Zebrafish prox1b Mutants Develop a Lymphatic Vasculature, and prox1b Does Not Specifically Mark Lymphatic Endothelial Cells

Recent discoveries of novel functions and diverse origins of lymphatic vessels have drastically changed our view of lymphatic vasculature. Traditionally regarded as passive conduits for fluid and immune cells, lymphatic vessels now emerge as active, tissue-specific players in major physiological and pathophysiological processes. Lymphatic vessels show remarkable plasticity and heterogeneity, reflecting their functional specialization to control the tissue microenvironment. Moreover, alternative developmental origins of lymphatic endothelial cells in some organs may contribute to the diversity of their functions in adult tissues. This review aims to summarize the most recent findings of organotypic differentiation of lymphatic endothelial cells in terms of their distinct (patho)physiological functions in skin, lymph nodes, small intestine, brain, and eye. We discuss recent advances in our understanding of the heterogeneity of lymphatic vessels with respect to the organ-specific functional and molecular specialization of lymphatic endothelium, such as the hybrid blood-lymphatic identity of Schlemm’s canal, functions of intestinal lymphatics in dietary fat uptake, and discovery of meningeal lymphatic vasculature and perivascular brain lymphatic endothelial cells.

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Lymphatic endothelial progenitors bud from the cardinal vein and intersomitic vessels in mammalian embryos

Blood ◽

10.1182/blood-2012-05-428607 ◽

2012 ◽

Vol 120 (11) ◽

pp. 2340-2348 ◽

Cited By ~ 128

Author(s):

Ying Yang ◽

José Manuel García-Verdugo ◽

Mario Soriano-Navarro ◽

R. Sathish Srinivasan ◽

Joshua P. Scallan ◽

...

Keyword(s):

Endothelial Cells ◽

Blood Vessels ◽

Fluid Balance ◽

Tissue Fluid ◽

Lymphatic Endothelial Cells ◽

Cardinal Vein ◽

Cellular Processes ◽

Mammalian Embryos ◽

Lymphatic Vasculature ◽

Lymphatic Network

Abstract The lymphatic vasculature preserves tissue fluid balance by absorbing fluid and macromolecules and transporting them to the blood vessels for circulation. The stepwise process leading to the formation of the mammalian lymphatic vasculature starts by the expression of the gene Prox1 in a subpopulation of blood endothelial cells (BECs) on the cardinal vein (CV) at approximately E9.5. These Prox1-expressing lymphatic endothelial cells (LECs) will exit the CV to form lymph sacs, primitive structures from which the entire lymphatic network is derived. Until now, no conclusive information was available regarding the cellular processes by which these LEC progenitors exit the CV without compromising the vein's integrity. We determined that LECs leave the CV by an active budding mechanism. During this process, LEC progenitors are interconnected by VE-cadherin–expressing junctions. Surprisingly, we also found that Prox1-expressing LEC progenitors were present not only in the CV but also in the intersomitic vessels (ISVs). Furthermore, as LEC progenitors bud from the CV and ISVs into the surrounding mesenchyme, they begin expressing the lymphatic marker podoplanin, migrate away from the CV, and form the lymph sacs. Analyzing this process in Prox1-null embryos revealed that Prox1 activity is necessary for LEC progenitors to exit the CV.

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Liprin β1 is highly expressed in lymphatic vasculature and is important for lymphatic vessel integrity

Blood ◽

10.1182/blood-2009-03-212274 ◽

2010 ◽

Vol 115 (4) ◽

pp. 906-909 ◽

Cited By ~ 25

Author(s):

Camilla Norrmén ◽

Wouter Vandevelde ◽

Annelii Ny ◽

Pipsa Saharinen ◽

Massimiliano Gentile ◽

...

Keyword(s):

Endothelial Cells ◽

Lymphatic Vessel ◽

Lipid Absorption ◽

Tissue Fluid ◽

Lymphatic Endothelial Cells ◽

Isolation And Characterization ◽

Lymphatic Vasculature ◽

Vessel Integrity

Abstract The lymphatic vasculature is important for the regulation of tissue fluid homeostasis, immune response, and lipid absorption, and the development of in vitro models should allow for a better understanding of the mechanisms regulating lymphatic vascular growth, repair, and function. Here we report isolation and characterization of lymphatic endothelial cells from human intestine and show that intestinal lymphatic endothelial cells have a related but distinct gene expression profile from human dermal lymphatic endothelial cells. Furthermore, we identify liprin β1, a member of the family of LAR transmembrane tyrosine phosphatase-interacting proteins, as highly expressed in intestinal lymphatic endothelial cells in vitro and lymphatic vasculature in vivo, and show that it plays an important role in the maintenance of lymphatic vessel integrity in Xenopus tadpoles.

