Growing Role of 3D In Vitro Cell Cultures in the Study of Cellular and Molecular Mechanisms: Short Focus on Breast Cancer, Endometriosis, Liver and Infectious Diseases.

Over the past decade, the development of three-dimensional (3D) models has increased exponentially, facilitating the unravelling of fundamental and essential cellular mechanisms by which cells communicate with each other, assemble into tissues and organs and respond to biochemical and biophysical stimuli under both physiological and pathological conditions. This section presents a concise overview of the most recent updates on the significant contribution of different types of 3D cell cultures including spheroids, organoids and organ-on-chip and bio-printed tissues in advancing our understanding of cellular and molecular mechanisms. The case studies presented include the 3D cultures of breast cancer (BC), endometriosis, the liver microenvironment and infections. In BC, the establishment of 3D culture models has permitted the visualization of the role of cancer-associated fibroblasts in the delivery of exosomes, as well as the significance of the physical properties of the extracellular matrix in promoting cell proliferation and invasion. This approach has also become a valuable tool in gaining insight into general and specific mechanisms of drug resistance. Given the considerable heterogeneity of endometriosis, 3D models offer a more accurate representation of the in vivo microenvironment, thereby facilitating the identification and translation of novel targeted therapeutic strategies. The advantages provided by 3D models of the hepatic environment, in conjunction with the high throughput characterizing various platforms, have enabled the elucidation of complex molecular mechanisms underlying various threatening hepatic diseases. A limited number of 3D models for gut and skin infections have been developed. However, a more profound comprehension of the spatial and temporal interactions between microbes, the host and their environment may facilitate the advancement of in vitro, ex vivo and in vivo disease models. Additionally, it may pave the way for the development of novel therapeutic approaches in diverse research fields. The interested reader will also find concluding remarks on the challenges and prospects of using 3D cell cultures for discovering cellular and molecular mechanisms in the research areas covered in this review.

Surface Properties of a Biocompatible Thermoplastic Polyurethane and Its Anti-Adhesive Effect against E. coli and S. aureus.

Thermoplastic polyurethane (TPU) is a polymer used in a variety of fields, including medical applications. Here, we aimed to verify if the brush and bar coater deposition techniques did not alter TPU properties. The topography of the TPU-modified surfaces was studied via AFM demonstrating no significant differences between brush and bar coater-modified surfaces, compared to the un-modified TPU (TPU Film). The effect of the surfaces on planktonic bacteria, evaluated by MTT assay, demonstrated their anti-adhesive effect on E. coli, while the bar coater significantly reduced staphylococcal planktonic adhesion and both bacterial biofilms compared to other samples. Interestingly, Pearson's R coefficient analysis showed that Ra roughness and Haralick's correlation feature were trend predictors for planktonic bacterial cells adhesion. The surface adhesion property was evaluated against NIH-3T3 murine fibroblasts by MTT and against human fibrinogen and human platelet-rich plasma by ELISA and LDH assay, respectively. An indirect cytotoxicity experiment against NIH-3T3 confirmed the biocompatibility of the TPUs. Overall, the results indicated that the deposition techniques did not alter the antibacterial and anti-adhesive surface properties of modified TPU compared to un-modified TPU, nor its bio- and hemocompatibility, confirming the suitability of TPU brush and bar coater films in the biomedical and pharmaceutical fields.

Photodynamic toluidine blue-gold nanoconjugates as a novel therapeutic for Staphylococcal biofilms.

Staphylococci are among the most frequent bacteria known to cause biofilm-related infections. Pathogenic biofilms represent a global healthcare challenge due to their high tolerance to antimicrobials. In this study, water soluble polyethylene glycol (PEG)-coated gold nanospheres (28 ppm) and nanostars (15 ppm) with electrostatically adsorbed photosensitizer (PS) Toluidine Blue O (TBO) ∼4 µM were successfully synthesized and characterized as PEG-GNPs@TBO and PEG-GNSs@TBO. Both nanoconjugates and the TBO 4 µM solution showed remarkable, if similar, antimicrobial photodynamic inactivation (aPDI) effects at 638 nm, inhibiting the formation of biofilms by two Staphylococcal strains: a clinical methicillin-resistant Staphylococcus aureus (MRSA) isolate and Staphylococcus epidermidis (S. epidermidis) RP62A. Alternatively in biofilm eradication treatments, the aPDI effects of PEG-GNSs@TBO were more effective and yielded a 75% and 50% reduction in viable count of MRSA and S. epidermidis RP62A preformed biofilms, respectively and when compared with untreated samples. This reduction in viable count was even greater than that obtained through aPDI treatment using a 40 µM TBO solution. Confocal laser microscopy (CLSM) and scanning electron microscope (SEM) images of PEG-GNSs@TBO's aPDI treatments revealed significant changes in the integrity and morphology of biofilms, with fewer colony masses. The generation of reactive oxygen species (ROS) upon PEG-GNSs@TBO's aPDI treatment was detected by CLSM using a specific ROS fluorescent probe, demonstrating bright fluorescence red spots across the surfaces of the treated biofilms. Our findings shine a light on the potential synergism between gold nanoparticles (AuNPs) and photosensitizers in developing novel nanoplatforms to target Staphylococcal biofilm related infections.

