Abstract

Abstract

Patients with cancer and bone metastases can have an increased risk of fracturing their femur. Treatment is based on the impending fracture risk: patients with a high fracture risk are considered for prophylactic surgery, whereas low fracture risk patients are treated conservatively with radiotherapy to decrease pain. Current clinical guidelines suggest to determine fracture risk based on axial cortical involvement of the lesion on conventional radiographs, but that appears to be difficult. Therefore, we developed a patient-specific finite element (FE) computer model that has shown to be able to predict fracture risk in an experimental setting and in patients. The goal of this study was to determine whether patient-specific finite element (FE) computer models are better at predicting fracture risk for femoral bone metastases compared to clinical assessments based on axial cortical involvement on conventional radiographs, as described in current clinical guidelines.

45 patients (50 affected femurs) affected with predominantly lytic bone metastases who were treated with palliative radiotherapy for pain were included. CT scans were made and patients were followed for six months to determine whether or not they fractured their femur. Non-linear isotropic FE models were created with the patient-specific geometry and bone density obtained from the CT scans. Subsequently, an axial load was simulated on the models mimicking stance. Failure loads normalized for bodyweight (BW) were calculated for each femur. High and low fracture risks were determined using a failure load of 7.5 × BW as a threshold. Experienced assessors measured axial cortical involvement on conventional radiographs. Following clinical guidelines, patients with lesions larger than 30 mm were identified as having a high fracture risk. FE predictions were compared to clinical assessments by means of diagnostic accuracy values (sensitivity, specificity and positive (PPV) and negative predictive values (NPV)).

Seven femurs (14%) fractured during follow-up. Median time to fracture was 8 weeks. FE models were better at predicting fracture risk in comparison to clinical assessments based on axial cortical involvement (sensitivity 100% vs. 86%, specificity 74% vs. 42%, PPV 39% vs. 19%, and NPV 100% vs. 95%, for the FE computer model vs. axial cortical involvement, respectively). We concluded that patient-specific FE computer models improve fracture risk predictions of femoral bone metastases in advanced cancer patients compared to clinical assessments based on axial cortical involvement, which is currently used in clinical guidelines. Therefore, we are initiating a pilot for clinical implementation of the FE model.

Infections are among the main complications connected to implantation of biomedical devices, having high incidence rate and severe outcome. Since their treatment is challenging, prevention must be preferred. For this reason, solutions capable of exerting suitable efficacy while not causing toxicity and/or development of resistant bacterial strains are needed. To address infection, inorganic antibacterial coatings, and in particular silver coatings, have been extensively studied and used in the clinical practice, but some drawbacks have been evidenced, such as scarce adhesion to the substrate, delamination, or scarce control over silver release.

Here, antibacterial nanostructured silver-based thin films are proposed, obtained by a novel plasma-assisted technique, Ionized Jet Deposition (IJD). Coatings are obtained by deposition of metallic silver targets. Films thickness is selected based on previous results aimed at measuring extent and duration of silver release and at evaluating toxicity to host cells (fibroblasts). Here, composition (grazing incidence XRD) and morphology (SEM) of the obtained coatings are characterized for deposition onto different substrates, both metallic and polymeric. For heat sensitive substrates, possible alterations caused by coatings deposition in terms of morphology (SEM) and composition (FT-IR) is assessed. Then, a proof-of-concept study of the capability of these films to inhibit microbial biofilm formation is performed by using two different supports i.e., the Calgary Biofilm Device and the microplates. To the best of the Authors knowledge, this is the first study describing the application of specific anti-biofilm analyses to nanostructured coatings. In particular, anti-biofilm activities are tested against the following pathogenic strains: Escherichia (E.) coli NCTC12923, Staphylococcus (S.) aureus ATCC29213 and S. aureus 86. Among these, the strain 86 is not only pathogen but it also possesses several antibiotic resistance genes, allowing the evaluation of the utilization of nanostructured coatings as an alternative anti-microbial system to face the global threat of antibiotic resistance.

Results indicate that films deposited from silver targets are composed of nanosized aggregates of metallic silver, indicating a perfect transfer of composition from the deposition target to the coatings.

Results obtained here indicate that the films have significant antibacterial and antibiofilm activity. In addition, they prove that the system can be successfully applied for evaluation of coatings antibacterial efficacy for biomedical applications.

