Hypoxic tumors are radioresistant stemming from the fact that oxygen promotes reactive oxygen species (ROS) propagation after water radiolysis and stabilizes irradiation-induced DNA damage. Therefore, an attractive strategy to radiosensitize solid tumors is to increase tumor oxygenation at the time of irradiation, ideally above a partial pressure of 10 mm-Hg at which full radiosensitization can be reached. Historically, the many attempts to increase vascular O₂ delivery have had limited efficacy, but mathematical models predicted that inhibiting cancer cell respiration would be more effective. Here, we report that mitochondria-targeted antioxidant MitoQ can radiosensitize human breast tumors in mice. This was not a class effect, as neither MitoTEMPO nor SKQ1 shared this property. At clinically relevant nanomolar concentrations, MitoQ completely abrogated the oxygen consumption of several human cancer cell lines of different origins, which was associated with a glycolytic switch. Using orthotopic breast cancer models in mice, we observed that pretreating hypoxic MDA-MB-231 tumors with MitoQ delayed tumor growth with both single dose irradiation and clinically relevant fractionated radiotherapy. Oxygenated MCF7 tumors were not radiosensitized, suggesting an oxygen enhancement effect of MitoQ. Because MitoQ already successfully passed Phase I clinical trials, our findings foster its clinical evaluation in combination with radiotherapy.

Three-dimensional (3D) bioprinting technology enables the controlled deposition of cells and biomaterials (i.e., bioink) to easily create complex 3D biological microenvironments. Silk fibroin (SF) has recently emerged as a compelling bioink component due to its advantageous mechanical and biological properties. This study reports on the development and optimization of a novel bioink for extrusion-based 3D bioprinting and compares different bioink formulations based on mixtures of alginate methacrylate (ALMA), gelatin and SF. The rheological parameters of the bioink were investigated to predict printability and stability, and the optimal concentration of SF was selected. The bioink containing a low amount of SF (0.002% w/V) was found to be the best formulation. Light-assisted gelation of ALMA was exploited to obtain the final hydrogel matrix. Rheological analyses showed that SF-enriched hydrogels exhibited greater elasticity than SF-free hydrogels and were more tolerant to temperature fluctuations. Finally, MG-63 cells were successfully bioprinted and their viability and proliferation over time were analyzed. The SF-enriched bioink represents an excellent biomaterial in terms of printability and allows high cell proliferation over a period of up to 3 weeks. These data confirm the possibility of using the selected formulation for the successful bioprinting of cells into extracellular matrix-like microenvironments.

Gelatin methacryloyl (GelMA) is a polypeptide derived from the methacryloylation of gelatin and retains the thermoresponsive behavior of gelatin. When cooled, GelMA undergoes a sol-gel transition. By photo-cross-linking GelMA in a heated ("Hot") or cooled ("Cold") state, it results in a set of hydrogels with distinct properties. To date, the mechanical properties of these resulting hydrogels have not been fully elucidated. Here, we show that "Cold" hydrogels are viscoelastic, while "Hot" hydrogels are almost completely elastic. These features have remarkable implications for cell-substrate interactions in vitro: here, we show that fibroblasts, when cultured on these different substrates, adhere preferentially to Cold hydrogels. These results suggest that efficient cell adhesion requires specific mechanical properties of the substrate. This novel platform enables the precise control of different mechanical properties of GelMA by simply adjusting the cross-linking temperature, providing a flexible approach for the design of biologically inspired microenvironments in vitro.

The crosstalk between the cells and the extracellular matrix (ECM) is bidirectional and consists of a pushing/pulling stretch exerted by the cells and a mechanical resistance counteracted by the surrounding microenvironment. It is widely recognized that the stiffness of the ECM, its viscoelasticity, and its overall deformation are the most important traits influencing the response of the cells. Here these three parameters are combined into a concept of elastic energy, which in biological terms represents the mechanical feedback that cells perceive when the ECM is deformed. It is shown that elastic energy is a stress factor that influences the response of cells in three‐dimensional (3D) cultures. Strikingly, the higher the elastic energy of the matrix and thus the mechanical feedback, the higher the stress state of the cells, which correlates with the formation of G3BP‐mediated stress granules. This condition is associated with an increase in alkaline phosphatase (ALP) activity but a decrease in gene expression and is mediated by the nuclear translocation of Yes‐associated protein (YAP). This work supports the importance of considering the elastic energy as mechano‐controller in regulating cellular stress state in 3D cultures.

Dental caries represents one of the most prevalent oral diseases worldwide, and conventional treatments rely on the use of inert restorative materials. Yet, restoration failure rates remain frequent, as current in vitro testing platforms fail to reproduce the complexity of native dental tissues. Here, we present a bioprinted dual-mechanical dentin-pulp platform generated using a methacrylated alginate (ALMA)-based biomaterial ink that can be either single or dual crosslinked (i.e. SC and DC models). These platforms, which differ in terms of stiffness and viscoelasticity, were developed to probe how biophysical cues govern cell-specific functions. When bioprinted within these matrices, HDPSCs-derived odontoblasts showed upregulation of lineage-specific markers and tissue mineralization within the DC models. In contrast, Human Umbilical Vein Endothelial Cells developed a more complex and interconnected vessel-like network within the SC constructs. By integrating both compartments within a single platform, we propose an in vitro dentin-pulp model that mimics the mechanical heterogeneity of native dental tissues.