Effect of Low Copper Doping on the Optical, Cytocompatible, Antibacterial, and SARS-CoV-2 Trapping Properties of Calcium Phosphate Glasses.

Calcium phosphate glasses (CPGs) are acquiring great importance in the biomedical field because of their thermomechanical and bioresorbable properties. In this study, optically transparent copper (1 mol %)-doped calcium phosphate glasses (CPGs_Cu) were prepared through the melt-quenching method, and their biocompatibility and antibacterial and antiviral properties were evaluated and compared with undoped CPGs. Biocompatibility was evaluated on murine fibroblast NIH-3T3 cells as a preliminary study of cytocompatibility. The in vitro tests were performed through indirect and direct cytotoxicity analyses by MTT and Alamar Blue assays and supported by electron microscopy observations. Microbiological analyses were performed against the most common Gram-negative and Gram-positive pathogens that cause nosocomial infections: Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae, Staphylococcus aureus, and the methicillin-resistant Staphylococcus aureus strain. In addition, the bioglass samples were exposed to SARS-CoV-2 to assess their effects on viral survival. The obtained results assessed the biocompatibility of both bioglass types and their ability to reduce the viral load and trap the virus. In addition, Cu2+-doped bioglass was found to be antibacterial despite its low content (1 mol %) of copper, making this a promising candidate material for biomedical applications, e.g., surgery probes, drug delivery, and photodynamic therapy.

Enhanced in vitro immersion behavior and antibacterial activity of NiTi orthopedic biomaterial by HAp-Nb2O5 composite deposits.

NiTi is a class of metallic biomaterials, benefit from superelastic behavior, high biocompatibility, and favorable mechanical properties close to that of bone. However, the Ni ion leaching, poor bioactivity, and antibacterial activity limit its clinical applications. In this study, HAp-Nb2O5 composite layers were PC electrodeposited from aqueous electrolytes containing different concentrations of the Nb2O5 particles, i.e., 0-1 g/L, to evaluate the influence of the applied surface engineering strategy on in vitro immersion behavior, Ni2+ ion leaching level, and antibacterial activity of the bare NiTi. Surface characteristics of the electrodeposited layers were analyzed using SEM, TEM, XPS, and AFM. The immersion behavior of the samples was comprehensively investigated through SBF and long-term PBS soaking. Gram-negative Escherichia coli (E. coli) and Gram-positive Staphylococcus aureus (S. aureus) infective reference bacteria were employed to address the antibacterial activity of the samples. The results illustrated that the included particles led to more compact and smoother layers. Unlike bare NiTi, composite layers stimulated apatite formation upon immersion in both SBF and PBS media. The concentration of the released Ni2+ ion from the composite layer, containing 0.50 g/L Nb2O5 was ≈ 60% less than that of bare NiTi within 30 days of immersion in the corrosive PBS solution. The Nb2O5-reinforced layers exhibited high anti-adhesive activity against both types of pathogenic bacteria. The hybrid metallic-ceramic system comprising HAp-Nb2O5-coated NiTi offers the prospect of a potential solution for clinical challenges facing the orthopedic application of NiTi.

Strontium-doped apatitic bone cements with tunable antibacterial and antibiofilm ability.

Injectable calcium phosphate cements (CPCs) represent promising candidates for the regeneration of complex-shape bone defects, thanks to self-hardening ability, bioactive composition and nanostructure offering high specific surface area for cell attachment and conduction. Such features make CPCs also interesting for functionalization with various biomolecules, towards the generation of multifunctional devices with enhanced therapeutic ability. In particular, strontium-doped CPCs have been studied in the last years due to the intrinsic antiosteoporotic character of strontium. In this work, a SrCPC previously reported as osteointegrative and capable to modulate the fate of bone cells was enriched with hydroxyapatite nanoparticles (HA-NPs) functionalized with tetracycline (TC) to provide antibacterial activity. We found that HA-NPs functionalized with TC (NP-TC) can act as modulator of the drug release profile when embedded in SrCPCs, thus providing a sustained and tunable TC release. In vitro microbiological tests on Escherichia coli and Staphylococcus aureus strains proved effective bacteriostatic and bactericidal properties, especially for the NP-TC loaded SrCPC formulations. Overall, our results indicate that the addition of NP-TC on CPC acted as effective modulator towards a tunable drug release control in the treatment of bone infections or cancers.

Gold Nanoparticles: Can They Be the Next Magic Bullet for Multidrug-Resistant Bacteria?