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Cell Fate Determination of Lymphatic Endothelial Cells

International Journal of Molecular Sciences ◽

10.3390/ijms21134790 ◽

2020 ◽

Vol 21 (13) ◽

pp. 4790

Author(s):

Young Jae Lee

Keyword(s):

Endothelial Cells ◽

Immune Responses ◽

Cell Fate ◽

Dietary Fat ◽

Vascular System ◽

Lymphatic Endothelial Cells ◽

Lymphatic Vasculature ◽

Fat Uptake ◽

Mammalian System

The lymphatic vasculature, along with the blood vasculature, is a vascular system in our body that plays important functions in fluid homeostasis, dietary fat uptake, and immune responses. Defects in the lymphatic system are associated with various diseases such as lymphedema, atherosclerosis, fibrosis, obesity, and inflammation. The first step in lymphangiogenesis is determining the cell fate of lymphatic endothelial cells. Several genes involved in this commitment step have been identified using animal models, including genetically modified mice. This review provides an overview of these genes in the mammalian system and related human diseases.

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T1α/podoplanin is essential for capillary morphogenesis in lymphatic endothelial cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.90262.2008 ◽

2008 ◽

Vol 295 (4) ◽

pp. L543-L551 ◽

Cited By ~ 34

Author(s):

Angels Navarro ◽

Ricardo E. Perez ◽

Mo Rezaiekhaligh ◽

Sherry M. Mabry ◽

Ikechukwu I. Ekekezie

Keyword(s):

Endothelial Cells ◽

Capillary Tube ◽

Transmembrane Protein ◽

Tube Formation ◽

Small Gtpase ◽

Lymph Vessel ◽

Early Event ◽

Lymphatic Endothelial Cells ◽

Podoplanin Expression ◽

Lymphatic Vasculature

The lymphatic vasculature functions to maintain tissue perfusion homeostasis. Defects in its formation or disruption of the vessels result in lymphedema, the effective treatment of which is hampered by limited understanding of factors regulating lymph vessel formation. Mice lacking T1α/podoplanin, a lymphatic endothelial cell transmembrane protein, have malformed lymphatic vasculature with lymphedema at birth, but the molecular mechanism for this phenotype is unknown. Here, we show, using primary human lung microvascular lymphatic endothelial cells (HMVEC-LLy), that small interfering RNA-mediated silence of podoplanin gene expression has the dramatic effect of blocking capillary tube formation in Matrigel. In addition, localization of phosphorylated ezrin/radixin/moesin proteins to plasma membrane extensions, an early event in the capillary morphogenic program in lymphatic endothelial cells, is impaired. We find that cells with decreased podoplanin expression fail to properly activate the small GTPase RhoA early (by 30 min) after plating on Matrigel, and Rac1 shows a delay in its activation. Further indication that podoplanin action is linked to RhoA activation is that use of a cell-permeable inhibitor of Rho inhibited lymphatic endothelial capillary tube formation in the same manner as did podoplanin gene silencing, which was not mimicked by treatment with a Rac1 inhibitor. These data clearly demonstrate that early activation of RhoA in the lymphangiogenic process, which is required for the successful establishment of the capillary network, is dependent on podoplanin expression. To our knowledge, this is the first time that a mechanism has been suggested to explain the role of podoplanin in lymphangiogenesis.

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Ligand-directed targeting of lymphatic vessels uncovers mechanistic insights in melanoma metastasis

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.1424994112 ◽

2015 ◽

Vol 112 (8) ◽

pp. 2521-2526 ◽

Cited By ~ 10

Author(s):

Dawn R. Christianson ◽

Andrey S. Dobroff ◽

Bettina Proneth ◽

Amado J. Zurita ◽

Ahmad Salameh ◽

...

Keyword(s):

Endothelial Cells ◽

Lymphatic Vessels ◽

Melanoma Cells ◽

Cell Interactions ◽

Melanoma Metastasis ◽

Lymphatic Endothelium ◽

Lymphatic Endothelial Cells ◽

Cell Surfaces ◽

Surface Binding ◽

Lymphatic Vasculature

Metastasis is the most lethal step of cancer progression in patients with invasive melanoma. In most human cancers, including melanoma, tumor dissemination through the lymphatic vasculature provides a major route for tumor metastasis. Unfortunately, molecular mechanisms that facilitate interactions between melanoma cells and lymphatic vessels are unknown. Here, we developed an unbiased approach based on molecular mimicry to identify specific receptors that mediate lymphatic endothelial–melanoma cell interactions and metastasis. By screening combinatorial peptide libraries directly on afferent lymphatic vessels resected from melanoma patients during sentinel lymphatic mapping and lymph node biopsies, we identified a significant cohort of melanoma and lymphatic surface binding peptide sequences. The screening approach was designed so that lymphatic endothelium binding peptides mimic cell surface proteins on tumor cells. Therefore, relevant metastasis and lymphatic markers were biochemically identified, and a comprehensive molecular profile of the lymphatic endothelium during melanoma metastasis was generated. Our results identified expression of the phosphatase 2 regulatory subunit A, α-isoform (PPP2R1A) on the cell surfaces of both melanoma cells and lymphatic endothelial cells. Validation experiments showed that PPP2R1A is expressed on the cell surfaces of both melanoma and lymphatic endothelial cells in vitro as well as independent melanoma patient samples. More importantly, PPP2R1A-PPP2R1A homodimers occur at the cellular level to mediate cell–cell interactions at the lymphatic–tumor interface. Our results revealed that PPP2R1A is a new biomarker for melanoma metastasis and show, for the first time to our knowledge, an active interaction between the lymphatic vasculature and melanoma cells during tumor progression.

