Treatment of cortical bone defects is a clinical challenge. Guided bone regeneration (GBR), commonly used in oral in maxillofacial dental surgery, may show promise for orthopedic application in repair of cortical defects. However, a limitation in the use of GBR for cortical bone defects is the lack of an ideal scaffold that provides sufficient mechanical support to bridge the cortical bone with minimal interference in the repair process. We have developed a new collagen membrane, CelGro™, for use in GBR. We report the material characterisation of CelGro™, and evaluate the performance of CelGro™ in translational preclinical and clinical studies. The results show CelGro™ has a bilayer structure of different fibre alignment and is composed almost exclusively of type I collagen. CelGro™ was found to be completely acellular and free from xenoantigen, α-gal. In the preclinical study of a rabbit cortical bone defect model, CelGro™ demonstrated enhanced bone-modelling activity and cortical bone healing. Micro-CT evaluation showed early bony bridging over the defect area 30 days post-operatively, and nearly complete restoration of mature cortical bone at the bone defect site 60 days post-operatively. Histological analysis at day 60 after surgery further confirmed that CelGro™ enables bridging of the cortical bone defect by induction of newly-formed cortical bone. Compared to a commercially available collagen membrane, Bio-Gide®, CelGro™ showed much better cortical alignment and reduced porosity at the defect interface. Owning the fact that selection of orthopedic patients with cortical bone defects is complex, we conducted a clinical study in a total of 16 dental implants which were placed in 10 participants receiving GBR. The results showed that Celgro™ restore bone defect with no complications or adverse events. Cone-beam computed tomography (CBCT) images show significantly increased bone formation in both horizontally and vertically, which provids sufficient support to the implants within 4 months. Together, the the findings of our study demonstrate that CelGro™ is an ideal membrane for GBR not only in oral maxillofacial reconstructive surgery but also in orthopedic applications.

There is a high risk of injury from damage to the force-bearing tissue of the tendon. Due to its poor self-healing ability, clinical interventions for tendon injuries are limited and yield unsatisfying results. Tissue engineering might supply an alternative to this obstacle. As one of the key elements of tissue engineering, various cell sources have been used for tendon engineering, but there is no consensue concerning a single optimal source. In this review, we summarized the development of tendon tissue from the embryonic stage and categorized the used cell sources in tendon engineering. By comparing various cell sources as the candidates for tendon regeneration, each cell type was found to have its advantages and limitations; therefore, it is difficult to define the best cell source for tendon engineering. The microenvironment cells located is also crucial for cell growth and differentiation; so, the optimal cells are unlikely to be the same for each patient. In the future, the clinical application of tendon engineering might be more precise and customized in contrast to the current use of a standardized/generic one-size-fits-all procedure. The best cell source for tendon engineering will require a case-based assessment.

Tendons are the force transferring tissue that enable joint movement. Excessive mechanical loading is commonly considered as a primary factor causing tendinopathy, however, an increasing body of evidence supports the hypothesis that overloading creates microdamage of collagen fibers resulting in a localized decreased loading on the cell population within the damaged site. Heterotopic ossification is a complication of late stage tendinopathy, which can significantly affect the mechanical properties and homeostasis of the tendon. Here, we the examine the effect of mechanical underloading on tendon ossification and investigate its underlying molecular mechanism. Rabbit Achilles tendons were dissected and cultured in an underloading environment (3% cyclic tensile stain,0.25 ​Hz, 8 ​h/day) for either 10, 15 or 20 days. Using isolated tendon-derived stem cells (TDSCs) 3D constructs were generated, cultured and subjected to an underloading environment for 6 days. Histological assessments were performed to evaluate the structure of the 3D constructs; qPCR and immunohistochemistry were employed to study TDSC differentiation and the β-catenin signal pathway was investigated by Western blotting. Mechanical testing was used to determine ability of the tendon to withstand force generation. Tendons cultured for extended times in an environment of underloading showed progressive heterotopic ossification and a reduction in biomechanical strength. qPCR revealed that 3D TDSCs constructs cultured in an underloading environment exhibited increased expression of several osteogenic genes: these include RUNX2, ALP and osteocalcin in comparison to tenogenic differentiation markers (scleraxis and tenomodulin). Immunohistochemical analysis further confirmed high osteocalcin production in 3D TDSCs constructs subject to underloading. Western blotting of TDSC constructs revealed that β-catenin accumulation and translocation were associated with an increase in phosphorylation at Ser552 and decrease phosphorylation at Ser33. These findings unveil a potential mechanism for heterotopic ossification in tendinopathy due to the underloading of TDSCs at the damage sites, and also that β-catenin could be a potential target for treating heterotopic ossification in tendons. Tendon heterotopic ossification detrimentally affect quality of life especially for those who has atheletic career. This study reveals the possible mechanism of heterotpic ossification in tendon related to mechanical loading. This study provided the possible to develop a mechanical stimulation protocol for preventive and therapeutic purpose for tendon heterotopic ossification.

