Thiolene- and Polycaprolactone Methacrylate-Based Polymerized High Internal Phase Emulsion (PolyHIPE) Scaffolds for Tissue Engineering.

Highly porous emulsion templated polymers (PolyHIPEs) provide a number of potential advantages in the fabrication of scaffolds for tissue engineering and regenerative medicine. Porosity enables cell ingrowth and nutrient diffusion within, as well as waste removal from, the scaffold. The properties offered by emulsion templating alone include the provision of high interconnected porosity, and, in combination with additive manufacturing, the opportunity to introduce controlled multiscale porosity to complex or custom structures. However, the majority of monomer systems reported for PolyHIPE preparation are unsuitable for clinical applications as they are nondegradable. Thiol-ene chemistry is a promising route to produce biodegradable photocurable PolyHIPEs for the fabrication of scaffolds using conventional or additive manufacturing methods; however, relatively little research has been reported on this approach. This study reports the groundwork to fabricate thiol- and polycaprolactone (PCL)-based PolyHIPE materials via a photoinitiated thiolene click reaction. Two different formulations, either three-arm PCL methacrylate (3PCLMA) or four-arm PCL methacrylate (4PCLMA) moieties, were used in the PolyHIPE formulation. Biocompatibility of the PolyHIPEs was investigated using human dermal fibroblasts (HDFs) and human osteosarcoma cell line (MG-63) by DNA quantification assay, and developed PolyHIPEs were shown to be capable of supporting cell attachment and viability.

Fabrication of Biodegradable Synthetic Vascular Networks and Their Use as a Model of Angiogenesis.

One of the greatest challenges currently faced in tissue engineering is the incorporation of vascular networks within tissue-engineered constructs. The aim of this study was to develop a technique for producing a perfusable, 3-dimensional, cell-friendly model of vascular structures that could be used to study the factors affecting angiogenesis and vascular biology in engineered systems in more detail. Initially, biodegradable synthetic pseudovascular networks were produced via the combination of robocasting and electrospinning techniques. The internal surfaces of the vascular channels were then recellularized with human dermal microvascular endothelial cells (HDMECs) with and without the presence of human dermal fibroblasts (HDFs) on the outer surface of the scaffold. After 7 days in culture, channels that had been reseeded with HDMECs alone demonstrated irregular cell coverage. However, when using a co-culture of HDMECs inside and HDFs outside the vascular channels, coverage was found to be continuous throughout the internal channel. Using this cell combination, collagen gels loaded with vascular endothelial growth factor were deposited onto the outer surface of the scaffold and cultured for a further 7 days. After this, endothelial cell outgrowth from within the channels into the collagen gel was observed, showing that the engineered vasculature maintains its capacity for angiogenesis. Furthermore, the HDMECs appeared to have formed perfusable tubules within the gel. These results show promising steps towards the development of an in vitro platform for studying angiogenesis and vascular biology in a tissue engineering context.

Investigating Neovascularization in Rat Decellularized Intestine: An In Vitro Platform for Studying Angiogenesis.

One of the main challenges currently faced by tissue engineers is the loss of tissues postimplantation due to delayed neovascularization. Several strategies are under investigation to create vascularized tissue, but none have yet overcome this problem. In this study, we produced a decellularized natural vascular scaffold from rat intestine to use as an in vitro platform for neovascularization studies for tissue-engineered constructs. Decellularization resulted in almost complete (97%) removal of nuclei and DNA, while collagen, glycosaminoglycan, and laminin content were preserved. Decellularization did, however, result in the loss of elastin and fibronectin. Some proangiogenic factors were retained, as fragments of decellularized intestine were able to stimulate angiogenesis in the chick chorioallantoic membrane assay. We demonstrated that decellularization left perfusable vascular channels intact, and these could be repopulated with human dermal microvascular endothelial cells. Optimization of reendothelialization of the vascular channels showed that this was improved by continuous perfusion of the vasculature and further improved by infusion of human dermal fibroblasts into the intestinal lumen, from where they invaded into the decellularized tissue. Finally we explored the ability of the perfused cells to form new vessels. In the absence of exogenous angiogenic stimuli, Dll4, a marker of endothelial capillary-tip cell activation during sprouting angiogenesis, was absent, indicating that the reformed vasculature was largely quiescent. However, after addition of vascular endothelial growth factor A, Dll4-positive endothelial cells could be detected, demonstrating that this engineered vascular construct maintained its capacity for neovascularization. In summary, we have demonstrated how a natural xenobiotic vasculature can be used as an in vitro model platform to study neovascularization and provide information on factors that are critical for efficient reendothelialization of decellularized tissue.

Vascularization strategies for tissue engineers.

All tissue-engineered substitutes (with the exception of cornea and cartilage) require a vascular network to provide the nutrient and oxygen supply needed for their survival in vivo. Unfortunately the process of vascular ingrowth into an engineered tissue can take weeks to occur naturally and during this time the tissues become starved of essential nutrients, leading to tissue death. This review initially gives a brief overview of the processes and factors involved in the formation of new vasculature. It then summarizes the different approaches that are being applied or developed to overcome the issue of slow neovascularization in a range of tissue-engineered substitutes. Some potential future strategies are then discussed.

Fabrication of biodegradable synthetic perfusable vascular networks via a combination of electrospinning and robocasting.

Biodegradable synthetic vascular networks are produced via the combination of robocasting and electrospinning techniques. Preliminary revascularization studies using microvascular endothelial cells and human dermal fibroblasts show good attachment and uniform distribution within the vascular networks, highlighting their potential use in vascular tissue engineering applications.