In 2021 the bone grafting market was worth €2.72 billion globally. As allograft bone has a limited supply and risk of disease transmission, the demand for synthetic grafting substitutes (BGS) continues to grow while allograft bone grafts steadily decrease. Synthetic BGS are low in mechanical strength and bioactivity, inspiring the development of novel grafting materials, a traditionally laborious and expensive process. Here a novel BGS derived from sustainably grown coral was evaluated. Coral-derived scaffolds are a natural calcium carbonate bio-ceramic, which induces osteogenesis in bone marrow mesenchymal stem cells (MSCs), the cells responsible for maintaining bone homeostasis and orchestrating fracture repair. By 3D printing MSCs in coral-laden bioinks we utilise high throughput (HT) fabrication and evaluation of osteogenesis, overcoming the limitations of traditional screening methods.

MSC and coral-laden GelXA (CELLINK) bioinks were 3D printed in square bottom 96 well plates using a CELLINK BIO X printer with pneumatic adapter Samples were non-destructively monitored during the culture period, evaluating both the sample and the culture media for metabolism (PrestoBlue), cytotoxicity (lactose dehydrogenase (LDH)) and osteogenic differentiation (alkaline phosphatase (ALP)). Endpoint, destructive assays used included qRT-PCR and SEM imaging.

The inclusion of coral in the printed bioink was biocompatable with the MSCs, as reflected by maintained metabolism and low LDH release. The inclusion of coral induced osteogenic differentiation in the MSCs as seen by ALP secretion and increased RUNX2, collagen I and osteocalcin transcription.

Sustainably grown coral was successfully incorporated into bioinks, reproducibly 3D printed, non-destructively monitored throughout culture and induced osteogenic differentiation in MSCs. This HT fabrication and monitoring workflow offers a faster, less labour-intensive system for the translation of bone substitute materials to clinic.

Acknowledgements: This work was co-funded by Enterprise Ireland and Zoan Biomed through Innovation Partnership IP20221024.

Recent studies have shown that bone mineral distribution is more heterogeneous in bone tissue from an animal model of osteoporosis and osteoporotic human vertebral trabeculae. These tissue alterations may play a role in bone fragility seen in osteoporosis, albeit that they are not detectable by current diagnostic techniques (dual-energy X-ray absorptiometry, DXA). Type II Diabetes Mellitus (T2DM) also increases a patient's fracture risk beyond what can be explained or diagnosed by DXA, and is associated with impaired bone cell function, compromised collagen structure and reduced mechanical properties. However, it is not currently known whether osteoporosis or T2DM leads to an increased mineral heterogeneity in the femoral head of humans, a common osteoporotic fracture site. In this study, we examine bone microarchitecture, mineralisation and mechanical properties of trabecular bone from osteoarthritic, diabetic and osteoporotic patients. We report that while osteoporotic trabecular bone has significantly deteriorated mechanical properties and microarchitecture compared to the other groups, there is also a significant increase in mean mineral content. Moreover, the heterogeneity of the mineral content in osteoporotic bone is significantly higher than osteoarthritic (+35%) and diabetic (+13%) groups. We propose that the compromised architecture following bone loss at the onset of osteoporosis alters the mechanical environment, which initiates compensatory changes in mineral content. We show for the first time that trabecular bone mineralisation is significantly more heterogeneous (+20%) in T2DM compared to osteoarthritic controls. Interestingly, bone microarchitecture and mechanical properties are not significantly different between diabetic and osteoarthritic groups despite this increase in mineral heterogeneity.

The international literature base demonstrates that individuals living with diabetes mellitus (DM) are at increased risk of mortality and post-operative complications following hip fracture surgery (HFS) than non-diabetics. Studies investigating databases in American, European or Asiatic populations highlight the impact geography can have on the resultant investigation. We aim to quantify the impact DM has on HFS patients in a single university hospital. The HIPE dataset of fragility fractures occurring in Galway University Hospital from 2014–2016 were analysed and cross referenced with hospital laboratory and public databases. A database of 759 individuals was created including 515 females and 237 males, with a mean age of 78+/−12.2 years, of which 110 patients had DM. The patient length-of-stay (PLOS) was comparable in all groups with patient age being the primary influencing factor. An extended PLOS correlated with an increased long-term mortality. A trend toward increased occurrence of sub-trochanteric fractures was observed in diabetics with fewer periprosthetic and intertrochanteric fractures. Patients with DM had a significant increased risk of post-operative mortality compared to non-diabetics. Males with DM where at a greater risk of death after HFS [HR 2.29, 95% CI 1.26–4.17. p=0.006] than females with DM [HR 1.69, 95% CI 0.99–2.91. p=0.056]. The presence of DM did not directly impact a patient's PLOS or increase the need for a re-operation. DM is associated with increased post-operative patient mortality and may influence the anatomical fracture pattern. This observation will support further investigation into the mechanical and biochemical changes occurring in the femur in individuals living with DM.

