Journal: bioRxiv

Article Title: Lymphatic Endothelial Cells Regulate Neutrophil Phenotypes and Function in a Microphysiological Model of Infection

doi: 10.64898/2026.03.24.714048

Figure Lengend Snippet: (A) PDMS devices fabricated from soft lithography molds assembled onto MatTek glass-bottom culture dishes with integrated removable PDMS rods, and resultant lumen channels following rod removal after collagen polymerization. (B) Schematic showing the lumen seeding arrangement with one lumen seeded with human dermal lymphatic endothelial cells (HdLECs, yellow) to form a biomimetic lymphatic vessel and the adjacent lumen loaded with neutrophils (purple). pHrodo-conjugated bacterial bioparticles (green) are embedded within the surrounding collagen matrix. (C) Representative fluorescence image of a biomimetic lymphatic vessel formed within the collagen matrix.

Article Snippet: Adult primary human dermal lymphatic endothelial cells (HdLECs) were purchased from PromoCell (C-12217).

Techniques: Fluorescence

Journal: Materials Today Bio

Article Title: Three-dimensional modelling of lymphangiogenesis in-vitro using bioorthogonal click-crosslinked gelatin hydrogels

doi: 10.1016/j.mtbio.2025.102367

Figure Lengend Snippet: Three-dimensional (3D) distribution of HDLEC within GelTN hydrogels. (A) Orthogonal section view of Z-stack confocal images (700 μm) of HDLEC encapsulated in GelTN (6–12 % w/v), stained with calcein-AM, presented with depth coding to indicate the z-position (depth) of HDLEC within GelTN network. The scale bar indicates the depth range of 0–700 μm, as a reference for the z-axis positioning of HDLEC within GelTN hydrogels. (B) Schematic illustration of HDLEC encapsulation in GelTN hydrogel, followed by compatibility assessments. (C) Top view (X – Y plane) of z-stack confocal images of HDLEC embedded in GelTN (6–12 %) hydrogels for up to 14 days, stained with calcein AM (green) and propidium iodide (red). Scale bar = 200 μm.

Article Snippet: Primary human dermal lymphatic endothelial cells (HDLEC) isolated from single donor juvenile foreskin, were purchased from PromoCell® (Heidelberg, Germany).

Techniques: Staining, Encapsulation

Journal: Materials Today Bio

Article Title: Three-dimensional modelling of lymphangiogenesis in-vitro using bioorthogonal click-crosslinked gelatin hydrogels

doi: 10.1016/j.mtbio.2025.102367

Figure Lengend Snippet: Quantification of HDLEC network formation, viability and metabolic activity in response to 3D encapsulation in GelTN. (A) Percentage of viable cells after 24 h of encapsulation. (B) Quantification of (i) branches, (ii) isolated elements and (C) total network formation of HDLEC-laden in GelTN hydrogels (6–12 %) after 14 days. (D) Metabolic activity of HDLEC encapsulated in GelTN (6–12 %) at day 1, 5, 10 and 14, presented as relative fluorescence unit (RFU) of AlamarBlue. Data are presented as mean ± SD (n = 3).

Article Snippet: Primary human dermal lymphatic endothelial cells (HDLEC) isolated from single donor juvenile foreskin, were purchased from PromoCell® (Heidelberg, Germany).

Techniques: Activity Assay, Encapsulation, Isolation, Fluorescence

Journal: Materials Today Bio

Article Title: Three-dimensional modelling of lymphangiogenesis in-vitro using bioorthogonal click-crosslinked gelatin hydrogels

