Journal: Microcirculation (New York, N.Y. : 1994)

Article Title: A bioengineered lymphatic vessel model for studying lymphatic endothelial cell-cell junction and barrier function

doi: 10.1111/micc.12730

Figure Lengend Snippet: (A) A schematic of an organotypic 3D lymphatic vessel model (LV-on-chip). Prox-1 (green) and CD31 (red) expression confirms lymphatic endothelial identity and cell morphology in the channel. (B) Morphologic changes in human dermal microvascular blood endothelial cells (BECs) with lymphatic endothelial cells (LECs) after one day of cell seeding. BECs become more contractile than LECs, forming a smaller vessel diameter compared to LECs. (C) BVs and LVs observed in mouse ear tissues. mLYVE-1, anti-mouse LYVE-1 antibody; mCD31, anti-mouse CD31 antibody. (D) Phalloidin (red) and anti-VE-cad (VE-cadherin) antibody (green) staining to visualize F-actin and adherens junctions. (E) Lymphatic and blood vessel barrier function. 70 kDa dextran was introduced into the vessel lumens and dextran diffusion was observed in real time under microscopy. Superimposed red dashed lines represent the edges of the vessel lumens. (F) Quantification of the permeability of BEC-generated engineered BVs and LEC-generated LVs. ** p = 0.0016, two tailed unpaired Student t-test, n = 5 per group. Data are expressed as mean ± S.E.M.

Article Snippet: In the hollow channel, we seeded human dermal microvascular lymphatic endothelial cells (LECs) to form a biomimetic lymphatic vessel ( ).

Techniques: Expressing, Staining, Diffusion-based Assay, Microscopy, Permeability, Generated, Two Tailed Test

Journal: Microcirculation (New York, N.Y. : 1994)

Article Title: A bioengineered lymphatic vessel model for studying lymphatic endothelial cell-cell junction and barrier function

doi: 10.1111/micc.12730

Figure Lengend Snippet: (A) Lymphatic endothelial cells (LECs) in different ECM hydrogels (2D): 2.5 mg/ml collagen 1, 2.5 mg/ml collagen 1 and 150 μg/ml Fibronectin, and no gel (plastic). F-actin and VE-cad were visualized to assess cytoskeletal arrangement and adherens junction formation in each condition. (B) Quantification of the relative junction area was performed, illustrating a significantly lower junction area in cells grown on the 2.5 mg/ml collagen 1 compared to the cells grown directly on plastic. ** p = 0.0017 (Collagen 1 vs. plastic); higher junction area in cells grown on the 2.5 mg/ml collagen 1 + fibronectin compared to the cells grown on collagen 1. * p = 0.0151 (Collagen 1 + fibronectin vs. Collagen 1); not-significant (ns) p = 0.5292 (Collagen 1 + fibronectin vs plastic). One-way ANOVA with Tukey’s HSD tests , n = 6 per group. Data are expressed as mean ± S.E.M. (C) Dynamics of fibronectin on LECs in collagen 1 or collagen 1 + fibronectin gel. On collagen 1 gel, LEC islands with VE-cad expression lacks fibronectin expression. On collagen 1 + fibronectin, fibronectin connects separate LEC islands. (D) At day 4 on Collagen 1 + fibronectin, LECs showed tightened junctions and fibronectin was localized in the junctional area.

Article Snippet: In the hollow channel, we seeded human dermal microvascular lymphatic endothelial cells (LECs) to form a biomimetic lymphatic vessel ( ).

