Development of a bone tissue-engineered construct to enhance new bone formation in revision total hip replacement

The main issue associated with revision total hip replacements (rTHRs) is how to generate new bone and restore bone stock for fixation of the revision stem. Bone tissue engineering (BTE) seeks the generation of constructs ex vivo in order to replace damaged or lost bone. The aim of this thesis was to develop a bone tissue-engineered construct with a calcium-phosphate (CaP) coated porous metal scaffold seeded throughout its structure with mesenchymal stem cells (MSCs) in order to enhance new bone formation at rTHRs. The study had in vitro and in vivo phases. For the in vitro phase, CaP coatings by biomimetic and electrochemical methods on the surface of titanium and tantalum discs were investigated and seeded with MSCs under static culture conditions. Different coating methods produced different morphologies and compositions with biomimetic coatings enhancing MSCs growth while the electrochemical ones enhanced their osteogenic potential. An electrochemically CaP coated porous titanium cylinder was seeded with MSCs and dynamically cultured in a perfusion bioreactor, showing an increased MSCs proliferation and osteogenic differentiation and an even distribution of cells throughout the scaffolds compared to statically cultured constructs. Tissue-engineered constructs in the perfusion bioreactor were evaluated in vivo by implantation in the medial femoral condyle of sheep with and without gap. Their osseointegration and implant-bone fixation strength were compared to non tissue-engineered constructs. The results showed that the addition of MSCs to the scaffolds did not significantly increase osseointegration or implant-bone fixation strength. However, in the defects with gap the tissue-engineered constructs showed a higher implant-bone contact area and therefore higher forces were necessary to push the tissue-engineered implants out of the bone in the defects with gap than for the non tissue-engineered ones. In conclusion, BTE can be applied in order to develop constructs with a clinical application in rTHRs where a lack of bone stock is problematic

Decellularization of tumours: A new frontier in tissue engineering

Cancer is one of the leading causes of death worldwide. The tumour extracellular matrix (ECM) has unique features in terms of composition and mechanical properties, resulting in a structurally and chemically different ECM to that of native, healthy tissues. This paper reviews to date the efforts into decellularization of tumours, which in the authors’ view represents a new frontier in the ever evolving field of tumour tissue engineering. An overview of the ECM and its importance in cancer is given, ending with examples of research using decellularized tumours, which has already indicated potential therapeutic targets, unravelled malignancy mechanisms or response to chemotherapy agents. The review highlights that more research is needed in this area, which can answer important questions related to tumour formation and progression to ultimately identify new and effective therapeutic targets. Within the near-future of personalized medicine, this research can create patient-specific tumour models and therapeutic regimes

Chronic Leg Ulcers: Are Tissue Engineering and Biomaterials Science the Solution?

Chronic leg ulcers (CLUs) are full thickness wounds that usually occur between the ankle and knee, fail to heal after 3 months of standard treatment, or are not entirely healed at 12 months. CLUs present a considerable burden on patients, subjecting them to severe pain and distress, while healthcare systems suffer immense costs and loss of resources. The poor healing outcome of the standard treatment of CLUs generates an urgent clinical need to find effective solutions for these wounds. Tissue Engineering and Biomaterials Science offer exciting prospects for the treatment of CLUs, using a broad range of skin substitutes or scaffolds, and dressings. In this review, we summarize and discuss the various types of scaffolds used clinically in the treatment of CLUs. Their structure and therapeutic effects are described, and for each scaffold type representative examples are discussed, supported by clinical trials. Silver dressings are also reviewed due to their reported benefits in the healing of leg ulcers, as well as recent studies on new dermal scaffolds, reporting on clinical results where available. We conclude by arguing there is a further need for tissue-engineered products specifically designed and bioengineered to treat these wounds and we propose a series of properties that a biomaterial for CLUs should possess, with the intention of focusing efforts on finding an effective treatment

