scholarly journals Investigating lymphangiogenesis in vitro and in vivo using engineered human lymphatic vessel networks

2021 ◽  
Vol 118 (31) ◽  
pp. e2101931118
Author(s):  
Shira Landau ◽  
Abigail Newman ◽  
Shlomit Edri ◽  
Inbal Michael ◽  
Shahar Ben-Shaul ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Vessel ◽  
Blood Vessels ◽  
Lymphatic Vessel ◽  
Lymphatic Vessels ◽  
Lymphatic Endothelial Cells ◽  
Supporting Cells ◽  
Vessel Formation ◽  
Vessel Networks

The lymphatic system is involved in various biological processes, including fluid transport from the interstitium into the venous circulation, lipid absorption, and immune cell trafficking. Despite its critical role in homeostasis, lymphangiogenesis (lymphatic vessel formation) is less widely studied than its counterpart, angiogenesis (blood vessel formation). Although the incorporation of lymphatic vasculature in engineered tissues or organoids would enable more precise mimicry of native tissue, few studies have focused on creating engineered tissues containing lymphatic vessels. Here, we populated thick collagen sheets with human lymphatic endothelial cells, combined with supporting cells and blood endothelial cells, and examined lymphangiogenesis within the resulting constructs. Our model required just a few days to develop a functional lymphatic vessel network, in contrast to other reported models requiring several weeks. Coculture of lymphatic endothelial cells with the appropriate supporting cells and intact PDGFR-β signaling proved essential for the lymphangiogenesis process. Additionally, subjecting the constructs to cyclic stretch enabled the creation of complex muscle tissue aligned with the lymphatic and blood vessel networks, more precisely biomimicking native tissue. Interestingly, the response of developing lymphatic vessels to tensile forces was different from that of blood vessels; while blood vessels oriented perpendicularly to the stretch direction, lymphatic vessels mostly oriented in parallel to the stretch direction. Implantation of the engineered lymphatic constructs into a mouse abdominal wall muscle resulted in anastomosis between host and implant lymphatic vasculatures, demonstrating the engineered construct's potential functionality in vivo. Overall, this model provides a potential platform for investigating lymphangiogenesis and lymphatic disease mechanisms.

Angiopoietin-1 promotes LYVE-1-positive lymphatic vessel formation

Blood ◽  
2005 ◽  
Vol 105 (12) ◽  
pp. 4649-4656 ◽  
Author(s):  
Tohru Morisada ◽  
Yuichi Oike ◽  
Yoshihiro Yamada ◽  
Takashi Urano ◽  
Masaki Akao ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Lymphatic Vessel ◽  
Matrix Protein ◽  
Lymphatic Vessels ◽  
Vascular Endothelial ◽  
Vessel Formation ◽  
Lymphatic Vasculature ◽  
In Vitro Effects

Abstract Angiopoietin (Ang) signaling plays a role in angiogenesis and remodeling of blood vessels through the receptor tyrosine kinase Tie2, which is expressed on blood vessel endothelial cells (BECs). Recently it has been shown that Ang-2 is crucial for the formation of lymphatic vasculature and that defects in lymphangiogenesis seen in Ang-2 mutant mice are rescued by Ang-1. These findings suggest important roles for Ang signaling in the lymphatic vessel system; however, Ang function in lymphangiogenesis has not been characterized. In this study, we reveal that lymphatic vascular endothelial hyaluronan receptor 1-positive (LYVE-1+) lymphatic endothelial cells (LECs) express Tie2 in both embryonic and adult settings, indicating that Ang signaling occurs in lymphatic vessels. Therefore, we examined whether Ang-1 acts on in vivo lymphatic angiogenesis and in vitro growth of LECs. A chimeric form of Ang-1, cartilage oligomeric matrix protein (COMP)-Ang-1, promotes in vivo lymphatic angiogenesis in mouse cornea. Moreover, we found that COMP-Ang-1 stimulates in vitro colony formation of LECs. These Ang-1-induced in vivo and in vitro effects on LECs were suppressed by soluble Tie2-Fc fusion protein, which acts as an inhibitor by sequestering Ang-1. On the basis of these observations, we propose that Ang signaling regulates lymphatic vessel formation through Tie2. (Blood. 2005;105:4649-4656)

scholarly journals A novel pro-lymphangiogenic function for Th17/IL-17

Blood ◽  
2011 ◽  
Vol 118 (17) ◽  
pp. 4630-4634 ◽  
Author(s):  
Sunil K. Chauhan ◽  
Yiping Jin ◽  
Sunali Goyal ◽  
Hyun Soo Lee ◽  
Thomas A. Fuchsluger ◽  
...  
Keyword(s):  
Cell Culture ◽  
Endothelial Cells ◽  
Autoimmune Diseases ◽  
Blood Vessels ◽  
Th17 Cells ◽  
Lymphatic Vessels ◽  
Preclinical Model ◽  
Lymphatic Endothelial Cells ◽  
Corneal Lymphangiogenesis