Background: Botulinum toxin (Botox) injection is in widespread clinical use for the treatment of muscle spasms and tendinopathy but the mechanism of action is poorly understood.Hypothesis: We hypothesised that the reduction of patellar-tendon mechanical-loading following intra-muscular injection of Botox results in tendon atrophy that is at least in part mediated by the induction of senescence of tendon-derived stem cells (TDSCs).Study design: Controlled laboratory studyMethods: A total of 36 mice were randomly divided into 2 groups (18 Botox-injected and 18 vehicle-only control). Mice were injected into the rightvastus lateralisof quadriceps muscles either with Botox (to induce mechanical stress deprivation of the patellar tendon) or with normal saline as a control. At 2 weeks post-injection, animals were euthanized prior to tissues being harvested for either evaluation of tendon morphology or in vitro studies. TDSCs were isolated by cell-sorting prior to determination of viability, differentiation capacity or the presence of senescence markers, as well as assessing their response to mechanical loading in a bioreactor. Finally, to examine the mechanism of tendon atrophy in vitro, the PTEN/AKT-mediated cell senescence pathway was evaluated in TDSCs from both groups.Results: Two weeks after Botox injection, patellar tendons displayed several atrophic features including tissue volume reduction, collagen fibre misalignment and increased degradation. A colony formation assay revealed a significantly reduced number of colony forming units of TDSCs in the Botox-injected group compared to controls. Multipotent differentiation capacities of TDSCs were also diminished after Botox injection. To examine if mechanically deprived TDSC are capable of forming tendon tissue, we used an isolated bioreactor system to culture tendon constructs using TDSC. These results showed that TDSCs from the Botox-treated group failed to restore tenogenic differentiation after appropriate mechanical loading. Examination of the signalling pathway revealed that injection of Botox into quadriceps muscles causes PTEN/AKT-mediated cell senescence of TDSCs.Conclusion: Intramuscular injection of Botox interferes with tendon homeostasis by inducing tendon atrophy and senescence of TDSCs. Botox injection may have long-term adverse consequences for the treatment of tendinopathy.Clinical relevance: Intramuscular Botox injection for tendinopathy or tendon injury could result in adverse effects in human tendons and evaluation of its long-term efficacy is warranted.

Tendinopathy is a common chronic tendon disease relating to inflammation and degeneration in an orthopaedic area. With high morbidity, limited self-repairing capacity and, most importantly, no definitive treatments, tendinopathy still influences patients’ life quality negatively. Tendon-derived stem cells (TDSCs), as primary precursor cells of tendon cells, play an essential role in both the development of tendinopathy, and functional and structural restoration after tendinopathy. Thus, a method that can in vitro mimic the in vivo differentiation of TDSCs into tendon cells would be useful. Here, the present protocol describes a method based on a three-dimensional (3D) uniaxial stretching system to stimulate the TDSCs to differentiate into tendon-like tissues. There are seven stages of the present protocol: isolation of mice TDSCs, culture and expansion of mice TDSCs, preparation of stimulation culture medium for cell sheet formation, cell sheet formation by culturing in stimulation medium, preparation of 3D tendon stem cell construct, assembly of the uniaxial-stretching mechanical stimulation complex, and evaluation of the mechanical stimulated in vitro tendon-like tissue. The effectiveness was demonstrated by histology. The entire procedure takes less than 3 weeks. To promote extracellular matrix deposition, 4.4 mg/mL ascorbic acid was used in the stimulation culture medium. A separated chamber with a linear motor provides accurate mechanical loading and is portable and easily adjusted, which is applied for the bioreactor. The loading regime in the present protocol was 6% strain, 0.25 Hz, 8 h, followed by 16 h rest for 6 days. This protocol could mimic cell differentiation in the tendon, which is helpful for the investigation of the pathological process of tendinopathy. Moreover, the tendon-like tissue is potentially used to promote tendon healing in tendon injury as an engineered autologous graft. To sum up, the present protocol is simple, economic, reproducible and valid.