7th Dutch Bio-Medical Engineering Conference
January 24th & 25th 2019, Egmond aan Zee, the Netherlands
13:00   Joints
Chair: Marloes Groot
15 mins
Estimation of Ankle Joint Stiffness Via System Identification Techniques During Functional Movement
Ronald C. van 't Veld, Alejandro Moya Esteban, Alfred C. Schouten
Abstract: Introduction Human limb mechanical properties can be quantified using joint impedance, the dynamic relation between joint angle deviations and corresponding joint torque, with joint stiffness as its position-dependent component. Joint stiffness is key in postural control and movement and well understood within (quasi-)static conditions [1]. However, muscle and joint mechanics change during functional movements, consequently also the joint stiffness changes [2,3]. Monitoring and understanding the continuous modulation of joint stiffness during functional movements in able-bodied subjects has several technical and clinical applications. For example, it can be applied to the design of next-generation biomimetic prostheses and exoskeletons or to advance clinical assessment and decision making. Therefore, the goal of this study is to develop an experimental protocol to assess how humans modulate ankle joint stiffness in time-varying conditions. Methods Six healthy adults (5 male, 24.2±1.0y) participated in this study. Participants were seated with their right foot connected to a single axis actuator using a footplate. Muscle activity of the soleus, gastrocnemius and tibialis anterior muscles was recorded using electromyography. The experiment consisted of three conditions: a 5Nm amplitude sinusoidal torque tracking task against a rigid actuator (static posture), and a 0.15rad amplitude slow (0.3Hz) and fast (0.6Hz) sinusoidal position tracking task against a virtual spring resulting in a torque amplitude of 9Nm (dynamic posture). Additionally, pseudo-random binary sequence (PRBS) perturbations were applied during the tasks (±0.015 rad, 0.15s switching time). System identification was performed using an ensemble-based algorithm, which produced ankle joint stiffness estimates by means of multiple short segments of the torque and angle recordings [2]. Results and Discussion Across subjects and conditions two maximum stiffness peaks, corresponding to plantarflexion and dorsiflexion phases, were observed followed by periods of decreased stiffness. The periods of high and low stiffness matched periods of high and low muscle activation. The average estimated joint stiffness magnitude across conditions ordered from low to high was: fast dynamic, static and slow dynamic posture. Our results highlight the variability in stiffness modulation strategies across conditions, especially across movement frequency. References [1] Kearney RE, Hunter IW. Crit Rev Biomed Eng. 1990; 18(I):55-87. [2] Ludvig D et al. Exp Brain Res. 2017; 235(10):2959-70. [3] Dietz V, Sinkjaer T. Lancet Neurol. 2007; 6(8):725-33.
15 mins
Self-Defending Bone Implants
Ingmar van Hengel, Niko Eka Putra, Melissa Tierolf, Francisca Gelderman, Vito Valerio, Stefanos Athanasiadis, Raisa Grotenhuis, Ad Fluit, Bram van der Eerden, Lidy Fratila-Apachitei, Iulian Apachitei, Amir Zadpoor
Abstract: Everyone knows someone who received a hip or knee implant. Together with the number of patients requiring such an implant, the number of complications is on the rise: implant-associated infections and aseptic loosening have great impact on quality of life and functioning in society. We aim to prevent these complications through the generation of self-defending, multifunctional bone implants that simultaneously prevent infection and enhance fixation of the implant by stimulation of bone stem cells. For this purpose we designed highly porous titanium implants that were additively manufactured by selective laser melting (SLM) resulting in implants with a 4 times enhanced surface area compared to solid implants. Subsequently, the implant’s surface was biofunctionalized using an electrochemical surface modification method, namely plasma electrolytic oxidation (PEO). Through addition of calcium and phosphate species as well as silver nanoparticles (AgNPs) to the PEO electrolyte these elements were incorporated in the growing TiO2 layer during the PEO process. It is important to note that the AgNPs become fully immobilized in the surface, thereby preventing the NPs from free circulation and potential nanotoxic effects. Following PEO processing, the surface morphology, phase and chemical composition as well as ion release kinetics were studied. The antibacterial capacity was evaluated against methicillin-resistant Staphylococcus aureus (MRSA), a highly resistant bacteria that is frequently involved in implant-associated infections. Meanwhile, osteogenesis was studied using human mesenchymal stem cells (MSC). The PEO treatment of the highly porous implants resulted in a bioactive surface with interconnected micropores in which hydroxyapatite phases were demonstrated. Release of silver ions from the implant surface was demonstrated for at least 28 days, indicating a long-lasting infection prevention capacity. Furthermore, the surface proved to stimulate the differentiation of mesenchymal stem cells (MSC) towards osteoblasts, thereby stimulating osseointegration and proper fixation between bone tissue and implant. At the same time, the porous implants demonstrated enhanced in vitro and ex vivo antibacterial activity against MRSA compared to solid implants. Currently, we are testing other antimicrobial and osteogenic elements to further improve these capacities and generate a self-defending implant applicable for the clinic.
