Recombinant Human Collagen-Based Bioinks for the 3D Bioprinting of Full-thickness Human Skin Equivalent

Abstract

As a major extracellular matrix component within the skin, collagen has been widely used to engineer human skin tissues. However, most collagen is extracted from animals. Here, we introduced recombinant human type III collagen (rhCol3) as a bioactive component to formulate bioinks for the bioprinting of a full-thickness human skin equivalent. Human dermal fibroblasts were encapsulated in the gelatin methacryloyl-rhCol3 composite bioinks and printed on a transwell to form the dermis layer, on which human epidermal keratinocytes were seeded to perform an air-liquid interface culture for 6 weeks. After optimizing the bioink formulation and bioprinting process, we investigated the effect of rhCol3 on skin tissue formation. The results suggest that a higher concentration of rhCol3 would enhance the growth of both cells, resulting in a more confluent (~100%) spreading of the epidermal keratinocytes at an early stage (3 days), compared to the rhCol3-free counterpart. Moreover, in an in vivo experiment, adding rhCol3 in the hydrogel formulation would contribute to the skin wound healing process. Taken together, we conclude that rhCol3 could act as a functional bioink component to promote basic skin cellular processes for skin tissue engineering.

Engineered-Skin of Single Dermal Layer Containing Printed Hybrid Gelatin-Polyvinyl Alcohol Bioink via 3D-Bioprinting: In Vitro Assessment under Submerged vs. Air-Lifting Models

“…Nevertheless, other studies comparing submerged and air-lifting models against other types of hydrogels reported a similar cell viability trend as our findings. However, a previous study by Vila et al, 2018, showed a similar cell viability trend of encapsulated fibroblasts of >85% for air-lifting model [36], and a study by Yang et al, 2022 unraveled that the encapsulated HDFs for submerged hydrogels could maintain cell viability up to 90% [37]. Although the HDFs showed high cell in the GPVA hydrogels, the cells maintained their cell morphology in spherical and rounded shapes after a few days of incubation.…”

Marine Collagen-Based Bioink for 3D Bioprinting of a Bilayered Skin Model

“…The mechanical characterisation was performed after the crosslinking of biofabricated samples using each one of bioink formulations proposed. The marine collagen concentration increases the construct stiffness resulting in more robust and easily handled samples; however, all the formulations showed a compression modulus in the range of characteristics of a soft hydrogel for skin tissue engineering or soft tissue bioprinting described in the literature [ 30 , 42 ]. Yang et al, in their study on the development of a recombinant human collagen-based bioinks for the 3D bioprinting of a skin equivalent, reported a compression modulus of 22.6 kPa that is similar to the compression modulus of both collagen-based bioinks proposed [ 42 ].…”

Application of Collagen-Based Hydrogel in Skin Wound Healing

“…This approach usually involves the reaction between the photosensitizer and UV light to produce intra- and intermolecular links within the collagen fibers. The UV–riboflavin or UV–GelMA-induced crosslinking reaction of collagen is commonly used in the treatment of skin tissue [ 49 , 50 , 51 ].…”

“…Gelatin methacryloyl (GelMA) is usually mixed with collagen to create bio-inks for photoinitiated 3D bioprinting because of its great biocompatibility and quick photocrosslinking capabilities. Yang et al blended recombinant human type III collagen (rhCol3) with GelMA to configure hybrid bio-inks [ 50 ] ( Figure 6 B). Although adding rhCol3 to GelMA slightly slows down the kinetics of thermal and photocrosslinking, as well as the mechanical properties after gelation, they successfully constructed an in vitro 3D human skin equivalent with human epidermal keratinocytes (HaCaTs) and dermal fibroblasts (HDFs) using extrusion-based 3D bioprinting.…”

“…2021, American Chemical Society) ( B ) Preparation of the rhCol3-based bio-ink formulations consisting of the base component GelMA, bioactive rhCol3, and photoinitiator LAP; 3D bioprinting of the HDF-laden dermal constructs on a transwell, followed by photocrosslinking; seeding of HaCaTs on top of printed dermal constructs; submerged culturing of the in vitro 3D skin tissue; air–liquid interface culturing process to obtain the differentiated epidermal layer (Adapted with permission from Ref. [ 50 ] 2022 Yang et al). ( C )The bio-ink formulation contained alginate–tyramine polymer chains and collagen, which can be crosslinked to form a mechanically stable, high-resolution structure with homogeneously distributed cells (Adapted with permission from Ref [ 98 ].…”