Abstract Th17 cells, in addition to their proinflammatory functions, have been recognized as potent inducers of angiogenesis in autoimmune diseases and malignancies. In the present study, we demonstrate distinct mechanisms by which IL-17 induces lymphangiogenesis. Using the mouse cornea micropocket and cell culture assays, our data demonstrate that IL-17 directly promotes growth of lymphatic vessels by inducing increased expression of prolymphangiogenic VEGF-D and proliferation of lymphatic endothelial cells. However, IL-17–induced growth of blood vessels is primarily mediated through IL-1β secretion by IL-17–responsive cells. Furthermore, in vivo blockade of IL-17 in a preclinical model of Th17-dominant autoimmune ocular disease demonstrates a significant reduction in the corneal lymphangiogenesis and in the progression of clinical disease. Taken together, our findings demonstrate a novel prolymphangiogenic function for Th17/IL-17, indicating that IL-17 can promote the progression and amplification of immunity in part through its induction of lymphangiogenesis.

scholarly journals Generation of specialized blood vessels through transdifferentiation of lymphatic endothelial cells

2021 ◽  
Author(s):  
Rudra N. Das ◽  
Ivan Bassi ◽  
Yanchao Han ◽  
Giuseppina Lambiase ◽  
Yaara Tevet ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Vessel ◽  
Blood Vessels ◽  
Cell Fate ◽  
Lymphatic Vessels ◽  
Developmental Trajectory ◽  
Negative Regulator ◽  
Blood Vessel Formation ◽  
Vessel Formation ◽  
Cell Ontogeny

AbstractThe lineage and developmental trajectory of a cell are key determinants of cellular identity. Yet, the functional relevance of deriving a specific cell type from ontologically distinct progenitors, remains an open question. In the case of the vascular system, blood and lymphatic vessels are composed of endothelial cells (ECs) that differentiate and diversify to cater the different physiological demands of each organ. While lymphatic vessels have been shown to originate from multiple cell sources, lymphatic ECs (LECs) themselves seem to have a unipotent cell fate. In this work we uncover a novel mechanism of blood vessel formation through transdifferentiation of LECs. Using advanced long-term reiterative imaging and lineage-tracing of ECs in zebrafish, from embryonic development through adulthood, we reveal a hitherto unknown process of LEC-to-BEC transdifferentiation, underlying vascularization of the anal fin (AF). Moreover, we demonstrate distinct functional implications for deriving AF vessels from either LECs or BECs, uncovering for the first time a clear link between cell ontogeny and functionality. Molecularly, we identify Sox17 as a negative regulator of lymphatic fate specification, whose specific expression in AF LECs suppresses its lymphatic cell fate. Finally, we show that akin to the developmental process, during adult AF regeneration the vasculature is re-derived from lymphatics, demonstrating that LECs in the mature fish retain both potency and plasticity for generating specialized blood vessels. Overall, our work highlights a novel mechanism of blood vessel formation through LEC trans-differentiation, and provides the first in vivo evidence for a link between cell ontogeny and functionality in ECs.

scholarly journals Lymphovenous hemostasis and the role of platelets in regulating lymphatic flow and lymphatic vessel maturation

Blood ◽  
2016 ◽  
Vol 128 (9) ◽  
pp. 1169-1173 ◽  
Author(s):  
John D. Welsh ◽  
Mark L. Kahn ◽  
Daniel T. Sweet
Keyword(s):  
Endothelial Cells ◽  
Lymphatic Vessel ◽  
Vascular System ◽  
Lymphatic Vessels ◽  
Lymph Flow ◽  
The Body ◽  
Lymphatic Endothelial Cells ◽  
Lymphatic Circulation ◽  
Vessel Maturation

Abstract Aside from the established role for platelets in regulating hemostasis and thrombosis, recent research has revealed a discrete role for platelets in the separation of the blood and lymphatic vascular systems. Platelets are activated by interaction with lymphatic endothelial cells at the lymphovenous junction, the site in the body where the lymphatic system drains into the blood vascular system, resulting in a platelet plug that, with the lymphovenous valve, prevents blood from entering the lymphatic circulation. This process, known as “lymphovenous hemostasis,” is mediated by activation of platelet CLEC-2 receptors by the transmembrane ligand podoplanin expressed by lymphatic endothelial cells. Lymphovenous hemostasis is required for normal lymph flow, and mice deficient in lymphovenous hemostasis exhibit lymphedema and sometimes chylothorax phenotypes indicative of lymphatic insufficiency. Unexpectedly, the loss of lymph flow in these mice causes defects in maturation of collecting lymphatic vessels and lymphatic valve formation, uncovering an important role for fluid flow in driving endothelial cell signaling during development of collecting lymphatics. This article summarizes the current understanding of lymphovenous hemostasis and its effect on lymphatic vessel maturation and synthesizes the outstanding questions in the field, with relationship to human disease.