doi: 10.1016/j.mtbio.2025.102367

Figure Lengend Snippet: Sprouting of HDLEC spheroids in GelTN at different concentrations after 24 h of encapsulation . (A) Schematic illustration of HDLEC spheroid generation and encapsulation using hanging drop method, followed by spheroid encapsulation in GelTN. (B) Representative brightfield (BF) images of HDLEC spheroids (scale bar = 250 μm) and z-stack confocal images (projection of x – y planes) of HDLEC spheroids stained with Hoechst (Blue) and Phalloidin (actin filaments, red). Scale bar = 200 μm. (C) Schematic illustration of the quantification method of HDLEC spheroid sprouts. (D) The quantification of HDLEC spheroids (i) sprout length and (ii) number. ( E) Quantification of (i) actin intensity and (ii) nuclei count per sprout. The quantification was carried out on 6 random spheroids for each experimental group. Data are presented as mean ± SD (n = 3).

Article Snippet: Primary human dermal lymphatic endothelial cells (HDLEC) isolated from single donor juvenile foreskin, were purchased from PromoCell® (Heidelberg, Germany).

Techniques: Encapsulation, Staining

Journal: Materials Today Bio

Article Title: Three-dimensional modelling of lymphangiogenesis in-vitro using bioorthogonal click-crosslinked gelatin hydrogels

doi: 10.1016/j.mtbio.2025.102367

Figure Lengend Snippet: Sprouting of HDLEC spheroids encapsulated in GelTN_Lo (6 % w/v) and GelTN_Hi (12 % w/v) hydrogels in response to vascular endothelial growth factor C (VEGF-C). (A) BF images of HDLEC spheroids embedded in GelTN_Lo and GelTN_Hi, supplemented with complete media (CTRL), or serum starved media (0.1 % FCS) with different concentrations of VEGF-C (0, 50, 100, and 200 ng/mL). (B) Quantification of HDLEC spheroids sprout length (left) and number (right) was calculated for 6 randomly selected spheroids per experimental group. (C) Representative confocal images of HDLEC after 72 h of hydrogel encapsulation (nuclei = blue, actin = red; scale bar = 200 μm).

Article Snippet: Primary human dermal lymphatic endothelial cells (HDLEC) isolated from single donor juvenile foreskin, were purchased from PromoCell® (Heidelberg, Germany).

Techniques: Encapsulation

Journal: Materials Today Bio

Article Title: Three-dimensional modelling of lymphangiogenesis in-vitro using bioorthogonal click-crosslinked gelatin hydrogels

doi: 10.1016/j.mtbio.2025.102367

Figure Lengend Snippet: The effect of GelTN concentration on the hydrogel diffusion rate and mesh size and HDLEC on sprouting. GelTN_Lo and GelTN_Hi were prepared and incubated in 2 mg/mL FITC-Dextran (20 kDa) for up to 24 h with the (A) fluorescence measured post-digestion after 0, 2, 6 and 24 h, and (B) the percentage diffusion rate was calculated using the control hydrogel (containing 2 mg/mL FITC-Dextran). Statistical analysis represents the significant difference of diffusion rate between GelTN_Lo and GelTN_Hi. (C) Estimated mesh size of GelTN_Lo and GelTN_Hi (n = 3). (D) Bright field images of HDLEC spheroids encapsulated in: (Left) GelTN_Hi immersed in media containing 50 ng/mL VEGF-C, or (right) GelTN_Hi containing 50 ng/mL VEGF-C (GelTN_Hi_VC) for 24 h (scale bar = 100 μm). (E) The quantification of HDLEC sprout (i) number and (ii) length, embedded in GelTN_Hi and GelTN_Hi_VC compared to GelTN_Lo for 24 h.

Article Snippet: Primary human dermal lymphatic endothelial cells (HDLEC) isolated from single donor juvenile foreskin, were purchased from PromoCell® (Heidelberg, Germany).

