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Rationale: Elastin‐like polypeptides (ELPs) are elastic and thermoresponsive biopolymers composed of VPGXG repeats (X can be any amino acid except proline), used in biomedical applications, for example, tissue engineering and drug delivery. As different variants of ELP are mostly produced fermentatively, there is a need for the development of analysis methods that allow for absolute protein quantification in both complex matrices and purified samples and MW determination of the final products. Methods: ELPs were intracellularly expressed in Escherichia coli quantified after cell lysis and enzymatic digestion using a proline‐specific protease ProAlanase (Promega) at acidic conditions. Resulting peptides were separated by liquid chromatography, and mass spectrometry analysis was conducted by electrospray ionization high‐resolution mass spectrometry using an Orbitrap mass spectrometer. The addition of a stable isotopically labeled internal standard enabled quantification in complex matrices. Prior to intact mass analysis, ELPs were purified from fermentation broth by inverse temperature cycling. Intact protein analysis was performed using reversed‐phase liquid chromatography, and mass spectrometry analysis was conducted by electrospray ionization high‐resolution mass spectrometry using a time‐of‐flight mass spectrometer. Results: Absolute quantification of ELPs was achieved by utilizing ELP‐specific properties, that is, proline‐rich, soluble at low pH and low temperature. The repetitive nature of ELPs allows for sensitivity increase and use of higher dilution factors to minimize the matrix effects. Despite the lack of amino acids with charged side chains (Arg, His, Lys, Asp, and Glu) in ELP, we demonstrated successful intact protein analysis using reversed‐phase LC coupled to electrospray ionization TOF MS. Moreover, truncated protein forms could be chromatographically separated and characterized as well as N‐terminal modifications. Conclusions: Both methods combined enabled quantitative and qualitative characterization of fermentatively produced ELPs. [ABSTRACT FROM AUTHOR]

Currently available focal knee resurfacing implants (FKRIs) are fully or partially composed of metals, which show a large disparity in elastic modulus relative to bone and cartilage tissue. Although titanium is known for its excellent osseointegration, the application in FKRIs can lead to potential stress‐shielding and metal implants can cause degeneration of the opposing articulating cartilage due to the high resulting contact stresses. Furthermore, metal implants do not allow for follow‐up using magnetic resonance imaging (MRI).To overcome the drawbacks of using metal based FKRIs, a biomimetic and MRI compatible bi‐layered non‐resorbable thermoplastic polycarbonate‐urethane (PCU)‐based FKRI was developed. The objective of this preclinical study was to evaluate the mechanical properties, biocompatibility and osteoconduction of a novel Bionate® 75D ‐ zirconium oxide (B75D‐ZrO2) composite material in vitro and the osseointegration of a B75D‐ZrO2 composite stem PCU implant in a caprine animal model. The tensile strength and elastic modulus of the B75D‐ZrO2 composite were characterized through in vitro mechanical tests under ambient and physiological conditions. In vitro biocompatibility and osteoconductivity were evaluated by exposing human mesenchymal stem cells to the B75D‐ZrO2 composite and culturing the cells under osteogenic conditions. Cell activity and mineralization were assessed and compared to Bionate® 75D (B75D) and titanium disks. The in vivo osseointegration of implants containing a B75D‐ZrO2 stem was compared to implants with a B75D stem and titanium stem in a caprine large animal model. After a follow‐up of 6 months, bone histomorphometry was performed to assess the bone‐to‐implant contact area (BIC). Mechanical testing showed that the B75D‐ZrO2 composite material possesses an elastic modulus in the range of the elastic modulus reported for trabecular bone. The B75D‐ZrO2 composite material facilitated cell mediated mineralization to a comparable extent as titanium. A significantly higher bone‐to‐implant contact (BIC) score was observed in the B75D‐ZrO2 implants compared to the B75D implants. The BIC of B75D‐ZrO2 implants was not significantly different compared to titanium implants. A biocompatible B75D‐ZrO2 composite approximating the elastic modulus of trabecular bone was developed by compounding B75D with zirconium oxide. In vivo evaluation showed an significant increase of osseointegration for B75D‐ZrO2 composite stem implants compared to B75D polymer stem PCU implants. The osseointegration of B75D‐ZrO2 composite stem PCU implants was not significantly different in comparison to analogous titanium stem metal implants. [ABSTRACT FROM AUTHOR]

