Microfabrication of a biomimetic arcade-like electrospun scaffold for cartilage tissue engineering applications.

In recent years, the engineering of biomimetic cellular microenvironments has emerged as a top priority for regenerative medicine, being the in vitro recreation of the arcade-like cartilaginous tissue one of the most critical challenges due to the notorious absence of cost- and time-efficient microfabrication techniques capable of building 3D fibrous scaffolds with precise anisotropic properties. Taking this into account, we suggest a feasible and accurate methodology that uses a sequential adaptation of an electrospinning-electrospraying set up to construct a hierarchical system comprising both polycaprolactone (PCL) fibres and polyethylene glycol sacrificial microparticles. After porogen leaching, the bi-layered PCL scaffold was capable of presenting not only a depth-dependent fibre orientation similar to natural cartilage, but also mechanical features and porosity proficient to encourage an enhanced cell response. In fact, cell viability studies confirmed the biocompatibility of the scaffold and its ability to guarantee suitable cell adhesion, proliferation and migration throughout the 3D anisotropic fibrous network during 21 days of culture. Additionally, likewise the hierarchical relationship between chondrocytes and their extracellular matrix, the reported PCL scaffold was able to induce depth-dependent cell-material interactions responsible for promoting a spatial modulation of the morphology, alignment and density of the cells in vitro.

Three-dimensional nanofibrous and porous scaffolds of poly(ε-caprolactone)-chitosan blends for musculoskeletal tissue engineering.

One of the established tissue engineering strategies relies on the fabrication of appropriate materials architectures (scaffolds) that mimic the extracellular matrix (ECM) and assist the regeneration of living tissues. Fibrous structures produced by electrospinning have been widely used as reliable ECM templates but their two-dimensional structure restricts, in part, cell infiltration and proliferation. A recent technique called thermally-induced self-agglomeration (TISA) allowed to alleviate this drawback by rearranging the 2D electrospun membranes into highly functional 3D porous-fibrous systems. Following this trend, the present research focused on preparing polycaprolactone/chitosan blends by electrospinning, to then convert them into 3D structures by TISA. By adding different amounts of chitosan, it was possible to accurately modulate the physicochemical properties of the obtained 3D nanofibrous scaffolds, leading to highly porous constructs with distinct morphologic and mechanical features. Viability and proliferation studies using adult human chondrocytes also revealed that the biocompatibility of the scaffolds was not impaired after 28 days of cell culture, highlighting their potential to be included into musculoskeletal tissue engineering applications, particularly cartilage repair.

Multi-layered electrospinning and electrospraying approach: Effect of polymeric supplements on chondrocyte suspension.

Articular cartilage was expected to be one of the first tissues to be successfully engineered, but replicating the complex fibril architecture and the cellular distribution of the native cartilage has proven difficult. While electrospinning has been widely used to reproduce the depth-dependent fibre architecture in 3D scaffolds, the chondrocyte-controlled distribution remains an unsolved problem. To incorporate cells homogeneously through the depth of scaffolds, a combination of polymer electrospinning and cell seeding is necessary. A multi-layer approach alternating between polymer electrospinning with chondrocyte electrospraying can be a solution. Still, the success of this process is related to the survival rate of the electrosprayed chondrocytes embedded within the electrospun mesh. In this regard, the present study investigated the impact of the multi-layered process and the supplementation of the electrospray chondrocyte suspension with different concentrations of Gelatin and Alginate on the viability of electrosprayed chondrocytes embedded within a Polycaprolactone/Gelatin electrospun mesh and on the mechanical properties of the resulting meshes. The addition of Gelatin in the chondrocyte suspension did not increase significantly (p > 0.05) the percentage of viable electrosprayed chondrocytes (25%), while 3 wt% Alginate addition led to a significant (p < 0.05) increase in chondrocyte viability (50%) relative to the case without polymer supplement (15%). Furthermore, the addition of both polymer supplements increased the mechanical properties of the multi-layer construct. These findings imply that this multi-layered approach can be applied to cartilage TE allowing for automated chondrocyte integration during scaffolds creation.

Boosting in vitro cartilage tissue engineering through the fabrication of polycaprolactone-gelatin 3D scaffolds with specific depth-dependent fiber alignments and mechanical stimulation.

Due to the limited self-healing ability of natural cartilage, several tissue engineering strategies have been explored to develop functional replacements. Still, most of these approaches do not attempt to recreate in vitro the anisotropic organization of its extracellular matrix, which is essential for a suitable load-bearing function. In this work, different depth-dependent alignments of polycaprolactone-gelatin electrospun fibers were assembled into three-dimensional scaffold architectures to assess variations on chondrocyte response under static, unconfined compressed and perfused culture conditions. The in vitro results confirmed that not only the 3D scaffolds specific depth-dependent fiber alignments potentiated chondrocyte proliferation and migration towards the fibrous systems, but also the mechanical stimulation protocols applied were able to enhance significantly cell metabolic activity and extracellular matrix deposition, respectively.

Electrospinning of bioactive polycaprolactone-gelatin nanofibres with increased pore size for cartilage tissue engineering applications.

Polycaprolactone (PCL) electrospun scaffolds have been widely investigated for cartilage repair application. However, their hydrophobicity and small pore size has been known to prevent cell attachment, proliferation and migration. Here, PCL was blended with gelatin (GEL) combining the favorable biological properties of GEL with the good mechanical performance of the former. Also, polyethylene glycol (PEG) particles were introduced during the electrospinning of the polymers blend by simultaneous electrospraying. These particles were subsequently removed resulting in fibrous scaffolds with enlarged pore size. PCL, GEL and PEG scaffolds formulations were developed and extensively structural and biologically characterized. GEL incorporation on the PCL scaffolds led to a considerably improved cell attachment and proliferation. A substantial pore size and interconnectivity increase was obtained, allowing cell infiltration through the porogenic scaffolds. All together these results suggest that this combined approach may provide a potentially clinically viable strategy for cartilage regeneration.