15 mins
Extracellular Matrix-Derived Bioink for Elastic Cartilage Bioprinting: Implications for Chondrogenesis
Ludo van Haasterecht, Dafydd Visscher, Yigitcan Sumbelli, M.N. Helder, J.P.J Don Griot, M.L. Groot, P.P.M van Zuijlen
Abstract: Background: Reconstruction of extensively damaged facial cartilage is a challenging task. As autologous and synthetic grafts have significant disadvantages, novel reconstructive strategies are required to provide patients with durable, patient-specific cartilage tissue. Three-dimensional bioprinting has the potential to solve several issues encountered in tissue engineering. However current bioinks, containing both cells and hydrogel, often miss the necessary chondrogenic cues for adequate cartilage formation. The addition of decellularized cartilage powder to the bioink may provide chondrocytes with the necessary signals required for adequate cell proliferation and chondrogenesis. Objective: The objectives were: i) To develop a cartilage-specific hydrogel with adequate bioprinting capabilities, and ii) to support and improve chondrocyte survival and post-printing neocartilage formation. Methods: Native caprine auricular cartilage was decellularized and enzymatically digested to produce extracellular matrix (ECM)-fragments. The effect of ECM addition to gelatin/alginate-based bioink of different concentrations was evaluated. Rheological properties, bioprintability, and swelling capacity, as well as cell survival were evaluated for both the gelatin/alginate bioink and the ECM-containing bioink. Following a culture time of three weeks, the bioprinted scaffolds were subjected to higher harmonic generation microscopy. Second Harmonic Generation (SHG) microscopy has a high specificity for non-centrosymmetric structures and was therefore used in evaluating whether deposition of collagen type I and II took place. Results: The bioink consisting of eight percent gelatin and five percent alginate, showed excellent bioprintability. Alterations in printability due to the addition of ECM fragments were resolved by adjusting the bioink components accordingly. Chondrocytes in bioprinted scaffolds displayed high cell survival, independent of ECM addition. Addition of ECM resulted in increased cell aggregation and collagen formation as shown using SHG microscopy. Conclusion: The addition of ECM fragments to 3D-printed bioink increases chondrocyte proliferation and neo-cartilage formation. Significance: The results in this study constitute a step towards bioprinted elastic cartilage for patient-specific reconstruction. Specifically, the addition of ECM to bioink increases cell proliferation and neocartilage formation necessary for the development of durable cartilage tissue.
15 mins
Improving Long-Term Culture of Cartilage Tissue in an Ex Vivo Osteochondral Model
Martina Puricelli, Edoardo Andreini, Dave Wanders, Linda M. Kock
Abstract: Ex vivo models represent an inexpensive and controllable way to explore osteochondral regeneration. We have developed an ex vivo osteochondral culture platform and using this we have shown that is possible to maintain physiological cartilage viability, matrix tissue content and structure for 56 days in culture [1]. As mechanical loading is known to be essential for the maintenance of osteochondral tissue viability and properties, the next step was to incorporate mechanical loading in our culture platform. Therefore, the aims of this study were to 1) adapt the platform such that we can apply mechanical loading to ex vivo porcine osteochondral biopsies in culture and 2) investigate the effect of a physiological mechanical loading regime on ex vivo cultured biopsies in a long-term study. We designed a platform that can simultaneously compress 6 osteochondral plugs through the vertical displacement of a linear actuator. In a custom designed graphical interface, we can set frequency, amplitude and intermittence of the stimulation. In a first study, we tested two different compression rates: 5% and 10% at 1Hz for 4 hours on, 20 hours off, during 6 weeks of culture. An unloaded group served as control. The mechanical properties of the cartilage tissue of loaded and unloaded groups were assessed through stress relaxation tests at day 0, and after 6 weeks of culture. The biological properties of the cartilage were evaluated through thionin staining for glycosaminoglycans (GAGs), quantification of GAG content and a viability (MTT) assay. We demonstrated that the designed platform can successfully stimulate osteochondral samples in a controlled, sterile and accurate manner. Preliminary results of the long-term culture showed that mechanical loading did preserve GAG content over time, while GAG content in unloaded samples decreased slightly over time. MTT staining shows that metabolic activity is preserved during the culture period similarly in all groups. In conclusion, this platform represents a valuable tool that can better preserve the osteochondral tissue viability and content during long-term culture by using relevant mechanical loading regimes. This offers opportunities for evaluating therapies and treatments for cartilage and/or bone repair in a relevant environment, mimicking the in vivo situation. This project has received funding from the European Union’s Horizon 2020 research and innovation programme under Marie Sklodowska-Curie grant agreement No 721432. References: [1] Schwab, A. et al, 2017. ALTEX - Alternatives to animal experimentation. [2] ML Vainieri, et al, 2018, Acta Biomaterialia.
