Journal: medRxiv

Article Title: Feasibility of Endothelial Cell Isolation from Routine Coronary Function Testing in ANOCA Patients

doi: 10.64898/2026.04.09.26350551

Figure Lengend Snippet: A) Method to isolate and culture ECs from catheterization material used during coronary function testing. B) Representative morphology (I, passage 0) and immunofluorescence images of cultured ECs (II and III, passage 5) showing positivity for VE-cadherin (II), von Willebrand Factor (vWF) (II) and CD31 (III). C) Flow-cytometric characterization of cultured ECs (passage 1) in comparison with multiple reference cell populations, including human dermal microvascular ECs (HDMVEC), human cardiac microvascular ECs (HCMEC), human coronary artery ECs (HCAEC), human plaque myofibroblasts and mesenchymal stem cells (MSC). The plotted histograms depict the ‘relative counts’ on the y-axis and the ‘relative intensity’ on the x-axis

Article Snippet: Reference populations included human dermal microvascular ECs (HDMVEC; iXCells Biotechnologies, REF#10HU-019), human cardiac microvascular ECs (HCMEC; Sigma-Aldrich, REF#C-12285), human coronary artery ECs (HCAEC; Lonza, REF#CC-2585), human plaque myofibroblasts and mesenchymal stem cells (MSC; Cell Therapy Facility, University Medical Center Utrecht; code: MSC053P3_AL-MSC071P3_R).

Techniques: Immunofluorescence, Cell Culture, Comparison

Journal: Journal of Biomedical Optics

Article Title: Functionalized erythrocyte-derived optical nanoparticles to target ephrin-B2 ligands

doi: 10.1117/1.JBO.24.8.085002

Figure Lengend Snippet: Ephrin-B2 Western blot expression. Western blot showing the expression of ephrin-B2 in hDMVECs infected with an ephrin-B2 lentivirus compared to the ephrin-B2 expression level of hDMVECs not infected with the ephrin-B2 lentivirus. Beta-actin levels are shown as the internal loading control.

Article Snippet: Human dermal microvascular endothelial cells (hDMVECs) were cultured in endothelial cell basal medium (ECBM) with growth supplement (Cell Applications, Inc.) as previously described.

Techniques: Western Blot, Expressing, Infection, Control

Journal: Journal of Biomedical Optics

Article Title: Functionalized erythrocyte-derived optical nanoparticles to target ephrin-B2 ligands

doi: 10.1117/1.JBO.24.8.085002

Figure Lengend Snippet: Cellular fluorescence of control and ephrin-B2 hDMVECs incubated with F-NETs at various ρ * values. (a) Fluorescent images of control and ephbin-B2 hDMVECs after 40 min of incubation at 4°C with F-NETs solutions. All images are falsely colored with the blue and red corresponding to DAPI and ICG NIR emission from the NETs, respectively. Scale bars are 30 μ m . (b) Averaged fluorescence intensity ( I ¯ ) [see Eq. (3)] as a function of ρ * . Cells from 3 to 4 images were analyzed, resulting in 16 to 41 measurements for each combination of nanoparticles and cells. Statistical significance of p < 0.001 is denoted by ***. Only statistically significant populations with the same ρ * value of F-NETs are indicated. (c) Sigmoidal fit to the I ¯ values of the ephrin-B2 hDMVECs versus the ρ * value of F-NETs. Error bars in (b) and (c) represent SDs.

Article Snippet: Human dermal microvascular endothelial cells (hDMVECs) were cultured in endothelial cell basal medium (ECBM) with growth supplement (Cell Applications, Inc.) as previously described.

Techniques: Fluorescence, Control, Incubation

Journal: Biofabrication

Article Title: A biopsy-sized 3D skin model with a perifollicular vascular plexus enables studying immune cell trafficking in the skin

doi: 10.1088/1758-5090/ad5d1a

Figure Lengend Snippet: Characterization of the vascular plexus in 3D-SoC. (A) The COMSOL model displays the calculated range of shear stresses in the vascular pattern. The design recapitulates the physiological range of shear stress levels found in the cutaneous capillaries, venules, and arterioles. We divide the vasculature into three shear rate zones as low-shear (vertical interconnecting channels), mid-shear (two outermost channels, top and bottom) and high-shear (two innermost, horizontal channels). (B) Imaging of the vascular network seeded with GFP-HDBECs confirms uniform coverage of the microchannel walls. Scale bar: 1 mm; (C) Immunofluorescent staining of primary HDBECs in 3D-SoC with VE-cadherin (VECAD; white). Scale bar: 5 µ m; (D) confocal imaging of the 3D-SoC seeded with HDBECs perfused with both 20 kDa and 40 kDa dextran at time zero and sixty minutes allowing for comparison of the permeability characteristics. Scale bar: 2 mm; (E) the graph shows increased leakage of dextran in the model without HDBECs (acellular control) for both molecular weights. (F) Time-lapse transport data integrated into a COMSOL model enabled the estimation of the average permeability of the vasculature. The permeability values were determined to be 0.62 µ m s −1 for 20 kDa and 0.41 µ m s −1 for 40 kDa respectively (** = p < 0.01).

Article Snippet: Human dermal blood endothelial cells (HDBECs) (PromoCell #C-12211) and GFP-tagged HDBECs (Angio-Proteomie #cAP-0005GFP-PM) were cultured up to passage 3 in Microvascular EC Growth Medium (PromoCell #C-22020).

