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. 2022 Mar 4;23(5):2836.
doi: 10.3390/ijms23052836.

Wound-Microenvironment Engineering through Advanced-Dressing Bioprinting

Cristina Del Amo  1 , Xabier Fernández-San Argimiro  2 , María Cascajo-Castresana  2 , Arantza Perez-Valle  1 , Iratxe Madarieta  2 , Beatriz Olalde  2 , Isabel Andia  1
Affiliations

Wound-Microenvironment Engineering through Advanced-Dressing Bioprinting

Cristina Del Amo et al. Int J Mol Sci. .

Abstract

In patients with comorbidities, a large number of wounds become chronic, representing an overwhelming economic burden for healthcare systems. Engineering the microenvironment is a paramount trend to activate cells and burst-healing mechanisms. The extrusion bioprinting of advanced dressings was performed with novel composite bioinks made by blending adipose decellularized extracellular matrix with plasma and human dermal fibroblasts. Rheological and microstructural assessments of the composite hydrogels supported post-printing cell viability and proliferation over time. Embedded fibroblasts expressed steady concentrations of extracellular matrix proteins, including type 1, 3 and 4 collagens and fibronectin. ELISA assessments, multiplex protein arrays and ensuing bioinformatic analyses revealed paracrine activities corresponding to wound-healing activation through the modulation of inflammation and angiogenesis. The two modalities of advanced dressings, differing in platelet number, showed differences in the release of inflammatory and angiogenic cytokines, including interleukin 8 (IL-8), monocyte chemotactic protein 1 (MCP-1), vascular endothelial growth factor (VEGF) and hepatocyte growth factor (HGF). The conditioned media stimulated human-dermal-cell proliferation over time. Our findings open the door to engineering the microenvironment as a strategy to enhance healing.

Keywords: 3D bioprinting; bioink; cytokines; decellularized adipose extracellular matrix; growth factors; plasma; platelet; wound healing.

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Conflict of interest statement

The authors declare no conflict of interest.

Figures

Figure 1
Figure 1
Rheological behavior: (A) flow curves of pDAM2: PRP/ALG and pDAM2: PPP/ALG inks and the individual components; (B) dynamic moduli at varying frequency at 37 °C of hydrogels (storage and loss moduli measured by frequency sweep tests of pDAM2: PRP/ALG and pDAM2: PPP/ALG inks).
Figure 2
Figure 2
Scanning electron microscopy (SEM) images of: (A) PRP/ALG, (B) pDAM2 and (C) pDAM2: PRP/ALG.
Figure 3
Figure 3
(A) Macroscopic image of cell-laden constructs bioprinted on 24-well plates. (B) HDF proliferation (DNA ng/mg construct) within a period of 11 days of culture. (C) HDF viability at 0, 4, 7 and 11 days of culture after bioprinting with pDAM2:PRP/ALG and pDAM2:PPP/ALG. Green corresponds to calcein-AM staining of life cells and red corresponds to Propidium iodide staining of dead cells (scale bar = 100 µm).
Figure 4
Figure 4
HDFs embedded within both modalities of advanced dressings showed moderate expression of COL1A1, COL1A2 and COL3A1 relative to GAPDH, without variations over time. The expression of fibronectin was high in both dressing types (RNA from the 11-day construct was lost), whereas the expression of COL4A1 and COL4A2 was low. There were no significant changes in gene expression over time and no differences between the two dressing modalities.
Figure 5
Figure 5
Signaling pathways enriched in the data obtained from the analysis of the conditioned media. (A) Signaling molecules synthesized by encapsulated cells over time that were involved in the wound-healing signaling pathway. (B,C) pattern of Z-score outcome over time predicting activation of the wound-healing signaling pathway (B), cell signaling and cell viability (C).
Figure 6
Figure 6
(A–D) Box plots show the median and 25–75 percentiles of relevant signaling cytokines involved in the modulation of inflammation (IL-8 and MCP-1, (A) and (B), respectively) and in angiogenesis (VEGF and HGF, (C) and (D), respectively) measured by ELISA in the conditioned media harvested over time. * Mean HGF concentration at Day 11 was 98,198 ± 3815 ng/mL (not shown in the graph). (E,F) RANTES and PDGF-BB released from the PRP component in the pDAM2: PRP/ALG dressings (as shown by acellular scaffolds cultured in the same conditions) were used up by embedded cells.
Figure 7
Figure 7
(A) Proliferative effect of conditioned media harvested from advanced dressings manufactured with PRP. (B) Proliferative effect of conditioned media harvested from advanced dressings manufactured with PPP.
Figure 8
Figure 8
Illustration of molecules (blue) and mechanisms (pink) involved in the wound-healing cascade and assessed in this study.
See this image and copyright information in PMC

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