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Summary: Background: While several commercial dermoepidermal scaffolds can promote wound healing of the skin, the achievement of complete skin regeneration still represents a major challenge. Objectives: To perform biological characterization of self‐assembled extracellular matrices (ECMs) from three different subpopulations of fibroblasts found in human skin: papillary fibroblasts (Pfi), reticular fibroblasts (Rfi) and dermal papilla fibroblasts (DPfi). Methods: Fibroblast subpopulations were cultured with ascorbic acid to promote cell‐assembled matrix production for 10 days. Subsequently, cells were removed and the remaining matrices characterized. Additionally, in another experiment, keratinocytes were seeded on the top of cell‐depleted ECMs to generate epidermal‐only skin constructs. Results: We found that the ECM self‐assembled by Pfi exhibited randomly oriented fibres associated with the highest interfibrillar space, reflecting ECM characteristics that are physiologically present within the papillary dermis. Mass spectrometry followed by validation with immunofluorescence analysis showed that thrombospondin 1 is preferentially expressed within the DPfi‐derived matrix. Moreover, we observed that epidermal constructs grown on DPfi or Pfi matrices exhibited normal basement membrane formation, whereas Rfi matrices were unable to support membrane formation. Conclusions: We argue that inspiration can be taken from these different ECMs, to improve the design of therapeutic biomaterials in skin engineering applications. [ABSTRACT FROM AUTHOR]

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Summary: Chronic wounds affect 1–2% of the world's population at any given time. These can be as a result of burns, or ulceration, and are essentially wounds which do not close. To facilitate closure, there are a number of biological products available which can be used as temporary skin replacements, or to promote tissue repair. These products usually replicate the two main layers found in human skin: the epidermis and dermis. Within the skin dermis the most abundant cell type are fibroblasts, whose primary role is to secrete extracellular matrix and support growth of cells in the adjacent epidermal layer. As fibroblasts within the skin are highly varied, the extracellular matrix in distinct locations of the dermis is also different; however skin substitutes do not usually reflect this diversity. In this study, from the UK, the researchers isolated three fibroblast sub‐types from human scalp skin dermis, and set about to characterise the extracellular matrix which the different sub‐types of fibroblasts synthesised in culture (i.e. developed in the lab, rather than on living skin). They found that the different fibroblast sub‐types produced extracellular matrix in culture reflective of the extracellular matrix found in distinct dermal locations in vivo (in living skin). They also found that certain fibroblast sub‐types were more proficient at supporting adjacent epithelial cells than others, which reflected the sub‐anatomical location from which the fibroblast sub‐types were originally isolated. The authors concluded that inspiration should be taken from the extracellular matrix which fibroblasts secrete to improve the design of biomimetic skin substitutes with improved therapeutic potential for skin tissue engineering. [ABSTRACT FROM AUTHOR]

Modification of tissue engineering scaffolds with bioactive molecules is a potential strategy for modulating cell behavior and guiding tissue regeneration. While adhesion to RGD peptides has been shown to inhibit in vitro chondrogenesis, the effects of extracellular matrix (ECM)-mimetic ligands with complex secondary and tertiary structures are unknown. This study aimed to determine whether collagen- and fibronectin-mimetic ligands would retain biologic functionality in three-dimensional (3D) hydrogels, whether different ECM-mimetic ligands differentially influence in vitro chondrogenesis, and if effects of ligands on differentiation depend on soluble biochemical stimuli. A linear RGD peptide, a recombinant fibronectin fragment containing the seven to ten Type III repeats (FnIII7-10) and a triple helical, collagen mimetic peptide with the GFOGER motif were covalently coupled to agarose gels using the sulfo-SANPAH crosslinker, and bone marrow stromal cells (BMSCs) were cultured within the 3D hydrogels. The ligands retained biologic functionality within the agarose gels and promoted density-dependent BMSC spreading. Interactions with all adhesive ligands inhibited stimulation by chondrogenic factors of collagen Type II and aggrecan mRNA levels and deposition of sulfated glycosaminoglycans. In medium containing fetal bovine serum, interactions with the GFOGER peptide enhanced mRNA expression of the osteogenic gene osteocalcin whereas FnIII7-10 inhibited osteocalcin expression. In conclusion, modification of agarose hydrogels with ECM-mimetic ligands can influence the differentiation of BMSCs in a manner that depends strongly on the presence and nature of soluble biochemical stimuli.

Objective: The goal of this study was to characterize the proteoglycan (PG) production and processing by bone marrow stromal cells (BMSCs) within a tissue engineered construct., Methods: Bovine BMSCs and articular chondrocytes (ACs) were isolated from an immature calf, seeded into agarose gels, and cultured up to 32 days in the presence of TGF-beta1. The localization of various PGs was examined by immunofluorescence and histological staining. The role of proteolytic enzymes in construct development was further investigated by examining the effects of aggrecanase and MMP inhibitors on PG accumulation, aggrecan processing, and construct mechanics., Results: BMSCs developed a matrix rich in sulfated-glycosaminoglycans (sGAG) and full-length aggrecan, but had low levels of versican. The BMSC constructs had less collagen II and aggrecan compared to the AC constructs cultured under identical conditions. AC constructs also had high levels of pericellular collagen VI, while BMSCs had a pericellular matrix containing little collagen VI and greater levels of decorin, biglycan, and fibronectin. Treatment with the aggrecanase inhibitor did not affect sGAG accumulation or the dynamic moduli of the BMSC constructs. The MMP inhibitor slightly but significantly inhibited sGAG accumulation and lowered the dynamic moduli of BMSC constructs., Conclusions: The results of this preliminary study indicate that long-term culture of BMSCs with TGF-beta1 promotes the development of an aggrecan-rich matrix characteristic of native articular cartilage; however, BMSCs accumulate significantly lower levels of sGAG and assemble distinct pericellular microenvironments compared to ACs. PG turnover does not appear to play a major role in the development of tissue engineered cartilage constructs by BMSCs.