Optimising viability at the core of large bone reconstructions

The management of critical-size bone defects in oral and maxillofacial surgery consistently faces a major biological barrier: central hypoxia. As soon as the volume of the substitute exceeds natural diffusion capacities, cell death compromises regeneration. For the practitioner, the challenge is therefore to transform a simple passive scaffold into a dynamic environment capable of ensuring oxygen transport while delivering the mechanical stimuli essential for osteocyte activity.

This study is based on a dual technological approach: the 3D printing of functionalised polycaprolactone (PCL) matrices and the use of perfusion bioreactors. The author tested the hypothesis that a specific surface functionalisation — combining carboxymethyl κ-carrageenan, carbonated nanohydroxyapatite, collagen and chemical etching — would optimise osteogenesis. In parallel, the work postulates that finite element modelling of fluid shear stresses helps prevent central necrosis. The objective is to demonstrate that the coupling between fluid dynamics and oxygen transport constitutes the missing link to successfully achieve personalised and viable grafts without systematically resorting to autogenous harvesting.

Methodology: Tissue engineering and numerical modelling

This doctoral research is based on a dual approach combining in vitro experimentation and finite element modelling (FEM) to address the issue of hypoxia in critical-sized bone defects. The study used polycaprolactone (PCL) scaffolds designed by 3D printing, whose biochemical and mechanical properties were optimised via several surface functionalisation protocols.

The experimental design compared and analysed the following variables:

• Surface treatments: Comparison of wet-chemical etching (wet-chemical etching) versus plasma-assisted methods.
• Bio-activation: Immobilisation of carboxymethyl κ-carrageenan and incorporation of high concentrations of carbonated nanohydroxyapatite associated with collagen.
• Culture dynamics: Use of perfusion bioreactors to ensure oxygen supply and simulate mechanical stress.

Quantitative analysis was based on finite element modelling to accurately map fluid flow-induced shear stress and oxygen diffusion gradients. These models enabled the quantification of the impact of perfusion on the survival of cells seeded at the core of the 3D structures, while correlating mechanical stimuli with the mechanotransduction signalling pathways of osteocytes.

Optimising the regeneration of critical bone defects: the balance between perfusion and oxygenation

Faced with critical-size bone defects, the intrinsic regenerative capacity of bone reaches its limits, necessitating the use of tissue engineering. The major challenge remains cell survival at the core of large-volume substitutes: without adequate oxygen diffusion, hypoxia condemns the central cells, compromising graft integration. This doctoral study explored the interaction between fluid dynamics and oxygen transport within 3D-printed PCL scaffolds, using perfusion bioreactors to transform these passive structures into active and mechanically stimulating biological environments.

Results: Functionalisation and fluid dynamics

Studies conducted on polycaprolactone (PCL) scaffolds reveal that surface modification is a determining factor for osteogenesis. The study compared several functionalisation approaches and quantified the impact of perfusion on cell viability.

The use of finite element modelling (FEM) enabled the quantification of the shear stresses induced by the flow and oxygen diffusion. The main observations show that:

• Prevention of necrosis: Perfusion bioreactors are identified as essential tools, preventing hypoxia-induced cell death in the core of critical-sized scaffolds.
• Mechanotransduction: Fluid flow generates specific mechanical stimuli that osteocytes sense, activating the signalling pathways necessary for bone remodelling.
• Structure-function synergy: The combination of a high concentration of nanohydroxyapatite and an optimised 3D-printed architecture maintains stable oxygen transport while supporting mechanical loads.

From cellular survival to active osseointegration

The regeneration of critical-size bone defects remains a major challenge in maxillofacial surgery, where insufficient oxygen diffusion at the core of biomaterials compromises cell survival. This research explores how the interaction between fluid dynamics and oxygen transport within 3D-printed and functionalised polycaprolactone (PCL) scaffolds helps to overcome this biological barrier through the use of perfusion bioreactors.

The results of this study demonstrate that osteoblast survival in large-volume structures depends on precise fluid mechanical stimulation (shear stress) and constant oxygenation, making perfusion bioreactors essential to prevent central hypoxia. The integration of carbonated nanohydroxyapatite and collagen, combined with wet-chemical etching, outperforms conventional plasma treatments by significantly improving biochemical properties and osteogenic activity. The study highlights that osteocyte mechanotransduction is the essential driver of this remodelling process.

The weak point lies in the technological transition: although finite element modelling validates the efficacy of the system, direct clinical application remains complex due to bioreactor logistics. Nevertheless, by proving that customised constructs can equal, or even surpass, traditional autologous grafts, this work marks a turning point towards precision tissue engineering in oral and maxillofacial surgery.

Summary of results

This study demonstrates that the functionalisation of PCL scaffolds by wet chemical etching, coupled with the integration of carbonated nanohydroxyapatite and collagen, significantly optimises osteogenic activity. The use of perfusion bioreactors proves essential to prevent hypoxia-induced cell death at the core of critical-sized structures, while activating osteocyte mechanotransduction via a precisely quantified fluid shear stress.

In practical terms, for the practitioner:

• Go beyond simple diffusion: For critical-size bone defects, passive diffusion is insufficient; only active perfusion (via a bioreactor or early neovascularisation) ensures cell survival at the centre of the graft.
• Optimise adhesion: Favour biomaterials whose surface has been chemically treated and enriched with nanohydroxyapatite, as these modifications are superior to plasma for promoting cell anchorage and differentiation.
• Harness the mechanics: Remember that fluid flow is not merely a nutrient carrier, but a crucial mechanical signal that biologically drives bone remodelling.

Technical glossary of the study

Critical-sized bone defects: Bone lesions whose extent exceeds the body's intrinsic regenerative capacities, necessitating the use of structured scaffolds to guide tissue reconstruction.

3D PCL scaffolds (3D-printed PCL scaffolds) : Three-dimensional printed polycaprolactone structures serving as a support matrix for cell proliferation and osteogenic differentiation in tissue engineering.

Perfusion bioreactors: Active culture devices ensuring continuous fluid circulation through the scaffold to overcome oxygen diffusion limitations and prevent hypoxia-induced cell death at the core of the structure.

Mechanotransduction: Biological process by which bone cells, notably osteocytes, convert mechanical stimuli (such as shear stress induced by fluid flow) into biochemical signals regulating bone remodelling.

Finite element modelling: Numerical simulation method used in this study to obtain quantitative information on fluid dynamics and oxygen distribution within the complex geometries of the scaffolds.

Surface functionalisation: Biochemical approaches consisting of immobilising molecules (e.g. carboxymethyl κ-carrageenan, collagen) onto the scaffold to improve its mechanical properties and stimulate osteogenic activity.

Wet-chemical etching: Surface treatment technique favoured in this research over plasma methods to optimise the interface between the biomaterial and the cells.

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