Techniques: Expressing

Journal: Microcirculation (New York, N.Y. : 1994)

Article Title: A bioengineered lymphatic vessel model for studying lymphatic endothelial cell-cell junction and barrier function

doi: 10.1111/micc.12730

Figure Lengend Snippet: (A) Activated integrin α5 was visualized in both ECM composition conditions by using anti-integrin α5 antibody (clone: SNAKA51) that can only detect the activated form of the integrin α5. F-actin was also observed in these conditions. (B) LECs in Collagen 1 were pre-treated with anti-integrin α5 antibodies (clone: SNAKA51) antibodies to activate integrin α5 in LECs. The fixed samples were stained with anti-VE-cadherin antibodies, anti-JAM-A antibodies, and phalloidin to visualize adherens junctions and F-actin. (C) Quantification of the relative junction area was performed, illustrating a significantly higher junction area in integrin α5 activated cells compared to the control LECs. ** p = 0.0020; Two tailed unpaired Student t-test, n = 6 per group. Data are expressed as mean ± S.E.M. (D) Control LECs or LECs with activated integrin α5 were seeded in LV-on-chip and cultured for 3 days on the rocking platform. 70 kDa dextran was introduced to the lymphatic lumens. Dextran diffusion was observed at 0 and 1 minutes under microscopy. Superimposed red dashed lines represent the edges of the vessel lumens. (E) Quantification of the permeability of LEC-generated engineered LVs in collagen 1 with and without integrin α5 activation. ** p = 0.0021. Two tailed unpaired Student t-test, n = 5 per group. Data are expressed as mean ± S.E.M. (F) This table summarizes our findings regarding LEC permeability and integrin α5 activity. LVs grown in Collagen 1 without any activator treatment showed high LEC permeability and low integrin α5 activity. In contrast, LVs grown in either Collagen 1 + Fibronectin or LVs grown in only Collagen 1 with integrin α5 activator pre-treatment both showed low LEC permeability and high integrin α5 activity.

Article Snippet: In the hollow channel, we seeded human dermal microvascular lymphatic endothelial cells (LECs) to form a biomimetic lymphatic vessel ( ).

Techniques: Staining, Control, Two Tailed Test, Cell Culture, Diffusion-based Assay, Microscopy, Permeability, Generated, Activation Assay, Activity Assay

Journal: PLoS ONE

Article Title: High Mobility Group Box-1 Promotes Inflammation-Induced Lymphangiogenesis via Toll-Like Receptor 4-Dependent Signalling Pathway

doi: 10.1371/journal.pone.0154187

Figure Lengend Snippet: (A): HMGB1 promoted VEGF-C-induced HDLECs proliferation in a dose-dependent manner. (B): TLR4 mediates HMGB1-induced LECs proliferation. (C-E): TLR4 mediates HMGB1-induced LECs tube formation.* p < 0.05, ** p < 0.01, *** p < 0.001

Article Snippet: Human dermal lymphatic endothelial cells (HDLECs) were purchased from ScienCell (Carlsbad, CA) and maintained in endothelial cell basal medium-2 with growth supplements (EBM-2 MV).

Techniques:

Journal: The Journal of Biological Chemistry

Article Title: Galectin-1 Regulates Tissue Exit of Specific Dendritic Cell Populations *

doi: 10.1074/jbc.M115.644799

Figure Lengend Snippet: Human lymphatic endothelial cells express and secrete galectin-1. Sections of skin from lymphedema patients were stained with polyclonal antibody against galectin-1 ( top ) or monoclonal antibody to the human LEC marker podoplanin ( bottom ). Bound antibody was detected with the corresponding secondary antibody and visualized using a 3-amino-9-ethylcarbazole chromogenic substrate system. Sections were counterstained with hematoxylin. Insets ( middle column ) show control antibody staining. Dilated lymphatic vessels are lined by LECs expressing galectin-1 ( arrow, top ) and podoplanin ( arrow, bottom ). Data are representative of six independent tissue samples. Note that the distribution of galectin-1 on LECs appears more dispersed than that of podoplanin, suggesting the localization of secreted galectin-1 in extracellular matrix ( arrowhead , top right panel ). Magnification is as follows: ×20 ( left ), ×40 ( middle ), and ×100 ( right ). Scale bar, 100 μm ( left ), 50 μm ( middle ), and 20 μm ( right ).

Article Snippet: Human dermal lymphatic endothelial cells (HMCV-DLyAd-Der Lym Endo) were purchased from Lonza (Walkersville) and maintained in EGM TM -2MV medium (Lonza) as described ( ).

Techniques: Staining, Marker, Control, Expressing