Intraosseous Transcutaneous Amputation Prosthesis (ITAP) is a new generation of limb replacements that can provide to amputees, an alternative solution to the main problems caused by the most common used external prosthesis such as pressure sores, infections and unnatural gait. ITAP is designed as one pylon osteointegrated into the bone and protruding through the skin, allowing both the mechanical forces to be directly transferred to the skeleton and the external skin being free from frictions and infections. The skin attachment to the implant is fundamental for the success of the ITAP, as it prevents the implant to move and consequently fail. In this study we wanted to test if cell viability and attachment was improved using TiO2 nanotubes. Human keratinocytes and human dermal fibroblasts were seeded for three days on TiO2 nanotubes with different sizes (18–30nm, 40–60nm and 60–110nm), compared with controls (smooth titanium) and tested for viability and attachment. A Mann-Whitney U test was used to compare groups where p values < 0.05 were considered significant. The results showed that the viability and cell attachment for keratinocytes were significantly higher after three days on controls comparing with all nanotubes (p=0.02), while attachment was higher on bigger nanotubes and controls. Cell viability for fibroblasts was significantly higher on nanotubes between 40 and 110nm comparing with smaller size and controls (p=0.03), while investigation of cell attachment is ongoing. From these early results, we can say that TiO2 nanotubes can improve the soft tissue attachment on ITAP. Further in-vitro and ex-vivo experiments on cell attachment will be carried out.
Re-attachment of tendon to bone is challenging with surgical repair failing in up to 90% of cases. Poor biological healing is common and characterised by the formation of weak scar tissue. Previous work has demonstrated that decellularised allogenic demineralised bone matrix (DBM) regenerates a physiologic enthesis. Xenografts offer a more cost-effective option but concerns over their immunogenicity have been raised. We hypothesised that augmentation of a healing tendon-bone interface with DBM incorporated with autologous mesenchymal stem cells (MSCs) would result in improved function, and restoration of the native enthesis, with no difference between xenogenic and allogenic scaffolds. Using an ovine model of tendon-bone retraction the patellar tendon was detached and a complete distal tendon defect measuring 1 cm was created. Suture anchors were used to reattach the shortened tendon and xenogenic DBM + MSCs (n=5) and allogenic DBM + MSCs (n=5) were used to bridge the defect. Functional recovery was assessed every 3 weeks and DBM incorporation into the tendon and its effect on enthesis regeneration was measured using histomorphometry.Background
Methods
Our results prove that Demineralised Cortical Bone (DCB) can be used as biological tendon graft substitute, combined with correct surgical technique and the use of suture bone anchor early mobilisation can be achieved. Surgical repair of tendon injuries aims to restore length, mechanical strength and function. In severe injuries with loss of tendon substance a tendon graft or a substitute is usually used to restore functional length. This is usually associated with donor site morbidity, host tissue reactions and lack of remodelling of the synthetic substitutes which may result in suboptimal outcome. In this study we hypothesise that DCB present in biological tendon environment with early mobilisation and appropriate tension will result in remodelling of the DCB into ligament tissue rather that ossification of the DCB at traditional expected. Our preparatory cadaveric study (abstract submitted to CORS 2013) showed that the repair model used in this animal study has sufficient mechanical strength needed for this animal study.Summary
Introduction
Our study shows that a tendon rupture can be successfully augmented with Demineralised Cortical Bone (DCB) giving initial appropriate mechanical strength suitable for in vivo use providing the biological reactions to the graft are favourable. Treatment of tendon and ligament injuries remains challenging; the aim is to find a biocompatible substance with mechanical and structural properties that replicate those of normal tendon and ligament. Because of its structural and mechanical properties, we proposed that DCB can be used in repair of tendon and ligament as well as regeneration of the enthesis. DCB is porous, biocompatible and has the potential to be remodelled by the host tissues. 2 studies were designed; in the first we examined the mechanical properties of DCB after gamma irradiation (GI) and freeze drying (FD). In the second we used different techniques for repairing bone-tendon-bone with DCB in order to measure the mechanical performance of the construct.Summary
Introduction
Osseointegrated Amputation Prostheses can be functionalised by both biological augmentation and structural augmentation. These augmentation techniques may aid the formation of a stable skin-implant interface. Current clinical options are limited in restoring function to amputees, and are associated with contact dermatitis and infection at the stump-socket interface. Osseointegrated Amputation Prosthesis attempts to solve issues at the stump-socket interface by directly transferring axial load to the prosthesis, via a skin-penetrating abutment. However, development is needed to achieve a seal at the skin-implant interface to limit infection. Fibronectin, an Extracellular Matrix protein, binds to integrins during wound healing, with the RGD tripeptide being part of the recognition sequence for its integrin binding domain. Summary
Introduction
