Three-dimensional printing is revolutionizing the development of scaffolds due to their rapid-prototyping characteristics. One of the most used techniques is fused filament fabrication (FFF), which is fast and compatible with a wide range of polymers, such as PolyLactic Acid (PLA). Mechanical properties of the 3D printed polymeric scaffolds are often weak for certain applications. A potential solution is the development of composite materials. In the present work, metal-PLA composites have been tested as a material for 3D printing scaffolds. Three different materials were tested: copper-filled PLA, bronze-filled PLA, and steel-filled PLA. Disk-shaped samples were printed with linear infill patterns and line spacing of 0.6, 0.7, and 0.8 mm, respectively. The porosity of the samples was measured from cross-sectional images. Biocompatibility was assessed by culturing Human Bone Marrow-Derived Mesenchymal Stromal on the surface of the printed scaffolds. The results showed that, for identical line spacing value, the highest porosity corresponded to bronze-filled material and the lowest one to steel-filled material. Steel-filled PLA polymers showed good cytocompatibility without the need to coat the material with biomolecules. Moreover, human bone marrow-derived mesenchymal stromal cells differentiated towards osteoblasts when cultured on top of the developed scaffolds. Therefore, it can be concluded that steel-filled PLA bioprinted parts are valid scaffolds for bone tissue engineering.

The extracellular matrix (ECM) of the lung is a filamentous network composed mainly of collagens, elastin, and proteoglycans that provides structural and physical support to its populating cells. Proliferation, migration and overall behaviour of those cells is greatly determined by micromechanical queues provided by the ECM. Lung fibrosis displays an aberrant increased deposition of ECM which likely changes filament organization and stiffens the ECM, thus upregulating the profibrotic profile of pulmonary cells. We have previously used AFM to assess changes in the Young’s Modulus (E) of the ECM in the lung. Here, we perform further ECM topographical, mechanical and viscoelastic analysis at the micro- and nano-scale throughout fibrosis development. Furthermore, we provide nanoscale correlations between topographical and elastic properties of the ECM fibres. Firstly, we identify a softening of the ECM after rats are instilled with media associated with recovery of mechanical homeostasis, which is hindered in bleomycin-instilled lungs. Moreover, we find opposite correlations between fibre stiffness and roughness in PBS- vs bleomycin-treated lung. Our findings suggest that changes in ECM nanoscale organization take place at different stages of fibrosis, with the potential to help identify pharmacological targets to hinder its progression.

Mechanical changes in tumors have long been linked to increased malignancy and therapy resistance and attributed to mechanical changes in the tumor extracellular matrix (ECM). However, to the best of our knowledge, there have been no mechanical studies on decellularized tumors. Here, we studied the biochemical and mechanical progression of the tumor ECM in two models of lung metastases: lung carcinoma (CAR) and melanoma (MEL). We decellularized the metastatic lung sections, measured the micromechanics of the tumor ECM, and stained the sections for ECM proteins, proliferation, and cell death markers. The same methodology was applied to MEL mice treated with the clinically approved anti-fibrotic drug nintedanib. When compared to healthy ECM (~0.40 kPa), CAR and MEL lung macrometastases produced a highly dense and stiff ECM (1.79 ± 1.32 kPa, CAR and 6.39 ± 3.37 kPa, MEL). Fibronectin was overexpressed from the early stages (~118%) to developed macrometastases (~260%) in both models. Surprisingly, nintedanib caused a 4-fold increase in ECM-occupied tumor area (5.1 ± 1.6% to 18.6 ± 8.9%) and a 2-fold in-crease in ECM stiffness (6.39 ± 3.37 kPa to 12.35 ± 5.74 kPa). This increase in stiffness strongly correlated with an increase in necrosis, which reveals a potential link between tumor hypoxia and ECM deposition and stiffness. Our findings highlight fibronectin and tumor ECM mechanics as attractive targets in cancer therapy and support the need to identify new anti-fibrotic drugs to abrogate aberrant ECM mechanics in metastases.

Traditional cell culture is experiencing a revolution moving toward physiomimetic approaches aiming to reproduce healthy and pathological cell environments as realistically as possible. There is increasing evidence demonstrating that biophysical and biochemical factors determine cell behavior, in some cases considerably. Alongside the explosion of these novel experimental approaches, different bioengineering techniques have been developed and improved. Increased affordability and popularization of 3D bioprinting, fabrication of custom-made lab-on-a chip, development of organoids and the availability of versatile hydrogels are factors facilitating the design of tissue-specific physiomimetic in vitro models. However, lower oxygen diffusion in 3D culture is still a critical limitation in most of these studies, requiring further efforts in the field of physiology and tissue engineering and regenerative medicine. During recent years, novel advanced 3D devices are introducing integrated biosensors capable of monitoring oxygen consumption, pH and cell metabolism. These biosensors seem to be a promising solution to better control the oxygen delivery to cells and to reproduce some disease conditions involving hypoxia. This review discusses the current advances on oxygen biosensors and control in 3D physiomimetic experimental models.

The use of physiomimetic decellularized extracellular matrix-derived hydrogels is attracting interest since they can modulate the therapeutic capacity of numerous cell types, including mesenchymal stromal cells (MSCs). Remarkably, extracellular vesicles (EVs) derived from MSCs display similar functions as their parental cells, mitigating tissue damage in lung diseases. However, recent data have shown that ECM-derived hydrogels could release other resident vesicles similar to EVs. Here, we aim to better understand the contribution of EVs and ECM-vesicles released from MSCs and/or lung-derived hydrogel (L-HG) in lung repair by using an in vitro lung injury model. L-HG derived-vesicles and MSCs EVs cultured either in L-HG or conventional plates were isolated and characterized. The therapeutic capacity of vesicles obtained from each experimental condition was tested by using an alveolar epithelial wound-healing assay. The number of ECM-vesicles released from acellular L-HG was 10-fold greater than EVs from conventional MSCs cell culture revealing that L-HG is an important source of bioactive vesicles. MSCs-derived EVs and L-HG vesicles have similar therapeutic capacity in lung repair. However, when wound closure rate was normalized by total proteins, the MSCs-derived EVs shows higher therapeutic potential to those released by L-HG. The EVs released from L-HG must be considered when HG is used as substrate for cell culture and EVs isolation.

Since the emergence of 3D bioprinting technology, both synthetic and natural materials have been used to develop bioinks for producing cell-laden cardiac grafts. To this end, extracellular-matrix (ECM)-derived hydrogels can be used to develop scaffolds that closely mimic the complex 3D environments for cell culture. This study presents a novel cardiac bioink based on hydrogels exclusively derived from decellularized porcine myocardium loaded with human-bone-marrow-derived mesenchymal stromal cells. Hence, the hydrogel can be used to develop cell-laden cardiac patches without the need to add other biomaterials or use additional crosslinkers. The scaffold ultrastructure and mechanical properties of the bioink were characterized to optimize its production, specifically focusing on the matrix enzymatic digestion time. The cells were cultured in 3D within the developed hydrogels to assess their response. The results indicate that the hydrogels fostered inter-cell and cell-matrix crosstalk after 1 week of culture. In conclusion, the bioink developed and presented in this study holds great potential for developing cell-laden customized patches for cardiac repair.