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

Blood ◽  
2010 ◽  
Vol 115 (4) ◽  
pp. 906-909 ◽  
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.

scholarly journals Induction of lymphatic endothelial cell differentiation in embryoid bodies

Blood ◽  
2006 ◽  
Vol 107 (3) ◽  
pp. 1214-1216 ◽  
Author(s):  
Ruediger Liersch ◽  
Filip Nay ◽  
Lingge Lu ◽  
Michael Detmar
Keyword(s):  
Endothelial Cells ◽  
Lymphatic Vessel ◽  
Molecular Mechanisms ◽  
Vascular System ◽  
Es Cells ◽  
Embryonic Stem ◽  
Lymphatic Endothelial Cell ◽  
Embryoid Bodies ◽  
Lymphatic Endothelial Cells ◽  
Vessel Formation

AbstractThe molecular mechanisms that regulate the formation of the lymphatic vascular system remain poorly characterized. Whereas studies in embryonic stem (ES) cells have provided major new insights into the mechanisms of blood vessel formation, the development of lymphatic endothelium has not been previously observed. We established embryoid bodies (EBs) from murine ES cells in the presence or absence of lymphangiogenic growth factors. We found that lymphatic endothelial cells develop at day 18 after EB formation. These cells express CD31 and the lymphatic lineage markers Prox-1 and Lyve-1, but not the vascular marker MECA-32, and they frequently sprout from preexisting blood vessels. Lymphatic vessel formation was potently promoted by VEGF-A and VEGF-C but not by bFGF. Our results reveal, for the first time, that ES cells can differentiate into lymphatic endothelial cells, and they identify the EB assay as a powerful new tool to dissect the molecular mechanisms that control lymphatic vessel formation.

scholarly journals Loss of Primary Cilia Protein IFT20 Dysregulates Lymphatic Vessel Patterning in Development and Inflammation

2021 ◽  
Vol 9 ◽  
Author(s):  
Delayna Paulson ◽  
Rebecca Harms ◽  
Cody Ward ◽  
Mackenzie Latterell ◽  
Gregory J. Pazour ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Lymphatic Vessel ◽  
Primary Cilia ◽  
Fluid Shear Stress ◽  
Lymphatic Vessels ◽  
Intraflagellar Transport ◽  
Membrane Receptors ◽  
Lymphatic Endothelial Cells ◽  
Corneal Lymphangiogenesis

Microenvironmental signals produced during development or inflammation stimulate lymphatic endothelial cells to undergo lymphangiogenesis, in which they sprout, proliferate, and migrate to expand the vascular network. Many cell types detect changes in extracellular conditions via primary cilia, microtubule-based cellular protrusions that house specialized membrane receptors and signaling complexes. Primary cilia are critical for receipt of extracellular cues from both ligand-receptor pathways and physical forces such as fluid shear stress. Here, we report the presence of primary cilia on immortalized mouse and primary adult human dermal lymphatic endothelial cells in vitro and on both luminal and abluminal domains of mouse corneal, skin, and mesenteric lymphatic vessels in vivo. The purpose of this study was to determine the effects of disrupting primary cilia on lymphatic vessel patterning during development and inflammation. Intraflagellar transport protein 20 (IFT20) is part of the transport machinery required for ciliary assembly and function. To disrupt primary ciliary signaling, we generated global and lymphatic endothelium-specific IFT20 knockout mouse models and used immunofluorescence microscopy to quantify changes in lymphatic vessel patterning at E16.5 and in adult suture-mediated corneal lymphangiogenesis. Loss of IFT20 during development resulted in edema, increased and more variable lymphatic vessel caliber and branching, as well as red blood cell-filled lymphatics. We used a corneal suture model to determine ciliation status of lymphatic vessels during acute, recurrent, and tumor-associated inflammatory reactions and wound healing. Primary cilia were present on corneal lymphatics during all of the mechanistically distinct lymphatic patterning events of the model and assembled on lymphatic endothelial cells residing at the limbus, stalk, and vessel tip. Lymphatic-specific deletion of IFT20 cell-autonomously exacerbated acute corneal lymphangiogenesis resulting in increased lymphatic vessel density and branching. These data are the first functional studies of primary cilia on lymphatic endothelial cells and reveal a new dimension in regulation of lymphatic vascular biology.

