Nanotechnology has been successfully used for the fabrication of targeted anti-cancer drug carriers. This study aimed to obtain Fe₃O₄ nanoparticles functionalized with Gemcitabine to improve the cytotoxic effects of the chemotherapeutic substance on cancer cells. The (un) functionalized magnetite nanoparticles were synthesized using a modified co-precipitation method. The nanoconjugate characterization was performed by XRD, SEM, SAED and HRTEM; the functionalizing of magnetite with anti-tumor substances has been highlighted through TGA. The interaction with biologic media has been studied by means of stability and agglomeration tendency (using DLS and Zeta Potential); also, the release kinetics of the drug in culture media was evaluated. Cytotoxicity of free-Gemcitabine and the obtained nanoconjugate were evaluated on human BT 474 breast ductal carcinoma, HepG2 hepatocellular carcinoma and MG 63 osteosarcoma cells by MTS. In parallel, cellular morphology of these cells were examined through fluorescence microscopy and SEM. The localization of the nanoparticles related to the cells was studied using SEM, EDX and TEM. Hemolysis assay showed no damage of erythrocytes. Additionally, an in vivo biodistribution study was made for tracking where Fe₃O₄@Gemcitabine traveled in the body of mice. Our results showed that the transport of the drug improves the cytotoxic effects in comparison with the one produced by free Gemcitabine for the BT474 and HepG2 cells. The in vivo biodistribution test proved nanoparticle accumulation in the vital organs, with the exception of spleen, where black-brown deposits have been found. These results indicate that our Gemcitabine-functionalized nanoparticles are a promising targeted system for applications in cancer therapy.

Biocompatibility, biodegradability, shear tinning behavior, quick gelation and an easy crosslinking process makes alginate one of the most studied polysaccharides in the field of regenerative medicine. The main purpose of this study was to obtain tissue-like materials suitable for use in bone regeneration. In this respect, alginate and several types of clay were investigated as components of 3D-printing, nanocomposite inks. Using the extrusion-based nozzle, the nanocomposites inks were printed to obtain 3D multilayered scaffolds. To observe the behavior induced by each type of clay on alginate-based inks, rheology studies were performed on composite inks. The structure of the nanocomposites samples was examined using Fourier Transform Infrared Spectrometry and X-ray Diffraction (XRD), while the morphology of the 3D-printed scaffolds was evaluated using Electron Microscopy (SEM, TEM) and Micro-Computed Tomography (Micro-CT). The swelling and dissolvability of each composite scaffold in phosfate buffer solution were followed as function of time. Biological studies indicated that the cells grew in the presence of the alginate sample containing unmodified clay, and were able to proliferate and generate calcium deposits in MG-63 cells in the absence of specific signaling molecules. This study provides novel information on potential manufacturing methods for obtaining nanocomposite hydrogels suitable for 3D printing processes, as well as valuable information on the clay type selection for enabling accurate 3D-printed constructs. Moreover, this study constitutes the first comprehensive report related to the screening of several natural clays for the additive manufacturing of 3D constructs designed for bone reconstruction therapy.

This study aims to investigate whether ionizing radiation combined with doxorubicin-conjugated iron oxide nanoparticles (NP-DOX) improves the internalization and cytotoxic effects of the nano-carrier-mediated drug delivery in MG-63 human osteosarcoma cells. NP-DOX was designed and synthesized using the co-precipitation method. Highly stable and crystalline nanoparticles conjugated with DOX were internalized in MG-63 cells through macropinocytosis and located in the perinuclear area. Higher nanoparticles internalization in MG-63 cells previously exposed to 1 Gy X-rays was correlated with an early accumulation of cells in G₂/M, starting at 12 h after treatment. After 48 h, the application of the combined treatment led to higher cytotoxic effects compared to the individual treatment, with a reduction in the metabolic capacity and unrepaired DNA breaks, whilst a low percent of arrested cells, contributing to the commitment of mitotic catastrophe. NP-DOX showed hemocompatibility and no systemic cytotoxicity, nor histopathological alteration of the main organs.

The use of charged particle radiotherapy is currently increasing, but combination therapy with DNA repair inhibitors remains to be exploited in the clinic. The high-linear energy transfer (LET) radiation delivered by charged particles causes clustered DNA damage, which is particularly effective in destroying cancer cells. Whether the DNA damage response to this type of damage is different from that elicited in response to low-LET radiation, and if and how it can be targeted to increase treatment efficacy, is not fully understood. Although several preclinical studies have reported radiosensitizing effects when proton or carbon ion irradiation is combined with inhibitors of, e.g., PARP, ATR, ATM, or DNA-PKcs, further exploration is required to determine the most effective treatments. Here, we examine what is known about repair pathway choice in response to high- versus low-LET irradiation, and we discuss the effects of inhibitors of these pathways when combined with protons and carbon ions. Additionally, we explore the potential effects of DNA repair inhibitors on antitumor immune signaling upon proton and carbon ion irradiation. Due to the reduced effect on healthy tissue and better immune preservation, particle therapy may be particularly well suited for combination with DNA repair inhibitors.

Besides the direct effects of radiations, indirect effects are observed within the surrounding non-irradiated area; irradiated cells relay stress signals in this close proximity, inducing the so-called radiation-induced bystander effect. These signals received by neighboring unirradiated cells induce specific responses similar with those of direct irradiated cells. To understand the cellular response of bystander cells, we performed a 2D gel-based proteomic study of the chondrocytes receiving the conditioned medium of low-dose irradiated chondrosarcoma cells. The conditioned medium was directly analyzed by mass spectrometry in order to identify candidate bystander factors involved in the signal transmission. The proteomic analysis of the bystander chondrocytes highlighted 20 proteins spots that were significantly modified at low dose, implicating several cellular mechanisms, such as oxidative stress responses, cellular motility, and exosomes pathways. In addition, the secretomic analysis revealed that the abundance of 40 proteins in the conditioned medium of 0.1 Gy irradiated chondrosarcoma cells was significantly modified, as compared with the conditioned medium of non-irradiated cells. A large cluster of proteins involved in stress granules and several proteins involved in the cellular response to DNA damage stimuli were increased in the 0.1 Gy condition. Several of these candidates and cellular mechanisms were confirmed by functional analysis, such as 8-oxodG quantification, western blot, and wound-healing migration tests. Taken together, these results shed new lights on the complexity of the radiation-induced bystander effects and the large variety of the cellular and molecular mechanisms involved, including the identification of a new potential actor, namely the stress granules.