In 2017 the World Health Organization (WHO) announced a list of the 12 multidrug-resistant (MDR) families of bacteria that pose the greatest threat to human health, and recommended that new measures should be taken to promote the development of new therapies against these superbugs. Few antibiotics have been developed in the last two decades. Part of this slow progression can be attributed to the surge in the resistance acquired by bacteria, which is holding back pharma companies from taking the risk to invest in new antibiotic entities. With limited antibiotic options and an escalating bacterial resistance there is an urgent need to explore alternative ways of meeting this global challenge. The field of medical nanotechnology has emerged as an innovative and a powerful tool for treating some of the most complicated health conditions. Different inorganic nanomaterials including gold, silver, and others have showed potential antibacterial efficacies. Interestingly, gold nanoparticles (AuNPs) have gained specific attention, due to their biocompatibility, ease of surface functionalization, and their optical properties. In this review, we will focus on the latest research, done in the field of antibacterial gold nanoparticles; by discussing the mechanisms of action, antibacterial efficacies, and future implementations of these innovative antibacterial systems.

Targeting the "Sweet Side" of Tumor with Glycan-Binding Molecules Conjugated-Nanoparticles: Implications in Cancer Therapy and Diagnosis.

Nanotechnology is in the spotlight of therapeutic innovation, with numerous advantages for tumor visualization and eradication. The end goal of the therapeutic use of nanoparticles, however, remains distant due to the limitations of nanoparticles to target cancer tissue. The functionalization of nanosystem surfaces with biological ligands is a major strategy for directing the actions of nanomaterials specifically to tumor cells. Cancer formation and metastasis are accompanied by profound alterations in protein glycosylation. Hence, the detection and targeting of aberrant glycans are of great value in cancer diagnosis and therapy. In this review, we provide a brief update on recent progress targeting aberrant glycosylation by functionalizing nanoparticles with glycan-binding molecules (with a special focus on lectins and anti-glycan antibodies) to improve the efficacy of nanoparticles in cancer targeting, diagnosis, and therapy and outline the challenges and limitations in implementing this approach. We envision that the combination of nanotechnological strategies and cancer-associated glycan targeting could remodel the field of cancer diagnosis and therapy, including immunotherapy.

Polyurethane-Based Coatings with Promising Antibacterial Properties.

In coatings technology, the possibility of introducing specific characteristics at the surface level allows for the manufacture of medical devices with efficient and prolonged antibacterial properties. This efficiency is often achieved by the use of a small amount of antibacterial molecules, which can fulfil their duty while limiting eventual releasing problems. The object of this work was the preparation and characterization of silver, titanium dioxide and chitosan polyurethane-based coatings. Coatings with the three antibacterials were prepared using different deposition techniques, using a brush or a bar coater automatic film applicator, and compared to solvent casted films prepared with the same components. For silver containing materials, an innovative strategy contemplating the use and preparation of silver nanoparticles in a single step-method was employed. This preparation was obtained starting from a silver precursor and using a single compound as the reducing agent and stabilizer. Ultraviolet-visible spectroscopy, scanning electron microscopy, energy dispersive X-ray spectroscopy, contact angle measurements and adhesion test experiments were used to characterize the prepared coatings. Promising antibacterial properties, measured via direct and indirect methods, were registered for all the silver-based materials.

Synergic Effect of Nanolignin and Metal Oxide Nanoparticles into Poly(l-lactide) Bionanocomposites: Material Properties, Antioxidant Activity, and Antibacterial Performance.

Binary and ternary poly(l-lactide) (PLLA)-based nanocomposites, containing nanolignin (1 wt %) and different metal oxide nanoparticles (0.5 wt %, Ag2O, TiO2, WO3, Fe2O3, and ZnFe2O4), were realized by solvent casting, and their morphological, thermal, surface, optical, antioxidant, and antimicrobial characterizations were performed. The presence of metal oxide nanoparticles at the selected weight concentration affects the surface microstructure of the PLLA polymer, and this outcome is particle-type dependent, according to the shape, morphology, and chemical properties of the selected nanoparticles (NPs). Analogously, wettability of PLLA-based nanocomposites was slightly modified by the presence of hydrophobic lignin nanoparticles and different shaped metal oxides. Results of differential scanning calorimetry (DSC) and X-ray powder diffraction (XRD) tests confirmed that nanoparticle addition confined the mobility of the amorphous phase, increasing at the same time the formation of more numerous but less perfect PLLA crystals. Interestingly, antioxidant activity was also obtained in ternary-based nanocomposites, where a synergic effect of lignin and metal oxide nanoparticles was obtained. Antibacterial tests showed manifest activity of TiO2 and Ag2O nanoparticles containing PLLA films, and the time dependence was more evident for Staphylococcus aureus than for Escherichia coli. Lignin nanoparticles are able to provide protection against UV light while still allowing visible light to pass and even surpass the UV-protection capacity provided by many inorganic nanoparticles. This makes them an attractive renewable additive for the realization of PLLA/metal oxide nanocomposites in the fields of food, drug packaging, and biomedical industry, where antibacterial and antioxidant properties are required.