The effect of temperature on the viability of human mesenchymal stem cells

Introduction Impaction allograft with cement is a common technique used in revision hip surgeries for the last 20 years. However, its clinical results are inconsistent. Recent studies have shown that mesenchymal stem cells (MSCs) seeded onto allograft can enhance bone formation. This in vitro study investigates whether the increase in temperature related to the polymerisation of bone cement will affect the viability of human MSCs. Methods The viability of human MSCs was measured after incubating them at temperatures of 38°C, 48°C and 58°C; durations 45 seconds, 80 seconds and 150 seconds. A control group was kept at 37°C and 5% carbon dioxide for the duration of the investigation (7 days). During the course of the study the human MSCs were analysed for cell metabolic activity using the alamarBlue™ assay, cell viability using both Trypan Blue dye exclusion and calcein staining under fluorescent microscopy, and necrosis and apoptosis using Annexin V and propidium iodide for flow cytometric analysis. A one-way analysis of variance with a priori Dunnett’s test was used to indicate the differences between the treatment groups, when analysed against the control. This identified conditions with a significant difference in cell metabolic activity (alamarBlue™) and cell viability (Trypan Blue). Results Results showed that cell metabolism was not severely affected up to 48°C/150 seconds, while cells in the 58°C group died. Similar results were shown using Trypan Blue and calcein analysis for cell viability. No significant difference in apoptosis and necrosis of the cells was observed when human MSCs treated at 48°C/150 seconds were compared with the control group. Conclusions The study suggests that human MSCs seeded onto allograft can be exposed to temperatures up to 48°C for 150 seconds. Exposure to this temperature for this time period is unlikely to occur during impaction allograft surgery when cement is used. Therefore, in many situations, the addition of human MSCs to cemented impaction grafting may be carried out without detrimental effects to the cells. Furthermore, previous studies have shown that this can enhance new bone formation and repair the defects in revision situations

Biofilm formation in total hip arthroplasty: Prevention and treatment

Biomaterials science is a very active area of research, which has allowed the successful use of implants in the orthopaedic field for over a century. However, implant infection remains a clinical concern as it is associated with extensive patient morbidity and a high economic burden, which is predicted to increase due to an ageing population. Bacteria are able to adhere, colonise and develop into biofilms on the surface of biomaterials making associated infections physiologically different to other post-surgical infections. Unfortunately, biofilms exert increased protection from the host immune system and an increased resistance to antibiotic therapy in comparison to their planktonic counterparts. The aim of this review is to assess the current knowledge on treatments, pathogenesis and the prevention of infections associated with orthopaedic implants, with a focus on total hip arthroplasty

Biomimetic surface functionalization of clinically relevant metals used as orthopaedic and dental implants

Titanium and its alloys or tantalum (Ta) are materials used in orthopaedic and dental implants due to their excellent mechanical properties and biocompatibility. However, their bioactivity and osteoconductivity is low. With a view to improving the bioactivity of these materials we hypothesised that the surface of Ta and TiAl6V4 can be functionalised with biomimetic, amorphous nano-sized calcium phosphate (CaP) apatite-like deposits, instead of creating uniform coatings, which can lead to flaking, delamination and poor adherence. We used Ta and TiAl6V4 metal discs with smooth and rough surfaces. Amorphous CaP apatite-like particles were deposited on the different surfaces by a biomimetic rapid two-step soaking method using concentrated simulated body fluid (SBF) solutions without a pre-treatment of the metal surfaces to induce CaP deposition. Immersion times in the second SBF solution of 48 and 18 h for Ta and TiAl6V4 respectively produced CaP deposits composed of amorphous globular nano-sized particles that also contained Mg, C and O. Longer immersion times produced more uniform coatings as well as an undesired calcite mineral phase. Prediction of in vivo behaviour by immersion in regular SBF showed that the obtained CaP deposits would act as a catalyst to rapidly form a Ca deficient CaP layer that also incorporates Mg. The amorphous CaP apatite-like deposits promoted initial attachment, proliferation and osteogenic differentiation of bone marrow derived mesenchymal stem cells. Finally, we used our method to functionalise 3D porous structures of titanium alloy made by selective laser sintering. Our study uses a novel and cost-effective approach to functionalise clinically relevant metal surfaces in order to increase the bioactivity of these materials, which could improve their clinical performance

In vitro hydrolytic degradation of polyester-based scaffolds under static and dynamic conditions in a customized perfusion bioreactor

