Critical-sized diaphysis defects are complicated by inherent sub-optimal healing conditions. The two-staged induced membrane technique has been used to treat these challenging defects since the 1980's. It involves temporary implantation of a membrane-inducing spacer and subsequent bone graft defect filling. A single-staged, graft-independent technique would reduce both socio-economic costs and patient morbidity. Our aim was to enable such single-staged approach through development of a strong bioactive glass scaffold that could replace both the spacer and the graft filling. We constructed amorphous porous scaffolds of the clinically used bioactive glass S53P4 and evaluated them in vivo using a critical-sized defect model in the weight-bearing femur diaphysis of New Zealand White rabbits. S53P4 scaffolds and standard polymethylmethacrylate spacers were implanted for 2, 4, and 8 weeks. Induced membranes were confirmed histologically, and their osteostimulative activity was evaluated through RT-qPCR of bone morphogenic protein 2, 4, and 7 (BMPs). Bone formation and osseointegration were examined using histology, scanning electron microscopy, energy-dispersive X-ray analysis, and micro-computed tomography imaging. Scaffold integration, defect union and osteosynthesis were assessed manually and with X-ray projections. We demonstrated that S53P4 scaffolds induce osteostimulative membranes and produce osseointegrative new bone formation throughout the scaffolds. We also demonstrated successful stable scaffold integration with early defect union at 8 weeks postoperative in critical-sized segmental diaphyseal defects with implanted sintered amorphous S53P4 scaffolds. This study presents important considerations for future research and the potential of the S53P4 bioactive glass as a bone substitute in large diaphyseal defects. STATEMENT OF SIGNIFICANCE: Surgical management of critical-sized diaphyseal defects involves multiple challenges, and up to 10% result in delayed or non-union. The two-staged induced membrane technique is successfully used to treat these defects, but it is limited by the need of several procedures and bone graft. Repeated procedures increase costs and morbidity, while grafts are subject to donor-site complications and scarce availability. To transform this two-staged technique into one graft-independent procedure, we developed amorphous porous scaffolds sintered from the clinically used bioactive glass S53P4. This work constitutes the first evaluation of such scaffolds in vivo in a critical-sized diaphyseal defect in the weight-bearing rabbit femur. We provide important knowledge and prospects for future development of sintered S53P4 scaffolds as a bone substitute.

Although the initial in vitro dissolution of bioactive glasses (BAG) is well characterized, the long-term behaviour of crystallized BAG scaffolds in a continuous fluid flow is incompletely understood. A detailed understanding of the long-term dissolution of scaffolds is vital for predicting their behaviour in clinical applications. Here, we explored the dissolution and reaction mechanisms of partly crystalline and glass–ceramic scaffolds based on the bioactive glasses S53P4 and 45S5 using a continuous flow-through method in Tris-buffer (Tris) and simulated body fluid (SBF) for up to 21 days. Granules of the parent glasses were used as references. The main crystalline phase in both scaffolds was sodium-calcium-silicate. The scaffolds’ dissolution suggested that the sodium-calcium-silicate crystals dissolved incongruently to yield hydrous silica. The silica phase then provided abundant nucleation sites for hydroxyapatite precipitation, resulting in fine-grained crystalline structures. When exposed to Tris, the scaffolds almost completely dissolved within the test period, leaving only highly porous remnant phases. For the 45S5 scaffolds, the calcium phosphate reaction layers that formed on their surfaces effectively slowed the dissolution in SBF. In contrast, this effect was less apparent for the S53P4 specimens.

The silicate-based bioactive glass S53P4 is clinically used in bone regenerative applications in granule form. However, utilization of the glass in scaffold form has been limited by the high tendency of the glass to crystallize during sintering. Here, careful optimization of sintering parameters enabled the manufacture of porous amorphous S53P4 scaffolds with a strength high enough for surgical procedures in bone applications (5 MPa). Sintering was conducted in a laboratory furnace for times ranging from 25 to 300 min at 630 °C, i.e., narrowly below the commencement of the crystallization. The phase composition of the scaffolds was verified with XRD, and the ion release was tested in vitro and compared with granules in continuous flow of Tris buffer and simulated body fluid (SBF). The amorphous, porous S53P4 scaffolds present the possibility of using the glass composition in a wider range of applications.

Fabrication of porous tissue-engineering scaffolds from bioactive glasses (BAG) is complicated by the tendency of BAG compositions to crystallize in thermal treatments during scaffold manufacture. Here, experimental biocompatible glass S59 (SiO₂ 59.7 wt%, Na₂O 25.5 wt%, CaO 11.0 wt%, P₂O₅ 2.5 wt%, B₂O₃ 1.3 wt%), known to be resistant to crystallization, was used in sintering of glass granules (300–500 µm) into porous scaffolds. The dissolution behavior of the scaffolds was then studied in vivo in rabbit femurs and under continuous flow conditions in vitro (14 days in vitro/56 days in vivo). The scaffolds were osteoconductive in vivo, as bone could grow into the scaffold structure. Still, the scaffolds could not induce sufficiently rapid bone ingrowth to replace the strength lost due to dissolution. The scaffolds lost their structure and strength as the scaffold necks dissolved. In vitro, S59 dissolved congruently throughout the 14-day experiments, resulting in only a slight reaction layer formation. Manufacturing BAG scaffolds from S59 that retain their amorphous structure was thus possible. The relatively rapid and stable dissolution of the scaffold implies that the glass S59 may have the potential to be used in composite implants providing initial strength and stable, predictable release of ions over longer exposure times. Graphical Abstract