Calcium phosphates-based coatings have been widely studied to favour a firm bonding between orthopaedic implants and the host bone. To this aim, thin films (thickness below 1 μm) having high adhesion to the substrate and a nanostructured surface texture are desired, capable of boosting platelet, proteins and cells adhesion. In addition, a tunable composition is required to resemble as closely as possible the composition of mineralized tissues and/or to intentionally substitute ions having possible therapeutic functions. The authors demonstrated nanostructured films having high surface roughness and a composition perfectly resembling the deposition target one can be achieved by Ionized Jet Deposition (IJD). Highly adhesive nanostructured coatings were obtained by depositing bone-apatite like thin films by ablation of deproteinized bovine bone, capable of promoting host cells attachment, proliferation and differentiation. Here, biomimetic films are deposited by IJD, using biogenic and synthetic apatite targets. Since IJD deposition can be carried out without heating the substrate, application on heat sensitive polymeric substrate, i.e. 3D printed porous scaffolds, is investigated.

Biogenic apatite coatings are obtained by deposition of deproteinized bone (bovine, ovine, equine, porcine) and compared to ones of stoichiometry hydroxyapatite (HAp). Coatings composition (FT-IR-ATR, FT-IR microscopy, XRD, EDS) and morphology (SEM, AFM) are tested for deposition onto metallic and 3D-printed polymeric substrates (polyurethane (PU)). Different post-treatment annealing procedures for metallic substrates are compared (350–425°C), to optimize crystallinity. Then, uniformity of substrate coverage and possible damage caused to the polymeric substrate are studied by SEM, DSC and FT-IR microscopy.

Biogenic coatings are composed by carbonated HAp (XRD, FT-IR). Trace ions Na⁺ and Mg²⁺ are transferred from deposition target to coating. All coatings are nanostructured, composed by nano-sized globular aggregates, of which morphology and dimensions depend on the target characteristics. As-deposited coatings are amorphous, but crystallinity can be tuned by post-treatment annealing. A bone-like crystallinity can be achieved for heating at ≥400°C, also depending on duration. When deposited on 3D-printed PU scaffolds, coatings, owing to sub-micrometric thickness, coat them entirely, without altering their fibre shape and porosity.

Obtained biomimetic bone apatite coatings can be deposited onto a variety of metallic and polymeric biomedical devices, thus finding several perspective applications in biomedical field.

Abstract

Osteoarthritis (OA) is a leading cause of joint deformity and functional limitation. An imbalance of anabolic and catabolic activity results in destruction of the extracellular matrix of articular cartilage. There is evidence to support the role of DNA methylation in the pathogenesis of OA, but the effect of other epigenetic modifiers is yet to be described. This study looks at the effect of novel epigenetic modulators, PFI-1, a bromodomain inhibitor, and SGC707, a histone methytransferase inhibitor, and their effects on gene expression in the pathogenesis of OA. Chondrocytes were extracted from OA femoral heads (n=6), cultured and incubated. Samples were treated with media alone (control), interleukin 1-beta (IL-1β) plus oncostatin M (OSM) alone, or in combination with increasing concentrations of PFI-1 or SGC707. Levels of expression of iNOS, COX2, IL8, IL1B, matrix metalloproteinase-13 (MMP13), RUNX2 and COL9A1 were measured using qRT-PCR, and expressed relative to GAPDH. PFI-1 (0.5 and 5µM) suppressed expression of catabolic genes in OA chondrocytes, at basal levels and when co-stimulated with IL-1β+OSM. Catabolic gene expression decreased (iNOS, COX2, IL-8, IL-1β and MMP), and RUNX2 expression was also supressed. There was no effect on expression of the anabolic gene COL9A1. SGC707 (0.1 and 1µM) did not induce a reduction in expression of all the catabolic genes. This study has demonstrated that PFI-1 has a potent protective effect against cartilage degradation, by modulating the expression of catabolic genes in OA chondrocytes. This further validates the role of epigenetics in OA, with implications for therapeutic interventions in the future.

BACKGROUND

Although osteoarthritis (OA) is not an inflammatory arthritis, a characteristic feature of OA is increased production of pro-inflammatory cytokines, such as interleukin 1beta (IL-1b), by articular chondrocytes. In fact, the degree of articular inflammation is often associated with disease progression; indicating that this process probably contributes to articular damage. Suppressor of cytokine signalling (SOCS) proteins are, as the name suggests, inhibitors of cytokine signalling that function via the JAK/STAT pathway (Janus kinase/signal transducers and activators of transcription). Eight SOCS proteins, SOCS1-SOCS7 and CIS-1 (cytokine-inducible SH2-domain-1 with similar structure to the other SOCS proteins) have been identified, of which, SOCS1-3 and CIS-1 are the best characterised. Reduced expression of SOCS proteins would be predicted to result in increased cytokine responsiveness and thereby could contribute to OA pathology.