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Local inhibition of elastase reduces EMILIN1 cleavage reactivating lymphatic vessel function in a mouse lymphoedema model

Clinical Science ◽

10.1042/cs20160064 ◽

2016 ◽

Vol 130 (14) ◽

pp. 1221-1236 ◽

Cited By ~ 11

Author(s):

Eliana Pivetta ◽

Bruna Wassermann ◽

Lisa Del Bel Belluz ◽

Carla Danussi ◽

Teresa Maria Elisa Modica ◽

...

Keyword(s):

Endothelial Cells ◽

Lymphatic Vessel ◽

Intercellular Junctions ◽

Lymphatic Endothelial Cells ◽

Lymph Drainage ◽

Regional Lymph Nodes ◽

Morphological Alterations ◽

Local Inhibition ◽

Lymphatic Vasculature ◽

Powerful Strategy

Lymphatic vasculature critically depends on the connections of lymphatic endothelial cells with the extracellular matrix (ECM), which are mediated by anchoring filaments (AFs). The ECM protein EMILIN1 is a component of AFs and is involved in the regulation of lymphatic vessel functions: accordingly, Emilin1−/− mice display lymphatic vascular morphological alterations, leading to functional defects such as mild lymphoedema, lymph leakage and compromised lymph drainage. In the present study, using a mouse post-surgical tail lymphoedema model, we show that the acute phase of acquired lymphoedema correlates with EMILIN1 degradation due to neutrophil elastase (NE) released by infiltrating neutrophils. As a consequence, the intercellular junctions of lymphatic endothelial cells are weakened and drainage to regional lymph nodes is severely affected. The local administration of sivelestat, a specific NE inhibitor, prevents EMILIN1 degradation and reduces lymphoedema, restoring a normal lymphatic functionality. The finding that, in human secondary lymphoedema samples, we also detected cleaved EMILIN1 with the typical bands of an NE-dependent pattern of fragmentation establishes a rationale for a powerful strategy that targets NE inhibition. In conclusion, the attempts to block EMILIN1 degradation locally represent the basis for a novel ‘ECM’ pharmacological approach to assessing new lymphoedema treatments.

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Isolation of Human Skin Lymphatic Endothelial Cells and 3D Reconstruction of the Lymphatic Vasculature In Vitro

Methods in Molecular Biology - Lymphangiogenesis ◽

10.1007/978-1-4939-8712-2_18 ◽

2018 ◽

pp. 279-290 ◽

Cited By ~ 1

Author(s):

Anita Rogic ◽

Francois Auger ◽

Mihaela Skobe

Keyword(s):

Endothelial Cells ◽

3D Reconstruction ◽

Human Skin ◽

Lymphatic Endothelial Cells ◽

Lymphatic Vasculature

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Single-Cell RNA Sequencing Reveals Heterogeneity and Functional Diversity of Lymphatic Endothelial Cells

International Journal of Molecular Sciences ◽

10.3390/ijms222111976 ◽

2021 ◽

Vol 22 (21) ◽

pp. 11976

Author(s):

Hannah den Braanker ◽

Astrid C. van Stigt ◽

Marc R. Kok ◽

Erik Lubberts ◽

Radjesh J. Bisoendial

Keyword(s):

Immune Response ◽

Endothelial Cells ◽

Single Cell ◽

Functional Diversity ◽

Immune Cell ◽

Inflammatory Diseases ◽

Lymphatic Endothelial Cells ◽

Chronic Inflammatory Diseases ◽

Single Cell Rna Sequencing ◽

Lymphatic Vasculature

Lymphatic endothelial cells (LECs) line the lymphatic vasculature and play a central role in the immune response. LECs have abilities to regulate immune transport, to promote immune cell survival, and to cross present antigens to dendritic cells. Single-cell RNA sequencing (scRNA) technology has accelerated new discoveries in the field of lymphatic vascular biology. This review will summarize these new findings in regard to embryonic development, LEC heterogeneity with associated functional diversity, and interactions with other cells. Depending on the organ, location in the lymphatic vascular tree, and micro-environmental conditions, LECs feature unique properties and tasks. Furthermore, adjacent stromal cells need the support of LECs for fulfilling their tasks in the immune response, such as immune cell transport and antigen presentation. Although aberrant lymphatic vasculature has been observed in a number of chronic inflammatory diseases, the knowledge on LEC heterogeneity and functional diversity in these diseases is limited. Combining scRNA sequencing data with imaging and more in-depth functional experiments will advance our knowledge of LECs in health and disease. Building the case, the LEC could be put forward as a new therapeutic target in chronic inflammatory diseases, counterweighting the current immune-cell focused therapies.

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