Creating biofunctional artificial scaffolds could potentially meet the demand of patients suffering from bone defects without having to rely on donors or autologous transplantation. Three-dimensional (3D) printing has emerged as a promising tool to fabricate, by computer design, biodegradable polymeric scaffolds with high precision and accuracy, using patient-specific anatomical data. Achieving controlled degradation profiles of 3D printed polymeric scaffolds is an essential feature to consider to match them with the tissue regeneration rate. Thus, achieving a thorough characterization of the biomaterial degradation kinetics in physiological conditions is needed. Here, 50:50 blends made of poly(e-caprolactone)–Poly(D, L-lactic-co-glycolic acid (PCL-PLGA) were used to fabricate cylindrical scaffolds by 3D printing (Ø 7 × 2 mm). Their hydrolytic degradation under static and dynamic conditions was characterized and quantified. For this purpose, we designed and in-house fabricated a customized bioreactor. Several techniques were used to characterize the degradation of the parent polymers: X-ray Photoelectron Spectroscopy (XPS), Gel Permeation Chro-matography (GPC), Scanning Electron Microscopy (SEM), evaluation of the mechanical properties, weigh loss measurements as well as the monitoring of the degradation media pH. Our results showed that flow perfusion is critical in the degradation process of PCL-PLGA based scaffolds implying an accelerated hydrolysis compared to the ones studied under static conditions, and up to 4 weeks are needed to observe significant degradation in polyester scaffolds of this size and chemical composition. Our degradation study and characterization methodology are relevant for an accurate design and to tailor the physicochemical properties of polyester-based scaffolds for bone tissue engineering

Role of hydrogen peroxide in intra-operative wound preparation based on an in vitro fibrin clot degradation model

Three per cent hydrogen peroxide (H2O2) is widely used to irrigate acute and chronic wounds in the surgical setting and clinical experience tells us that it is more effective at removing dried-on blood than normal saline alone. We hypothesise that this is due to the effect of H2O2 on fibrin clot architecture via fibrinolysis. We investigate the mechanisms and discuss the clinical implications using an in vitro model. Coagulation assays with normal saline (NaCl), 1% and 3% concentrations of H2O2 were performed to determine the effect on fibrin clot formation. These effects were confirmed by spectrophotometry. The effects of 1%, 3% and 10% H2O2 on the macroscopic and microscopic features of fibrin clots were assessed at set time intervals and compared to a NaCl control. Quantitative analysis of fibrin networks was undertaken to determine the fibre length, diameter, branch point density and pore size. Fibrin clots immersed in 1%, 3% and 10% H2O2 demonstrated volume losses of 0.09-0.25mm3/min, whereas those immersed in the normal saline gained in volume by 0.02±0.13 mm3/min. Quantitative analysis showed that H2O2 affects the structure of the fibrin clot in a concentration-dependent manner, with the increase in fibre length, diameter and consequently pore sizes. Our results support our hypothesis that the efficacy of H2O2 in cleaning blood from wounds is enhanced by its effects on fibrin clot architecture in a concentration- and time-dependent manner. The observed changes in fibre size and branch point density suggest that H2O2 is acting on the quaternary structure of the fibrin clot, most likely via its effect on cross-linking of the fibrin monomers and may therefore be of benefit for the removal of other fibrin-dependent structures such as wound slough

Collagen-laponite nanoclay hydrogels for tumor spheroid growth

The extracellular matrix (ECM) plays an important regulatory role in the development and progression of tumoral tissue. Its functions and properties are crucial in determining tumor cell behavior such as invasion, migration, and malignancy development. Our study explores the role of collagen type I in cancer development and spread using engineered tumor models like multicellular spheroids grown in collagen-based hydrogels to simulate early tumor formation. We employ microfluidic techniques to test the hypothesis that (i) adding Laponite nanoclay to collagen hydrogels modifies mechanical and rheological properties and (ii) changing the stiffness of the collagen microenvironment affects tumor spheroid growth. Our findings support our theories and suggest the use of ECM components and engineered tumor models in cancer research, offering a biocompatible and biomimetic method to tailor the mechanical properties of conventional collagen hydrogels

Biomimetic In vitro model of cell infiltration into skin scaffolds for pre-screening and testing of biomaterial-based therapies

Due to great clinical need, research where different biomaterials are tested as 3D scaffolds for skin tissue engineering has increased. In vitro studies use a cell suspension that is simply pipetted onto the material and cultured until the cells migrate and proliferate within the 3D scaffold, which does not mimic the in vivo reality. Our aim was to engineer a novel biomimetic in vitro model that mimics the natural cell infiltration process occurring in wound healing, thus offering a realistic approach when pre-screening and testing new skin substitutes. Our model consists of porous membrane cell culture inserts coated with gelatin and seeded with human dermal fibroblasts, inside which two different commercially available dermal substitutes were placed. Several features relevant to the wound healing process (matrix contraction, cell infiltration and proliferation, integration of the biomaterial with the surrounding tissue, and secretion of exogenous cytokines and growth factors) were evaluated. Our results showed that cells spontaneously infiltrate the materials and that our engineered model is able to induce and detect subtle differences between different biomaterials. The model allows for room for improvements or “adds-on” and miniaturization and can contribute to the development of functional and efficient skin substitutes for burns and chronic wounds