^{Department of Pathology and Laboratory Medicine and Institute for Medicine and Engineering, University of Pennsylvania, Philadelphia, Pennsylvania, USA.}

^{Cardiovascular Biology Research Program, Oklahoma Medical Research Foundation, Oklahoma City, Oklahoma, USA.}

Address correspondence to: Mark L. Kahn, University of Pennsylvania, Smilow Center for Translational Research – Room 11-123, 3400 Civic Center Blvd., Building 421, Philadelphia, Pennsylvania 19104, USA. Phone: 215.898.9007; E-mail: ude.nnepu.dem.liam@nhakkram.

Received 2014 Oct 6; Accepted 2015 May 22.

Abstract

Fluid shear forces have established roles in blood vascular development and function, but whether such forces similarly influence the low-flow lymphatic system is unknown. It has been difficult to test the contribution of fluid forces in vivo because mechanical or genetic perturbations that alter flow often have direct effects on vessel growth. Here, we investigated the functional role of flow in lymphatic vessel development using mice deficient for the platelet-specific receptor C-type lectin–like receptor 2 (CLEC2) as blood backfills the lymphatic network and blocks lymph flow in these animals. CLEC2-deficient animals exhibited normal growth of the primary mesenteric lymphatic plexus but failed to form valves in these vessels or remodel them into a structured, hierarchical network. Smooth muscle cell coverage (SMC coverage) of CLEC2-deficient lymphatic vessels was both premature and excessive, a phenotype identical to that observed with loss of the lymphatic endothelial transcription factor FOXC2. In vitro evaluation of lymphatic endothelial cells (LECs) revealed that low, reversing shear stress is sufficient to induce expression of genes required for lymphatic valve development and identified GATA2 as an upstream transcriptional regulator of FOXC2 and the lymphatic valve genetic program. These studies reveal that lymph flow initiates and regulates many of the key steps in collecting lymphatic vessel maturation and development.

Keywords: Angiogenesis, Cardiology, Development, Oncology, Vascular Biology

Abstract

Acknowledgments

The authors thank Dean Sheppard for donation of Itga9-null mice. We thank Lydia Sorokin for donation of the anti-laminin α5 antibody. We thank Kyle Kostesich for design of illustrations. We thank MinMin Lu and the University of Pennsylvania Histology Core Facility for histology support. This work was supported by the National Heart, Lung, and Blood Institute (NHLBI) grant T32HL007439 (to D.T. Sweet), NIH grant K25 HL107617 (to J.M. Jiménez), NIH grant P01 HL62250 (to P.F. Davies), NHLBI grant HL085607 and National Institute of General Medical Sciences grant GM103441 (to L. Xia), and by the Leducq Foundation and NIH grant R01 HL103432 (to M.L. Kahn).

Acknowledgments

Footnotes

Conflict of interest: The authors have declared that no conflict of interest exists.

Reference information:J Clin Invest. 2015;125(8):2995–3007. doi:10.1172/JCI79386.

See the related Commentary beginning on page 2924.

Footnotes

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