Techniques: Concentration Assay, Diffusion-based Assay, Incubation, Fluorescence, Control, Hi-C

Journal: Materials Today Bio

Article Title: Three-dimensional modelling of lymphangiogenesis in-vitro using bioorthogonal click-crosslinked gelatin hydrogels

doi: 10.1016/j.mtbio.2025.102367

Figure Lengend Snippet: Mechanical regulation of HDLEC lymphangiogenic activity in GelTN_Lo and GelTN_Hi via matrix metalloproteases (MMPs) and integrins. (A) RT-qPCR of the relative expression of (i) MMP2 , MMP9 and MMP14 in HDLEC encapsulated in GelTN_Lo vs GelTN_Hi (n = 3; scale bar = 250 μm). (B) Brightfield images of HDLEC spheroids 24 h post embedding in GelTN_Lo supplemented with 50 ng/mL VEGF-C in the presence or absence of 50 μM Marimastat (MMPi). (C) Relative expression of α5 ( ITGα5 ), β1 ( ITGβ1 ), and β3 ( ITGβ3 ) integrins in HDLEC encapsulated in 1GelTN_Lo vs GelTN_Hi (n = 3). (D) Brightfield images of siCtrl and siβ3 transfected HDLEC spheroids encapsulated in GelTN_Lo, cultured in 50 ng/mL VEGF-C containing media (i and ii), or 50 ng/mL VEGF-C containing media with 20 μM K34C (iii and iv). Scale bar = 100–250 μm. (E) Quantification of HDLEC spheroid sprouts length (left) and number (right) was calculated for 9 randomly selected spheroids per experimental group. Data are presented as mean ± SD.

Article Snippet: Primary human dermal lymphatic endothelial cells (HDLEC) isolated from single donor juvenile foreskin, were purchased from PromoCell® (Heidelberg, Germany).

Techniques: Activity Assay, Quantitative RT-PCR, Expressing, Transfection, Cell Culture

Journal: Materials Today Bio

Article Title: Three-dimensional modelling of lymphangiogenesis in-vitro using bioorthogonal click-crosslinked gelatin hydrogels

doi: 10.1016/j.mtbio.2025.102367

Figure Lengend Snippet: Evaluation of GelTN hydrogel injectability and its potential as a growth factor delivery platform. (A) Schematic illustration of the mechanical test setup to measure the injection force of GelTN primed in 1 mL syringe and 26G needle. (B) Representative Force – Time plots of GelTN (6 % w/v), ejected at 0 then every 5 min at 20 min post-crosslinking up to 60 min (n = 5). Water was used as a control. (C) VEGF-C release from GelTN in comparison to VEGF-C in PBS (control; n = 3). (D) Percentage VEGF-C release determined by cumulative VEGF-C concentration divided by the initial total VEGF-C concentration, multiplied by 100 (n = 3). (E) Representative images of western blot analysis showing p-Akt (60 kDa) and p-ERK1/2 (44 and 42 kDA) levels in HDLEC treated with GelTN_VC, V_100, CM, SM and GF-free GelTN (NC), for 20- and 30-min. Total Akt and ERK1/2 levels measured in comparison to the phosphorylation levels. GAPDH (36 kDa) was used as a loading control (Left panel). Quantification of p-Akt/total-Akt ratio in HDLEC treated with supernatants from SM, GelTN, GelTN_VC, VC100 and CM, for 20 and 30 min (right panel, n = 4). (F) Metabolic activity of HDLEC after treatment with supernatants from GelTN_V hydrogels, GelTN hydrogels, and controls: CM, SM and 100 ng/mL VEGF-C (V_100) for 72 h. Percentage HDLEC metabolic activity relative to the SM control (n = 3). (G) Representative brightfield images of HDLEC spheroids encapsulated in GelTN and treated with SM, VC100 and GelTN_VC for 24 h (Scale bar = 200 μm). Data are presented as mean ± SD.

Article Snippet: Primary human dermal lymphatic endothelial cells (HDLEC) isolated from single donor juvenile foreskin, were purchased from PromoCell® (Heidelberg, Germany).

Techniques: Injection, Control, Comparison, Concentration Assay, Western Blot, Phospho-proteomics, Activity Assay