Clinical translation of MSC therapies in orthopaedics has been hampered by heterogeneity and a lack of standardised and validated testing protocols for quality assurance. Although minimal criteria have been proposed¹, it is apparent that these do not predict performance in vivo. We used a combinatorial antibody profiling tool to probe the surface immunophenotype of human bone marrow derived MSCs and used this to define new marker panels. Cells were cultured from three marrow donors using specified expansion conditions and probed by high throughput flow cytometry using a panel of 230 antibodies. Analysis of expression of the surface proteins revealed significant variation in response to culture conditions and considerably less variation between donors. Of the panel of 230 markers 107 were negative, 24 had high expression in all samples, 1 had low expression and 98 displayed significant differences between cell preparations. Cluster analyses revealed that marker expression in one culture condition varied considerably from the other two. Phenotypic characterization of the cell preparations, assessed by analysis of differentiation propensity, showed similar patterns of variability between these samples. This suggests that the selected panel may be used as phenotypic MSC markers. Ongoing work involves the generation of novel antibody arrays which will be used as quality tests in a manufacturing environment. These tests will be used for in-process and product release applications for enhanced cell manufacturing and improved clinical outcomes.

Introduction and Aims: This study examined the feasibility and clinical response of treatment with the Cyberknife Stereotactic Radiosurgery system of patients with spine tumors not amenable to other types of treatment. These included patients with metastases recurrent after previous radiation, those resistant to radiation or those requiring extensive surgery for complete resection.

Method: Twenty-nine patients with tumors of the spine were treated with hypofractionated (one to four fractions) high dose radiation (CyberKnife), delivered using implanted fiducial markers for precise stereotactic localisaton. Patients had either recurrent spinal metastases (19), radio-resistant metastases (seven) or small lesions requiring extensive resection (three). After four to six fiducials were implanted, the patient was immobilised in a custom-moulded cradle and a CT scan was obtained with up to 300 slices at 1.25mm intervals. Inverse plannning was done to minimise dose to critical structures in close proximity to the tumor mass. Patients were followed-up with clinical pain scores, total pain medication, functional assessment and follow-up CT and/or MRI at three-month intervals to assess response to treatment.

Results: The tumors were located in all areas of the spine from C4 to the sacrum, with renal cell carcinoma being the most common diagnosis. The mean tumor volume was 253.4cc, with a range of 0.33 to 678.9 ccs. The maximum radiation dose prescribed to the tumor ranged from 1600cGy to 2500cGy delivered in one to four fractions. The number of fractions was determined by the tumor volume and whether the spinal lesion had been previously treated with radiation. The maximum allowable dose to the adjacent spinal cord was 800cGy and thus for the majority of the tumors prescribed to 2500cGy, 80% of the tumor volume received at least 2000cGy. Patients were treated in an outpatient setting with an average treatment time of 75 minutes. There were no new neurologic deficits or acute radiation toxicity. Patients with lesions in the lumbar spine or sacrum often experienced a brief period of nausea, which was easily controllable with one dose of anti-emetic. Some patients experienced a period of malaise or lethargy with no predictive factors. Pain was markedly improved in all patients with metastatic disease as demonstrated by pain scores, decreased use of narcotic medications and improved function. Repeat radiographic studies at three months generally demonstrated stable tumor volume, while those at six months showed decrease in tumor size.

Conclusion: Stereotactic radiosurgery has distinct advantages over external beam for patients with tumors of the spine, including less toxicity, ability to treat recurrences in previously radiated fields, and shorter treatment durations. While GammaKnife for cranial lesions is a widely accepted technique by neurosurgeons, the use of frameless stereotactic radiosurgery with the Cyberknife is new to the armamentarium of orthopaedic surgeons treating spinal tumors.