Tendinopathies negatively affect the life quality of millions of people in occupational and athletic settings, as well as the general population. Tendon healing is a slow process, often with insufficient results to restore complete endurance and functionality of the tissue. Tissue engineering, using tendon progenitors, artificial matrices and bioreactors for mechanical stimulation, could be an important approach for treating rips, fraying and tissue rupture. In our work, C3H10T1/2 murine fibroblast cell line was exposed to a combination of stimuli: a biochemical stimulus provided by Transforming Growth Factor Beta (TGF-β) and Ascorbic Acid (AA); a three-dimensional environment represented by PEGylated-Fibrinogen (PEG-Fibrinogen) biomimetic matrix; and a mechanical induction exploiting a custom bioreactor applying uniaxial stretching. In vitro analyses by immunofluorescence and mechanical testing revealed that the proposed combined approach favours the organization of a three-dimensional tissue-like structure promoting a remarkable arrangement of the cells and the neo-extracellular matrix, reflecting into enhanced mechanical strength. The proposed method represents a novel approach for tendon tissue engineering, demonstrating how the combined effect of biochemical and mechanical stimuli ameliorates biological and mechanical properties of the artificial tissue compared to those obtained with single inducement.

The myocardium behaves like a sophisticated orchestra that expresses its true potential only if each member performs the correct task harmonically. Recapitulating its complexity within engineered 3D functional constructs with tailored biological and mechanical properties, is one of the current scientific priorities in the field of regenerative medicine and tissue engineering. In this study, driven by the necessity of fabricating advanced model of cardiac tissue, we present an innovative approach consisting of heterogeneous, multi-cellular constructs composed of Human Umbilical Vein Endothelial Cells (HUVECs) and induced pluripotent cell-derived cardiomyocytes (iPSC-CMs). Cells were encapsulated within hydrogel strands containing alginate and PEG-Fibrinogen (PF) and extruded through a custom microfluidic printing head (MPH) that allows to precisely tailor their 3D spatial deposition, guaranteeing a high printing fidelity and resolution. We obtained a 3D cardiac tissue compose of iPSC-derived CMs with a high orientation index imposed by the different defined geometries and blood vessel-like shapes generated by HUVECs which, as demonstrated by in vivo grafting, better support the integration of the engineered cardiac tissue with host's vasculature.

In human dystrophies, progressive muscle wasting is exacerbated by ectopic deposition of fat and fibrous tissue originating from fibro/adipogenic progenitors (FAPs). In degenerating muscles, the ability of these cells to promote successful healing is attenuated, and FAPs aberrantly expand and differentiate into adipocytes and fibroblasts. Thus, arresting the fibro/adipogenic fate of FAPs, without affecting their physiological role, represents a valuable therapeutic strategy for patients affected by muscle diseases. Here, using a panel of adipose progenitor cells, including human-derived FAPs, coupled with pharmacological perturbations and proteome profiling, we report that LY2090314 interferes with a genuine adipogenic program acting as WNT surrogate for the stabilization of a competent β-catenin transcriptional complex. To predict the beneficial impact of LY2090314 in limiting ectopic deposition of fat in human muscles, we combined a poly-ethylene-glycol-fibrinogen biomimetic matrix with these progenitor cells to create a miniaturized 3D model of adipogenesis. Using this scalable system, we demonstrated that a two-digit nanomolar dose of this compound effectively represses adipogenesis at higher 3D scale, thus indicating the potential for LY2090314 to limit FAP-derived fat infiltrates in dystrophic muscles.

We present a new strategy for the fabrication of artificial skeletal muscle tissue with functional morphologies based on an innovative 3D bioprinting approach. The methodology is based on a microfluidic printing head coupled to a co-axial needle extruder for high-resolution 3D bioprinting of hydrogel fibers laden with muscle precursor cells (C2C12). To promote myogenic differentiation, we formulated a tailored bioink with a photocurable semi-synthetic biopolymer (PEG-Fibrinogen) encapsulating cells into 3D constructs composed of aligned hydrogel fibers. After 3-5 days of culture, the encapsulated myoblasts started migrating and fusing, forming multinucleated myotubes within the 3D bioprinted fibers. The obtained myotubes showed high degree of alignment along the direction of hydrogel fiber deposition, further revealing maturation, sarcomerogenesis, and functionality. Following subcutaneous implantation in the back of immunocompromised mice, bioprinted constructs generated organized artificial muscle tissue in vivo. Finally, we demonstrate that our microfluidic printing head allows to design three dimensional multi-cellular assemblies with an exquisite compartmentalization of the encapsulated cells. Our results demonstrate an enhanced myogenic differentiation with the formation of parallel aligned long-range myotubes. The approach that we report here represents a robust and valid candidate for the fabrication of macroscopic artificial muscle to scale up skeletal muscle tissue engineering for human clinical application.

Invasive intraneural electrodes can control advanced neural-interfaced prostheses in human amputees. Nevertheless, in chronic implants, the progressive formation of a fibrotic capsule can gradually isolate the electrode surface from the surrounding tissue leading to loss of functionality. This is due to a nonspecific inflammatory response called foreign-body reaction (FBR). The commonly used poly(ethylene glycol) (PEG)-based low-fouling coatings of implantable devices can be easily encapsulated and are susceptible to oxidative damage in long-term in vivo applications. Recently, sulfobetaine-based zwitterionic hydrogels have emerged as an important class of robust ultra-low fouling biomaterials, holding great potential to mitigate FBR. The aim of this proof-of-principle in vitro work was to assess whether the organic zwitterionic-poly(sulfobetaine methacrylate) [poly(SBMA)]-hydrogel could be a suitable coating for Polyimide (PI)-based intraneural electrodes to reduce FBR. We first synthesized and analyzed the hydrogel through a mechanical characterization (i.e., Young's modulus). Then, we demonstrated reduced adhesion and activation of fibrogenic and pro-inflammatory cells (i.e., human myofibroblasts and macrophages) on the hydrogel compared with PEG-coated and polystyrene surfaces using cell viability assays, confocal fluorescence microscopy and high-content analysis of oxidative stress production. Interestingly, we successfully coated PI surfaces with a thin film of the hydrogel through covalent bond and demonstrated its high hydrophilicity via water contact angle measurement. Importantly, we showed the long-term release of an anti-fibrotic drug (i.e., Everolimus) from the hydrogel. Because of the low stiffness, biocompatibility, high hydration and ultra-low fouling characteristics, our zwitterionic hydrogel could be envisioned as long-term diffusion-based delivery system for slow and controlled anti-inflammatory and anti-fibrotic drug release in vivo.