Bone-anchored devices have been used as skin-crossing conduits to record neuromuscular signals in sedated animals. Long-term recordings from cognisant subjects must be assessed. Hypothesis A bone-anchored device is suitable as a conduit for epimysial EMG (Electromyogram) recordings and is reliable in the long-term. The bone-anchored device was implanted into the medial aspect of an ovine tibia (n=1), and the epimysial electrode was sutured onto the peroneus tertius muscle. Epimysial and Surface EMG signals were recorded for 12 weeks.Introduction
Methods
Repair of tendon injuries aims to restore length, mechanical strength and function. We hypothesise that Demineralised Cortical Bone (DCB) present in biological tendon environment will result in remodelling of the DCB into ligament tissue. A cadaveric study was carried out to optimize the technique. The distal 1cm of the patellar tendon was excised and DCB was used to bridge the defect. 4 models were examined, Model-1: one anchor, Model-2: 2 anchors, Model-3: 2 anchors with double looped off-loading thread, Model-4: 2 anchors with 3 threads off-loading loop. 6 mature sheep undergone surgical resection of the distal 1cm of the right patellar tendon. Repair was done using DCB with 2 anchors. Immediate mobilisation was allowed, animals were sacrificed at 12 weeks. Force plate assessments were done at weeks 3, 6, 9 and 12. Radiographs were taken and pQCT scan was done prior to histological analysis. In the cadaveric study, the median failure force for the 4 models; 250N, 290N, 767N and 934N respectively. In the animal study, none of the specimens showed evidence of ossification of the DCB. One animal failed to show satisfactory progress, X-rays showed patella alta, on specimen retrieval there was no damage to the DCB and sutures and no evidence of anchor pullout. Functional weight bearing was 79% at week12. Histological analysis proved remodelling of the collagen leading to ligamentisation of the DCB. Results prove that DCB can be used as biological tendon substitute, combined with the use of suture bone anchor early mobilisation can be achieved.
Treatment of tendon and ligament injuries remains challenging; the aim is to find a biocompatible substance with mechanical and structural properties that replicate those of normal tendon and ligament. We examined the mechanical properties of Demineralised Cortical Bone (DCB) after gamma irradiation (GI) and freeze drying (FD). We also used different techniques for repairing bone-tendon-bone with DCB in order to measure the mechanical performance of the construct. DCB specimens were allocated into 4 groups; FD, GI, combination of both or none. The maximum tensile forces and stresses were measured. 4 cadaveric models of repair of 1cm patellar tendon defect using DCB were designed; model-1 using one bone anchor, Model-2 using 2 bone anchors, Model-3 off-loading by continuous thread looped twice through bony tunnels, Model-4 off-loading with 3 hand braided threads. Force to failure and mode were recorded for each sample. FD groups results were statistically higher (p=<0.05) compared to non-FD groups, while there was no statistical difference between GI and non-GI groups. The median failure force for model-1: 250N, model-2: 290N, model-3: 767N and model-4: 934N. There was no statistical significance between model-1 and model-2 (p=0.249), however statistical significance was found between other models (p=<0.006). GI has no significant effect on mechanical strength of the CDB while FD may have positive effect on its mechanical strength. Our study shows that a tendon rupture can be successfully augmented with CDB giving initial appropriate mechanical strength suitable for in vivo use providing the biological reactions to the graft are favourable.
Infection is the primary failure modality for transcutaneous implants because the skin breach provides a route for pathogens to enter the body. Intraosseous transcutaneous amputation prostheses (ITAP) are being developed to overcome this problem by creating a seal at the skin-implant interface to prevent bacterial invasion. Oral gingival epithelial cell adhesion creates an infection free seal around dental implants; however this has yet to be demonstrated outside the oral environment. All epithelial cells attach via hemidesmosomes (HD) and focal adhesions (FA) and their expression is an indicator of adhesion efficiency. The aim of this study was to compare epidermal keratinocyte with oral gingival epithelial cell adhesion on titanium alloy in vitro to determine whether these two cell types differ in their speed and strength of adhesion. It was hypothesised that oral gingival epithelial cells attach to titanium alloy earlier than epidermal keratinocytes; with greater expression of hemidesmosomes and focal adhesions. Human oral gingival epithelial cell (HGEP) and primary human epidermal keratinocyte (HPEK) adhesion to titanium alloy, was assessed at 4, 24, 48 and 72 hrs. Adhesion was measured by the number of FAs per unit cell area and expression of HDs using a semi-quantitative scale. At 4 and 24hrs, there was a significant increase in vinculin marker expression per unit cell area of 4.3 and 4.7 times in HGEP compared with HPEK (p=0.000). At 48 and 72hrs there were no significant differences. HD expression was significantly greater in HGEP at 4 and 24hrs (p=0.002) compared with HPEK. Up-regulation of HD expression in HPEK lagged that of HGEP until 48hrs, after which no significant differences were observed. This study has demonstrated that oral gingival cells up-regulate both focal adhesion and hemidesmosome expression at earlier time points compared with epidermal keratinocytes. Expression of hemidesmosomes lags that of focal adhesions, suggesting that focal adhesion formation is a prerequisite for hemidesmosome assembly. We postulate that early attachment of oral gingival epithelial cells to dental implant biomaterials may be responsible for the formation of an infection-free seal.
