Journal: Materials Today Bio

Article Title: Synergistic aligned neuronal and vascular growth inside 3D-PEG-Anisogels utilizing a triple-co-culture

doi: 10.1016/j.mtbio.2025.102737

Figure Lengend Snippet: Effect of NGF on the neurite length and the vascular-like structure formation in a co-culture of HUVECs and MSCs, or a single culture of a DRG after 7 days of culture inside the PEG-based hydrogels. A, B: Representative image of immunofluorescence staining for PECAM/CD31 (green) of structures formed by MSC:HUVEC co-cultures (1000 cells/μL, ratio 3:1) in a 1.25 w/v% hydrogel with 1 μM FN in EGM-2 without NGF (A) and with NGF (B); C,D: Representative image of immunofluorescence staining for NF-H (red) of DRG neurites in a 1.25 w/v% hydrogel with 1 μM FN cultured in EGM-2 without NGF (D) and with NGF (E); E, F: 3D Analysis of the total length of vascular-like structures (E) and of DRG neurites (F); Scale bar: 500 μm. n: 3–8 replicates per condition. (p∗ < 0.05). Error bars are given as standard error. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Human umbilical vein endothelial cells (HUVECs, passage 1–5, pooled donors, Lonza) were cultured in tissue culture flasks in endothelial growth medium (EGM-2 ready-to-use kit, Promocell) supplemented with FBS (2.00 w/v%), epidermal growth factor (recombinant human, 5 ng/mL), basic fibroblast growth factor (recombinant human, 10.0 ng/mL), insulin-like growth factor (Long R3 IGF, 20 ng/mL), vascular endothelial growth factor 165 (recombinant human, 0.5 ng/mL), ascorbic acid (1 μg/mL), heparin (22.5 μg/mL), and hydrocortisone (0.20 μg/mL).

Techniques: Co-Culture Assay, Immunofluorescence, Staining, Cell Culture

Journal: Materials Today Bio

Article Title: Synergistic aligned neuronal and vascular growth inside 3D-PEG-Anisogels utilizing a triple-co-culture

doi: 10.1016/j.mtbio.2025.102737

Figure Lengend Snippet: Effect of different proteins on vascular-like structures and single DRG. A,B: Respective images of immunofluorescence staining for PECAM/CD-31 (green) of structures formed by MSC:HUVEC co-cultures (1000 cells/μL, ratio 3 MSCs:1 HUVEC) with 300 μM RGD (A) and 200 μM IKVAV (B) C,D: Respective image of immunofluorescence staining for NF-H (red) of DRG neurites with 300 μM RGD (D) and 200 μM IKVAV (E); E, F: 3D Analysis of the total length vascular-like structures (E) and of DRG neurites (F); The PEG-QK gels were processed and analyzed after culturing for 7 days in EGM-2+NGF. Scale bar: 1 mm. n: 3–10 replicates per condition. (p∗ < 0.05). Error bars are given as standard error. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Human umbilical vein endothelial cells (HUVECs, passage 1–5, pooled donors, Lonza) were cultured in tissue culture flasks in endothelial growth medium (EGM-2 ready-to-use kit, Promocell) supplemented with FBS (2.00 w/v%), epidermal growth factor (recombinant human, 5 ng/mL), basic fibroblast growth factor (recombinant human, 10.0 ng/mL), insulin-like growth factor (Long R3 IGF, 20 ng/mL), vascular endothelial growth factor 165 (recombinant human, 0.5 ng/mL), ascorbic acid (1 μg/mL), heparin (22.5 μg/mL), and hydrocortisone (0.20 μg/mL).

Techniques: Immunofluorescence, Staining

Journal: Materials Today Bio

Article Title: Synergistic aligned neuronal and vascular growth inside 3D-PEG-Anisogels utilizing a triple-co-culture

doi: 10.1016/j.mtbio.2025.102737

Figure Lengend Snippet: Effect of different cell adhesive peptides and hydrogel stiffness on vascular-like structure formation and neurite growth using a tri-culture. A–F: Respective images of immunofluorescence staining for PECAM/CD-31 (green) of structures formed by HUVECs (1000 cells/μL, ratio 3 MSCs:1 HUVEC) with 200 μM IKVAV in a 1.25 w/v% (A) and a 2.00 w/v% (D) hydrogel, for NF-H (red) of DRG neurites in a 1.25 w/v% (B) and a 2.00 w/v% Hydrogel (E), and their respective overlays (C and F). G: 3D Analysis of the total length of vascular-like structures; H: 3D Analysis of the total length of DRG. The PEG-QK gels were processed and analyzed after 7 days of culture in EGM-2+NGF. Scale bar: 1 mm. n: 3–10 replicates per condition. Error bars are given as standard error. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Human umbilical vein endothelial cells (HUVECs, passage 1–5, pooled donors, Lonza) were cultured in tissue culture flasks in endothelial growth medium (EGM-2 ready-to-use kit, Promocell) supplemented with FBS (2.00 w/v%), epidermal growth factor (recombinant human, 5 ng/mL), basic fibroblast growth factor (recombinant human, 10.0 ng/mL), insulin-like growth factor (Long R3 IGF, 20 ng/mL), vascular endothelial growth factor 165 (recombinant human, 0.5 ng/mL), ascorbic acid (1 μg/mL), heparin (22.5 μg/mL), and hydrocortisone (0.20 μg/mL).

