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Defective valves and abnormal mural cell recruitment underlie lymphatic vascular failure in lymphedema distichiasis

Nature Medicine volume 10, pages 974–981 (2004)Cite this article

Abstract

Lymphatic vessels are essential for the removal of interstitial fluid and prevention of tissue edema. Lymphatic capillaries lack associated mural cells, and collecting lymphatic vessels have valves, which prevent lymph backflow. In lymphedema-distichiasis (LD), lymphatic vessel function fails because of mutations affecting the forkhead transcription factor FOXC2. We report that Foxc2−/− mice show abnormal lymphatic vascular patterning, increased pericyte investment of lymphatic vessels, agenesis of valves and lymphatic dysfunction. In addition, an abnormally large proportion of skin lymphatic vessels was covered with smooth muscle cells in individuals with LD and in mice heterozygous for Foxc2 and for the gene encoding lymphatic endothelial receptor, Vegfr3 (also known as Flt4). Our data show that Foxc2 is essential for the morphogenesis of lymphatic valves and the establishment of a pericyte-free lymphatic capillary network and that it cooperates with Vegfr3 in the latter process. Our results indicate that an abnormal interaction between the lymphatic endothelial cells and pericytes, as well as valve defects, underlie the pathogenesis of LD.

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Figure 1: Late development of the lymphatic vasculature is abnormal in Foxc2−/− embryos.
Figure 2: Expression of Pdgfrb, Pdgfb, endoglin and collagen IV in the lymphatic vessels of Foxc2−/− embryos.
Figure 3: Compound Foxc2+/−;Vegfr3+/lacZ mice show a lymphatic phenotype similar to that of Foxc2−/− mice.
Figure 4: Abnormal lymphatic vascular function and agenesis of lymphatic valves in Foxc2−/− mice.
Figure 5: Expression pattern of Foxc2 in lymphatic vessels.
Figure 6: Effects of human FOXC2 mutations in vivo and in cell culture.

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Acknowledgements

We thank St George's and St Thomas' lymphedema consortium (S. Jeffery, J.R. Levick, A. Child, K.G. Burnand, S. Mansour, G. Brice, M. Sarfarazi and C. Sholto-Douglas-Vernon) for help with the patient material; C. Betsholtz and C. Bondjers for providing probes for Pdgfb, Pdgfrb, Rgs5 and for discussions; P. Laakkonen and M. Jeltsch for providing anti-Lyve-1 sera; U. Ozerdem for providing the NG2 antibodies; M. Kimak for the preparation of DNA from individuals with LD; and T. Tainola, A. Parsons, M. Helanterä, T. Laakkonen, S. Lampi, K. Makkonen, P. Hyvärinen and M. Miller for technical assistance. This study was supported by grants from Finnish Academy of Sciences, Finnish Cancer Organizations, Sigrid Juselius Foundation, Helsinki University Central Hospital, Finnish Cultural Foundation (T.K.), Paulo Foundation (T.K.), Emil Aaltonen Foundation (T.K.), Aarne Koskelo Foundation (T.K.) and by the EU Integrated Project LSHG-CT-2004-503573.

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Author notes

  1. Tatiana V Petrova and Terhi Karpanen: These authors contributed equally to this work.

Authors and Affiliations

  1. Molecular/Cancer Biology Laboratory and Ludwig Institute for Cancer Research, Biomedicum Helsinki and Helsinki University Central Hospital, University of Helsinki, Haartmaninkatu 8, P.O.B. 63, Helsinki, 00014, Finland

    Tatiana V Petrova, Terhi Karpanen, Camilla Norrmén & Kari Alitalo

  2. Dermatology Unit, St. George's Hospital Medical School, Cranmer Terrace, London, SW17 0RE, UK

    Russell Mellor

  3. Department of Biochemistry, Hamamatsu University School of Medicine, Hamamatsu, 431-3192, Japan

    Tomoki Tamakoshi & Naoyuki Miura

  4. Department of Human Genetics, Graduate School of Public Health, University of Pittsburgh, Pittsburgh, 15261, Pennsylvania, USA

    David Finegold & Robert Ferrell

  5. Department of Pediatrics, School of Medicine, University of Pittsburgh, Pittsburgh, 15261, Pennsylvania, USA

    David Finegold

  6. Department of Pathology, University of Vienna Medical School, Vienna, 1090, Austria

    Dontscho Kerjaschki & Peter Mortimer

  7. A.I.Virtanen Institute, University of Kuopio, P.O.B. 1627, Kuopio, 70211, Finland

    Seppo Ylä-Herttuala

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Supplementary information

Supplementary Fig. 1

Analysis of lymphatic vasculature in adult Foxc2+/− mice. (PDF 310 kb)

Supplementary Fig. 2

Normal early development of lymphatic vasculature in Foxc2−/− mice. (PDF 506 kb)

Supplementary Fig. 3

Abnormal lymphatic patterning and PC/SMC recruitment in the skin of Foxc2−/− embryos. (PDF 295 kb)

Supplementary Fig. 4

Skin blood vessel development is normal in Foxc2−/− mice. (PDF 378 kb)

Supplementary Fig. 5

Mural cells associated with lymphatic vessels of Foxc2−/− mice express the PC marker NG2. (PDF 580 kb)

Supplementary Fig. 6

Pdgfb, Vegfr3, Prox1 and podoplanin are expressed in lymphatic endothelium lacking Foxc2. (PDF 320 kb)

Supplementary Fig. 7

Genetic relationship between Vegfr3 and Foxc2. (PDF 309 kb)

Supplementary Fig. 8

Expression pattern of FOXC2. (PDF 250 kb)

Supplementary Table 1

Use of antibodies against lymphatic endothelial specific markers in different experimental settings. (PDF 19 kb)

Supplementary Methods (PDF 26 kb)

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Petrova, T., Karpanen, T., Norrmén, C. et al. Defective valves and abnormal mural cell recruitment underlie lymphatic vascular failure in lymphedema distichiasis. Nat Med 10, 974–981 (2004). https://doi.org/10.1038/nm1094

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