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Hybrid constructs represent substantial progress in tissue engineering (TE) towards producing implants of a clinically relevant size that recapitulate the structure and multicellular complexity of the native tissue. They are created by interlacing printed scaffolds, sacrificial materials, and cell-laden hydrogels. A suitable biomaterial is a polycaprolactone (PCL); however, due to the higher viscosity of this biopolymer, three-dimensional (3D) printing of PCL is slow, so reducing PCL print times remains a challenge. We investigated parameters, such as nozzle shape and size, carriage speed, and print temperature, to find a tradeoff that speeds up the creation of hybrid constructs of controlled porosity. We performed experiments with conical, cylindrical, and cylindrical shortened nozzles and numerical simulations to infer a more comprehensive understanding of PCL flow rate. We found that conical nozzles are advised as they exhibited the highest shear rate, which increased the flow rate. When working at a low carriage speed, conical nozzles of a small diameter tended to form-flatten filaments and became highly inefficient. However, raising the carriage speed revealed shortcomings because passing specific values created filaments with a heterogeneous diameter. Small nozzles produced scaffolds with thin strands but at long building times. Using large nozzles and a high carriage speed is recommended. Overall, we demonstrated that hybrid constructs with a clinically relevant size could be much more feasible to print when reaching a tradeoff between temperature, nozzle diameter, and speed, This research was funded by FPI-CAR-2014, granted by the Government of La Rioja (Spain) and research funding Nos. 266486, 273689, and 273689 (FINSKIN) by the Academy of Finland (Finland). “Instituto de Estudios Riojanos” (Spain) also funded parts of this research.

Poly(L-lactic acid), PLLA, a synthetic biodegradable polyester, is widely accepted in tissue engineering. Hyaluronic acid (HA), a natural polymer, exhibits an excellent biocompatibility, influences cell signaling, proliferation, and differentiation. In this study, HA crosslinking was performed by immersion of the polysaccharide in water-acetone mixtures containing glutaraldehyde (GA). The objective of this work is to produce PLLA scaffolds with the pores coated with HA, that could be beneficial for bone tissue engineering applications. PLLA tridimensional scaffolds were prepared by compression molding followed by salt leaching. After the scaffolds impregnation with soluble HA solutions of distinct concentration, a GA-crosslinking reaction followed by inactivation of the unreacted GA with glycine was carried out. An increase on surface roughness is shown by scanning electron microscopy (SEM) with the addition of HA. Toluidine blue staining indicates the present of stable crosslinked HA. An estimation of the HA original weight in the hybrid scaffolds was performed using thermal gravimetric analyses. FTIR-ATR and XPS confirmed the crosslinking reaction. Preliminary in vitro cell culture studies were carried out using a mouse lung fibroblast cell line (L929). SEM micrographs of L929 showed that cells adhered well, spread actively throughout all scaffolds, and grew favorably. A MTS test indicated that cells were viable when cultured onto the surface of all scaffolds, suggesting that the introduction of crosslinked HA did not increase the cytotoxicity of the hybrid scaffolds., Contract grant sponsor: Portuguese Foundation for Science and Technology (FCT) through POCTIContract grant sponsor: FEDER programs including project ProteoLight; contract grant number: PTDC/FIS/68517/2006Contract grant sponsor: European Union funded STREP Project HIPPOCRATES; contract grant number: NMP3-CT-2003-505758Contract grant sponsor: European NoE EXPERTISSUES; contract grant number: NMP3-CT-2004-500283Contract grant sponsor: Spanish Ministry of Science (The FEDER financial support); contract grant number: MAT2007-66759-C03-01The authors acknowledge the funding for research in the field of Regenerative Medicine through the collaboration agreement from the Conselleria de Sanidad (Generalitat Valenciana) and the Instituto de Salud Carlos III (Ministry of Science and Innovation). The European Union Financing, as part of the SOCRA-TES/Erasmus program is also gratefully acknowledged.