The clinical success of osteochondral implants depends significantly on their surface properties. In vivo, an implant may roughen over time which can decrease its performance. The present study investigates whether changes in the surface texture of metal and two types of polycarbonate urethane (PCU) focal knee resurfacing implants (FKRIs) occurred after 6 and 12 months of in vivo articulation with native goat cartilage. PCU implants which differed in stem stiffness were compared to investigate whether the stem fixating the implant in the bone influences surface topography. Using optical profilometry, 19 surface texture parameters were evaluated, including spatial distribution and functional parameters obtained from the material ratio curve. For metal implants, wear during in vivo articulation occurred mainly via material removal, as shown by the significant decrease of the core‐valley transition from 91.5% in unused implants to 90% and 89.6% after 6 and 12 months, respectively. Conversely, for PCU implants, the wear mechanism consisted in either filling of the valleys or flattening of the surface by dulling of sharp peaks. This was illustrated in the change in roughness skewness from negative to positive values over 12 months of in vivo articulation. Implants with a softer stem experienced the most deformation, shown by the largest change in material ratio curve parameters. We therefore showed, using a detailed surface profilometry analysis, that the surface texture of metal and two different PCU FKRIs changes in a different way after articulation against cartilage, revealing distinct wear mechanisms of different implant materials. [ABSTRACT FROM AUTHOR]

Focal knee resurfacing implants (FKRIs) are intended to treat cartilage defects in middle‐aged patients. Most FKRIs are metal‐based, which hampers follow‐up of the joint using magnetic resonance imaging and potentially leads to damage of the opposing cartilage. The purpose of this study was to develop a nondegradable thermoplastic polyurethane (TPU) FKRI and investigate its osseointegration. Different surface roughness modifications and biphasic calcium phosphate (BCP) coating densities were first tested in vitro on TPU discs. The in vivo osseointegration of BCP‐coated TPU implants was subsequently compared to uncoated TPU implants and the titanium bottom layer of metal control implants in a caprine model. Implants were implanted bilaterally in stifle joints and animals were followed for 12 weeks, after which the bone‐to‐implant contact area (BIC) was assessed. Additionally, 18F‐sodium‐fluoride (18F‐NaF) positron emission tomography PET/CT‐scans were obtained at 3 and 12 weeks to visualize the bone metabolism over time. The BIC was significantly higher for the BCP‐coated TPU implants compared to the uncoated TPU implants (p =.03), and did not significantly differ from titanium (p =.68). Similar 18F‐NaF tracer uptake patterns were observed between 3 and 12 weeks for the BCP‐coated TPU and titanium implants, but not for the uncoated implants. TPU FKRIs with surface modifications could provide the answer to the drawbacks of metal FKRIs. [ABSTRACT FROM AUTHOR]

Background and Purpose: Corticosteroids are intra-articularly injected to relieve pain in joints with osteoarthritis (OA) or acute tissue damage such as ligament or tendon tears, despite its unverified contraindication in unstable joints. Biomaterial-based sustained delivery may prolong reduction of inflammatory pain, while avoiding harmful peak drug concentrations.Experimental Approach: The applicability of prolonged corticosteroid exposure was examined in a rat model of anterior cruciate ligament and medial meniscus transection (ACLT + pMMx) with ensuing degenerative changes.Key Results: Intra-articular injection of a bolus of the corticosteroid triamcinolone acetonide (TAA) resulted in enhanced joint instability in 50% of the joints, but neither instability-induced OA cartilage degeneration, synovitis, nor the OA-related bone phenotype was affected. However, biomaterial microsphere-based extended TAA release enhanced instability in 94% of the animals and induced dystrophic calcification and exacerbation of cartilage degeneration. In healthy joints, injection with TAA releasing microspheres had no effect at all. In vitro, TAA inhibited cell migration out of joint tissue explants, suggesting inhibited tissue healing in vivo as mechanisms for enhanced instability and subsequent cartilage degeneration.Conclusions and Implications: We conclude that short-term TAA exposure has minor effects on surgically induced unstable joints, but its extended presence is detrimental by extending instability and associated joint degeneration through compromised healing. This supports a contraindication of prolonged corticosteroid exposure in tissue damage-associated joint instability, but not of brief exposure. [ABSTRACT FROM AUTHOR]