Techniques: Shear, Imaging, Staining, Comparison, Permeability, Control

Journal: Biofabrication

Article Title: A biopsy-sized 3D skin model with a perifollicular vascular plexus enables studying immune cell trafficking in the skin

doi: 10.1088/1758-5090/ad5d1a

Figure Lengend Snippet: Incorporation and real-time monitoring of circulating T cells in 3D-SoC. (A) Schematic representation of the stages of T cell infiltration into human skin. (B) Live immunofluorescent images showing the naïve T cells labelled with CellTracker (red) on HDBECs in the first 1–2 min (left panel; the round morphology resembles the tethering/rolling stage); between 2–5 min (middle panel; the spread morphology resembles the firm adhesion stage); and between 5–15 min (right panel; the morphology and location relative to ECs resembles the diapedesis stage). The first two images show the top view, and the right-most image shows a cross-section of the 3D-SoC. Scale bars: 5 µ m; (C) High magnification image capturing a T cell (red) with its lamellipodia squeezing between two endothelial cells (green), resembling the morphology of T cells in vivo during their movement through capillary walls (namely diapedesis). Scale bar: 2 µ m; (D) characterization of Th1 cells polarized from Naïve T cells in vitro through flow cytometry showing expression of both Interferon γ and TNFα. (E) Comparison of the attachment of the T cells to the shear stress analysis for naive and Th1 cell population. (F) Total percentage of naïve T cells and Th1 cells retained after 5 and 10 mins of flow. (G) Percentage of cells retained for distinct shear zones; HS: high-shear, MS: mid-shear, LS: low-shear. (* = p < 0.05, ** = p < 0.01, *** = p < 0.005).

Article Snippet: Human dermal blood endothelial cells (HDBECs) (PromoCell #C-12211) and GFP-tagged HDBECs (Angio-Proteomie #cAP-0005GFP-PM) were cultured up to passage 3 in Microvascular EC Growth Medium (PromoCell #C-22020).

Techniques: In Vivo, In Vitro, Flow Cytometry, Expressing, Comparison, Shear

Journal: The Journal of investigative dermatology

Article Title: Nitric oxide-producing myeloid-derived suppressor cells inhibit vascular E- selectin expression in human squamous cell carcinomas

doi: 10.1038/jid.2012.190

Figure Lengend Snippet: (a,b) Co-staining of SCC cryosections demonstrated that iNOS + cells were evident in SCC tumors. (c,d) iNOS + cells lacked expression of the dendritic cell markers HLA-DR and CD11c, (e) the macrophage marker CD163, (f) the T cell marker CD3, and (g) the endothelial marker CD34. (h) iNOS + cells did express the myeloid marker CD11b. CD11b expression together with a lack of HLA-DR expression is a phenotype suggestive of MDSC. Similar findings were observed in a total of eight SCC tumors. Scale bar = 100 μM.

Article Snippet: Human dermal microvascular and umbilical vein endothelial cells (Lonza) were cultured in EGM-2 bulletkit growth medium (Lonza).

Techniques: Staining, Expressing, Marker

Journal: The Journal of investigative dermatology

Article Title: Nitric oxide-producing myeloid-derived suppressor cells inhibit vascular E- selectin expression in human squamous cell carcinomas

doi: 10.1038/jid.2012.190

Figure Lengend Snippet: (a) Human skin dermal microvascular cells (DMEC) were stimulated with either TLR7 agonist activated APC (APC), mimicking physiologic stimulation, or the potent endothelial activator TNFβ in the presence or absence of the NO donor spermine NONOate (NO). Treated cells were immunostained for E-selectin and analyzed by flow cytometry. NO potently suppressed E-selectin upregulation in response to stimulated APC but only partially inhibited expression when endothelial cells were directly stimulated with TNFβ. (b) Percent inhibition of E-selectin expression by NO after stimulation with APC or TNFβ. The mean and SEM of three different endothelial donors are shown. Human umbilical vein endothelial cells (HUVEC) responded similarly in that NO completely abrogated E-selectin expression in response to stimulated APC (APC+NO) but only partially inhibited expression after stimulation with TNFβ. (c-e) MDSC from SCC suppress endothelial E-selectin expression in vitro. (c) Human HUVEC were stimulated with TNFβ in the presence or absence of collagenase dispersed cells from SCC tumors. A modest but reproducible inhibition in E-selectin expression was observed. Representative histograms and individual results from four SCC tumors are shown, along with the mean and SEM of these measurements. (d,e) CD11b + MDSC from SCC tumors suppress endothelial E-selectin expression. CD11b + and CD11b − cell populations were obtained by magnetic bead separation from collagenase dispersed SCC tumors. HUVEC were stimulated with TNFβ in the presence or absence of CD11b + and CD11b − cells. (d) Representative histograms and (e) individual results from three SCC tumors are shown, along with the mean and SEM of these measurements. CD11b + MDSC reproducibly inhibited expression of endothelial E-selectin.

Article Snippet: Human dermal microvascular and umbilical vein endothelial cells (Lonza) were cultured in EGM-2 bulletkit growth medium (Lonza).

Techniques: Flow Cytometry, Expressing, Inhibition, In Vitro

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