scholarly journals Human Blood Vessel Organoids Penetrate Human Cerebral Organoids and Form a Vessel-Like System

Cells ◽  
2021 ◽  
Vol 10 (8) ◽  
pp. 2036
Author(s):  
Yujin Ahn ◽  
Ju-Hyun An ◽  
Hae-Jun Yang ◽  
Dong Gil Lee ◽  
Jieun Kim ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Vessel ◽  
Blood Brain Barrier ◽  
Human Blood ◽  
Cell Types ◽  
Brain Barrier ◽  
Vascular Cells ◽  
Blood Brain ◽  
Vessel Networks

Vascularization of tissues, organoids and organ-on-chip models has been attempted using endothelial cells. However, the cultured endothelial cells lack the capacity to interact with other somatic cell types, which is distinct from developing vascular cells in vivo. Recently, it was demonstrated that blood vessel organoids (BVOs) recreate the structure and functions of developing human blood vessels. However, the tissue-specific adaptability of BVOs had not been assessed in somatic tissues. Herein, we investigated whether BVOs infiltrate human cerebral organoids and form a blood–brain barrier. As a result, vascular cells arising from BVOs penetrated the cerebral organoids and developed a vessel-like architecture composed of CD31+ endothelial tubes coated with SMA+ or PDGFR+ mural cells. Molecular markers of the blood-brain barrier were detected in the vascularized cerebral organoids. We revealed that BVOs can form neural-specific blood-vessel networks that can be maintained for over 50 days.

scholarly journals Vasculogenesis and angiogenesis in modular collagen–fibrin microtissues

2014 ◽  
Vol 2 (10) ◽  
pp. 1497-1508 ◽  
Author(s):  
A. W. Peterson ◽  
D. J. Caldwell ◽  
A. Y. Rioja ◽  
R. R. Rao ◽  
A. J. Putnam ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Blood Vessel ◽  
Tissue Development ◽  
Blood Vessel Formation ◽  
Vessel Formation ◽  
Surrounding Matrix ◽  
Vasculogenesis And Angiogenesis ◽  
Vessel Networks

Vessel networks can be generated within modular protein microbeads containing endothelial cells and fibroblasts. Embedding these microtissues in a surrounding matrix emulates aspects of new blood vessel formation, a process that is critical in tissue development, remodeling, and regeneration.

scholarly journals Inflammation induces lymphangiogenesis through up-regulation of VEGFR-3 mediated by NF-κB and Prox1

Blood ◽  
2010 ◽  
Vol 115 (2) ◽  
pp. 418-429 ◽  
Author(s):  
Michael J. Flister ◽  
Andrew Wilber ◽  
Kelly L. Hall ◽  
Caname Iwata ◽  
Kohei Miyazono ◽  
...  
Keyword(s):  
Endothelial Cells ◽  
Time Course ◽  
Molecular Mechanisms ◽  
Lymphatic Vessels ◽  
Growth Factor Receptor ◽  
Receptor Expression ◽  
Lymphatic Endothelial Cells ◽  
Inflammatory Stimuli ◽  
And Migration

Abstract The concept of inflammation-induced lymphangiogenesis (ie, formation of new lymphatic vessels) has long been recognized, but the molecular mechanisms remained largely unknown. The 2 primary mediators of lymphangiogenesis are vascular endothelial growth factor receptor-3 (VEGFR-3) and Prox1. The key factors that regulate inflammation-induced transcription are members of the nuclear factor-kappaB (NF-κB) family; however, the role of NF-κB in regulation of lymphatic-specific genes has not been defined. Here, we identified VEGFR-3 and Prox1 as downstream targets of the NF-κB pathway. In vivo time-course analysis of inflammation-induced lymphangiogenesis showed activation of NF-κB followed by sequential up-regulation of Prox1 and VEGFR-3 that preceded lymphangiogenesis by 4 and 2 days, respectively. Activation of NF-κB by inflammatory stimuli also elevated Prox1 and VEGFR-3 expression in cultured lymphatic endothelial cells, resulting in increased proliferation and migration. We also show that Prox1 synergizes with the p50 of NF-κB to control VEGFR-3 expression. Collectively, our findings suggest that induction of the NF-κB pathway by inflammatory stimuli activates Prox1, and both NF-κB and Prox1 activate the VEGFR-3 promoter leading to increased receptor expression in lymphatic endothelial cells. This, in turn, enhances the responsiveness of preexisting lymphatic endothelium to VEGFR-3 binding factors, VEGF-C and VEGF-D, ultimately resulting in robust lymphangiogenesis.

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