15 mins
Biomechanical Functional Assessment of Hydrospacer to Serve as a Cartilage Implant
Rienk Schuiringa, René van Donkelaar, Keita Ito
Abstract: Cartilage defects occur frequently and can progress into osteoarthritis, which is painful and progressive, leading to serious functional limitations. Current treatments for defects are limited to specific age groups, lesion sites or sizes. The proposed solution is a biomimetic implant where artificial cartilage involves a HydroSpacer, existing of a knitted spacer fabric incorporated with a hydrogel that has strong swelling potential. The mechanical load-bearing function of the artificial cartilage originates from the restriction of the swelling by the spacer fabric fibres. The goal of this study is to develop an implant construct, which possess the mechanical properties of native cartilage. The swelling potential of the hydrogel is believed to have a strong influence on the mechanical properties of the HydroSpacer. Preliminary research involved HydroSpacers of poly 2-hydroxyethyl methacrylate (pHEMA) hydrogel synthesized with three different compositions and a poly lactic acid (PLA) spacer fabric. The hydrogel incorporated in the spacer fabric was polymerized using UV light and placed in PBS overnight to equilibrate in swollen state. The HydroSpacers and native porcine tibial plateau cartilage were mechanically tested using a Zwick tensile tester. Creep tests were performed using a 2 mm spherical-tip indenter. Loads of 10N, 25N and 0N were subsequently applied for 600 seconds, during which the indentation depth was monitored. Results demonstrate that the HydroSpacer stiffness is in the correct range (Table 1), while the time-dependent response of the HydroSpacer is quicker than that of cartilage. Load HydroSpacer (n=1) Cartilage (n=1) Maximum strain 10N 45%-55% 65% Maximum strain 25N 65%-83% 87% Residual strain 10N 9% 37% Residual strain 25N 12%-15% 51% Concluding, the first preliminary results are promising. Both the time-dependent and equilibrium responses of the HydroSpacer are within reasonable range from those of cartilage. This makes it likely that the properties of the construct can be sufficiently optimized by tuning the properties of either the hydrogel or the spacer fabric properties. Future developments will further characterize the constructs under a variety of mechanical conditions, optimize the construct to mimic the properties of cartilage, and use cell-seeded chondroitin sulfate methacrylate/hyaluronic acid methacrylate (CSMA/HAMA) hydrogels.
15 mins
Friction Characteristics of a Lubricated Implant for Articular Cartilage Defects
Alicia Damen, René van Donkelaar, Keita Ito
Abstract: Cartilage defects often progress into painful osteoarthritis with decreased range of motion and reduced mobility. Current treatments for cartilage defects depend on patient age and are limited to specific lesion sites or sizes. Especially for the middle-aged patient, no satisfying solutions exist. The MimCart InScite/RegMed project aims to develop a functional universal cartilage-resurfacing biomaterial. This implant involves a hydrogel with a strong swelling potential which is restricted by a knitted spacer fabric, thus mimicking the natural behaviour of respectively the proteoglycans and collagen type II in cartilage. Part of this project aims to minimize friction between implant and opposing cartilage, to prevent wear. The present study assesses the friction between the novel constructs and cartilage, and compares this to friction between cartilage and either cartilage or commercially available metal implants as controls. The coefficient of friction (CoF) was measured with a DHR-3 rheometer, expanded with a ring-on-disk accessory. Full thickness cartilage rings and disks were harvested from bovine patellae. Polylactic acid spacer fabric disks were filled with swelling pHEMA hydrogel. A cartilage ring rotated against a disk of either cartilage or implant material, submerged in PBS, FBS or synovial fluid (SF) at 25°C. Normal load was kept constant at 0.1 MPa and the angular velocity increased from 0.001 rad/s to 10 rad/s. CoF of cartilage-on-cartilage was highest in PBS and lowest in SF, agreeing with previous studies. Furthermore, the filled spacer fabric exhibited a CoF against cartilage which was 50 percent lower than CoCrMo against cartilage. Concluding, first measurements of the CoF using an in vitro cartilage-on-cartilage set-up were performed and the data look promising for further development of the implant. Upcoming, options for optimizing the implant surface using durable lubricating coatings will be explored, and the efficacy of such coatings will be demonstrated using the above presented approach.