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Porous collagen-glycosaminoglycan (collagen-GAG) scaffolds have shown promising clinical results for wound healing; however, these scaffolds do not replace the dermal and epidermal layer simultaneously and rely on local endogenous signaling to direct healing. Functionalizing collagen-GAG scaffolds with signaling factors, and/or additional matrix molecules, could help overcome these challenges. An ideal candidate for this is platelet-rich plasma (PRP) as it is a natural reservoir of growth factors, can be activated to form a fibrin gel, and is available intraoperatively. We tested the factors released from PRP (PRPr) and found that at specific concentrations, PRPr enhanced cell proliferation and migration and induced angiogenesis to a greater extent than fetal bovine serum (FBS) controls. This motivated us to develop a strategy to successfully incorporate PRP homogeneously within the pores of the collagen-GAG scaffolds. The composite scaffold released key growth factors for wound healing (FGF, TGFβ) and vascularization (VEGF, PDGF) for up to 14 days. In addition, the composite scaffold had enhanced mechanical properties (when compared to PRP gel alone), while providing a continuous upper surface of extracellular matrix (ECM) for keratinocyte seeding. The levels of the factors released from the composite scaffold were sufficient to sustain proliferation of key cells involved in wound healing, including human endothelial cells, mesenchymal stromal cells, fibroblasts, and keratinocytes; even in the absence of FBS supplementation. In functional in vitro and in vivo vascularization assays, our composite scaffold demonstrated increased angiogenic and vascularization potential, which is known to lead to enhanced wound healing. Upon pro-inflammatory induction, macrophages released lower levels of the pro-inflammatory marker MIP-1α when treated with PRPr; and released higher levels of the anti-inflammatory marker IL1-ra upon both pro-and anti-inflammatory induction when treated with the composite scaffold. Finally, our composite scaffold supported a co-culture system of human fibroblasts and do Amaral et al. PRP Incorporation Into Collagen-Based Scaffold keratinocytes that resulted in an epidermal-like layer, with keratinocytes constrained to the surface of the scaffold; by contrast, keratinocytes were observed infiltrating the PRPfree scaffold. This novel composite scaffold has the potential for rapid translation to the clinic by isolating PRP from a patient intraoperatively and combining it with regulatory approved scaffolds to enhance wound repair.

Bone tissue engineering scaffolds have two challenging functional tasks to fulfil: to encourage cell proliferation, differentiation and matrix synthesis and to provide suitable mechanical stability upon implantation. Composites of biopolymers and bioceramics combine the advantages of both types of materials, resulting in better processability and enhanced mechanical and biological properties through matrix reinforcement. In the present study, novel thick bone composite scaffolds were successfully fabricated using electrospun flat sheets of polyhydroxybutyrate-polyhydroxyvalerate/nanohydroxyapatite/silk fibroin essence (2% nanohydroxyapatite - 2% silk fibroin essence and 5% nanohydroxyapatite - 5% silk fibroin essence, respectively). Their potential asin vitrobone regeneration scaffolds was evaluated using mouse calvarian osteoblast cells (MC3T3-E1), in terms of morphology (scanning electron microscope), cell attachment, cell proliferation, Col type I, osteopontin and bone alkaline phosphatase activity (Quantitative Real Time Polymerase Chain Reaction [qRT-PCR], enzyme-linked immunosorbent assay, immunocytochemistry). Electrospun polyhydroxybutyrate-polyhydroxyvalerate scaffolds were used as reference constructs. The results showed that the compressive and tensile mechanical properties of the scaffolds are dependent on the change in their composition, and the treatment these underwent. Furthermore, methanol-treated and autoclaved (MA) P2 (2% nanohydroxyapatite, 2% silk fibroin essence) samples appeared to exhibit more promising tensile properties. Additionally, the compressive tests results confirmed that the methanol pre-treatment and the autoclaving step lead to an increase in the P2 secant modulus when compared to the non-methanol-treated ones, P2 and P5 (5% nanohydroxyapatite, 5% silk fibroin essence), respectively.Both formulations of polyhydroxybutyrate-polyhydroxyvalerate/nanohydroxyapatite/silk fibroin essence composite promoted greater cell adhesion and proliferation than the corresponding polyhydroxybutyrate-polyhydroxyvalerate control ones. Cells seeded on the composite fibrous scaffolds were extensively expanded and elongated on the fibre surface after one day in culture, whereas those seeded on the polyhydroxybutyrate-polyhydroxyvalerate scaffolds were not completely elongated. In addition, cells grown on P2 and P5 scaffolds had higher alkaline phosphatase activity when compared to those containing no nanohydroxyapatite/silk fibroin essence.

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The present work examines porous scaffolds preparation methods by combining salt leaching, gas forming, solvent evaporation and phase separation techniques.The scaffolds are intended for bone tissue regeneration. It aims to combine the biodegradability of poly (E-caprolactone) (PCL) with the osteoconductive properties of hydroxylapatite (HA) in order to create a composite which acts as a temporary support for bone cell attachment, growth and differentiation. Samples of different ceramic content were obtained. Scanning electron microscopy (SEM)/EDX), mechanical, hydrophylycity and degradation tests were carried out to characterize the composite. The prepared scaffolds have relatively homogenous pore structure throughout the matrix and the porosity was controlled by altering the initial volume fraction of salt particles and the amount of gas forming agents. The tensile strength of the HAlPCL composites decreased with increasing HA content, porosity and the technique being used. Future studies include bioactivity "in vitro" tests (samples will be immersed in simulated body fluid-SBF-solution for 30days at 37°C) and cell culturing.