildung und Forschung - Forschung unter Weltraumbedingungen (50WB0620), and from the Deutsche Forschungsgemeinschaft (Ti345/2) to J.T., from the sixth Framework Integrated Project Lymphangiogenomics (LSGH-2004-503573) to D.K., and from a Fresenius Nephro-Core Stipend to A.Z. F.C.L. and D.N.M. were supported by EuReGene; D.N.M. is a Helmholtz fellow. We thank N. Rakova for translating Russian articles and E. Prell, M. Klewer and B. Hausknecht for their technical assistance. We thank P. Uhrin (Department of Vascular Biology and Thrombosis Research, Medical University of Vienna) for the Swiss-129Sv mice and H.M. Kwon (Department of Medicine, University of Maryland) for pCMV-Tag2-TonEBP.

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Contributions

A.M. conducted the experiments and generated all experimental data, W.N. and F.-X.B. generated cells with stable TonEBP overexpression and contributed to the in vitro studies and writing of the manuscript, J.J. contributed to the in vitro studies, A.D., J.G. and A.Z. contributed to the animal studies, T.T. and K.A. provided adenoviruses and contributed to the adenoviral animal experiments, K.M. and A.K. did the three-dimensional resolution of the lymph capillary network, J.-K.P. analyzed and quantified eNOS expression, D.N.M. contributed to animal experiments and writing of the manuscript, W.D. provided human serum from subjects with refractory hypertension, P.D. and H.W. contributed to analysis of internal electrolyte redistribution in animal experiments by chemical analysis, N.v.R. generated clodronate liposomes for MPS depletion experiments, K.F.H. and K.-U.E. contributed to the conception of experimental design and the writing of the manuscript, F.C.L. provided serum samples from subjects and wrote the manuscript, D.K. contributed the methods and experimental design for quantification of lymph capillary network changes and the conception of macrophage–VEGF-C–lymph capillary interaction, J.T. planned and organized the experimental approach, supervised the project, analyzed the data statistically and wrote the manuscript.

Corresponding author

Correspondence to Jens Titze.

Supplementary information

Supplementary Text and Figures

Supplementary Figs. 1–4, Supplementary Table 1 and Supplementary Methods (PDF 775 kb)

Supplementary Video 1

Lymphcapillaries in rat ear, low-salt salt diet (MOV 4860 kb)

Supplementary Video 2

Lymphcapillaries in rat ear, high-salt diet (MOV 4086 kb)

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Machnik, A., Neuhofer, W., Jantsch, J. et al. Macrophages regulate salt-dependent volume and blood pressure by a vascular endothelial growth factor-C–dependent buffering mechanism. Nat Med 15, 545–552 (2009). https://doi.org/10.1038/nm.1960

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