Techniques: Adhesive, Immunofluorescence, Staining

Journal: Materials Today Bio

Article Title: Synergistic aligned neuronal and vascular growth inside 3D-PEG-Anisogels utilizing a triple-co-culture

doi: 10.1016/j.mtbio.2025.102737

Figure Lengend Snippet: Effect of alignment on the triculture. A–C: Respective images for immunofluorescence staining for NF-H (red) of DRGs and for PECAM/CD-31 (green) of structures formed by HUVECs (1000 cells/μL, ratio 3 MSCs:1 HUVEC) in a control hydrogel (A), a hydrogel with randomly oriented 1.0 v/v% 2.5x2.5 × 50 μm 3 microgels (B) and Anisogel with 1.0 v/v% 2.5x2.5 × 50 μm 3 microgels (C) with 1.25 w/v% PEG and 200 μM IKVAV; D–F: Respective radial plots for the depicted images' directionality of neurites and vascular-like structures and microgels for control (D), random (E) and aligned (F) conditions with 1.25 w/v% PEG; G–I: Respective images for immunofluorescence staining for NF-H (red) of DRG neurites and for PECAM/CD-31 (green) of structures formed by HUVECs (1000 cells/μL, ratio 3 MSCs: 1 HUVEC) in a control hydrogel (G), a hydrogel with randomly oriented 1.0 v/v% 2.5x2.5 × 50 μm 3 microgels (H) and Anisogel with 1.0 v/v% 2.5x2.5 × 50 μm 3 microgels (I) with 2.00 w/v% PEG and 200 μM IKVAV J: 3D Analysis of the total length of vascular-like structures. K: 3D Analysis of total neurite length from DRGs. The PEG-QK gels were processed and analyzed after culturing for 7 days in EGM-2+NGF. Scale bar: 1 mm. n: 7–12 replicates per condition (p∗∗ < 0.01, p∗∗∗ < 0.001). Error bars are given as standard error. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Human umbilical vein endothelial cells (HUVECs, passage 1–5, pooled donors, Lonza) were cultured in tissue culture flasks in endothelial growth medium (EGM-2 ready-to-use kit, Promocell) supplemented with FBS (2.00 w/v%), epidermal growth factor (recombinant human, 5 ng/mL), basic fibroblast growth factor (recombinant human, 10.0 ng/mL), insulin-like growth factor (Long R3 IGF, 20 ng/mL), vascular endothelial growth factor 165 (recombinant human, 0.5 ng/mL), ascorbic acid (1 μg/mL), heparin (22.5 μg/mL), and hydrocortisone (0.20 μg/mL).

Techniques: Immunofluorescence, Staining, Control

Journal: Materials Today Bio

Article Title: Synergistic aligned neuronal and vascular growth inside 3D-PEG-Anisogels utilizing a triple-co-culture

doi: 10.1016/j.mtbio.2025.102737

Figure Lengend Snippet: Effect of alignment on the triculture with single neurons. A, B: Respective images for immunofluorescence staining for NF-H (red) of single neurons (50/μL) and for PECAM/CD-31 (green) of structures formed by HUVECs (2000 cells/μL, ratio 3 MSCs: 1 HUVEC) in a control hydrogel (A) with their corresponding 40x image (B). C: Respective radial plots for the depicted control images' directionality of neurites and vascular-like structures. D,E: Respective images for immunofluorescence staining for NF-H (red) of single neurons (50/μL) and for PECAM/CD-31 (green) of structures formed by HUVECs (2000 cells/μL, ratio 3 MSCs: 1 HUVEC) in a a Anisogel with 1.00 v/v% 2.5x2.5 × 50 μm 3 microgels labeled with Rhodamine-B-acrylate (yellow) (D) and with their corresponding 40x image (E) with 2.00 w/v% PEG and 200 μM IKVAV; F: Respective radial plots for the depicted control images' directionality of neurites, vascular-like structures and microgels. G: 3D Analysis of the total length of vascular-like structures. H: 3D Analysis of the total neurite length from DRGs The PEG-QK gels were processed and analyzed after culturing for 7 days in EGM-2 + NGF. Scale bar: 1 mm (A, D); 200 μm (B, E). n: 3–11 replicates per condition (p∗ < 0.05, p∗∗ < 0.01, p∗∗∗ < 0.001). Error bars are given as standard error. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Article Snippet: Human umbilical vein endothelial cells (HUVECs, passage 1–5, pooled donors, Lonza) were cultured in tissue culture flasks in endothelial growth medium (EGM-2 ready-to-use kit, Promocell) supplemented with FBS (2.00 w/v%), epidermal growth factor (recombinant human, 5 ng/mL), basic fibroblast growth factor (recombinant human, 10.0 ng/mL), insulin-like growth factor (Long R3 IGF, 20 ng/mL), vascular endothelial growth factor 165 (recombinant human, 0.5 ng/mL), ascorbic acid (1 μg/mL), heparin (22.5 μg/mL), and hydrocortisone (0.20 μg/mL).

Techniques: Immunofluorescence, Staining, Control, Labeling