Characterization Platform of Biomaterial-Cell Interfaces in the context of bone tissue engineering – CharaBioC

Calcium-Phosphate bioceramics were produced with one-by-one variation of a chemical-physical parameter (e.g. chemical composition, porosity) with the aim to carry out comparisons of biological responses and to discriminate the impact of each of these parameters variation on bone tissue cells. Relevant cell culture methods and biological models were developed for the study of the cell-living interface, taking into account the features of bone tissue and the specific function of cell types essential for bone regeneration. Such an approach allowed to overcome two major barriers: 1) the limitations of static (standard) 2D cell culture, and 2) the complexity of interactions occurring between the different cell types involved in bone healing. A 3D flow cell culture system has been set up to mimic the mechanical constraints inherent to the circulation of body fluids with a significant influence on cell physiology. Models of co-culture with different cell types: bone-producing cells, cells involved in vascularization and cells from the immune system are currently being developed.

In addition to the scientific results, this project has enabled the team to acquire autonomy in the characterisation of bioceramics developed in the laboratory. A research activity in cell biology and bone tissue engineering has been fully integrated into the team's activities, reinforcing its multidisciplinary skills. This theme has been integrated into the pedagogic content of scientific courses taught at the University of Limoges. The project has led to a high success rate in several calls for projects, and international collaborations have been initiated or strengthened.

One of the direct consequences of the project is the maintenance of the laboratory's activity for evaluation of the biological properties of bioceramics and the understanding of the living-bioceramics interface. 4 research axis stem from this project and are currently followed through various projects: i) Assessment of the biological properties of biomaterials based on optimized calcium phosphate ceramics for applications in bone regenerative medicine. ii) Understanding of the living-material interface in order to identify relevant levers of action on which to act when designing these biomaterials to enhance their applicative properties. iii) The development of biological models and biophysical and chemical methods that come in addition to standard biological methods in order to acquire in-depth knowledge of the bioceramic/cell interface. iv) The development of hybrid living/bioceramic materials for bone tissue engineering.

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Challenges and approach followed in the project were published as a literature review (Magnaudeix, 2022) and an article describing the influence of bioceramic microarchitecture on angiogenesis have been published (Usseglio et al., 2022). An article on copper-doped bioceramics and another review are in preparation. The work carried out in the project is detailed in two thesis manuscripts (A. Brunel, 2023, J. Usseglio, 2024, Université de Limoges). This work has also been disseminated at conferences and vulgarization activities are being carried out, particularly in secondary schools (lectures, Scientibus action).

Calcium phosphate ceramics (CPC), including hydroxyapatite (HA), have been used as bone substitutes for more than 30 years. Their chemical composition, close to that of the bone mineral, confers them good biological properties. However, they are not sufficient to fit with all the needs in bone regenerative medicine, such as in the context of large bone defects. Therefore, it is essential to improve their biological performances in order to extend their application domains. Two approaches are mainly used in this way: 1) the substitution of the calcium phosphate crystalline lattice with chemical elements able to stimulate bone repair. 2) the modulation of the CPC architecture to optimize the cellular responses at the interface. However, a single action is sufficient to modify both their chemical physical characteristics and the associated biological properties. In addition, the variability of the methodologies for production of defined biomaterial and to characterize induce a lack of consensus from the results published in the literature. Theses observations highlight the need to develop consistent methods for characterizing the interface between living cells and materials and to set up relevant in vitro models. It will be integrated into the chain of ceramic materials manufacturing process, the historical and recognized core activity of the IRCER laboratory (University of Limoges, CNRS) in which it will be applied. CharaBioc will bring new skills in the field of life sciences for a better knowledge of material/living interactions and the development of innovative biomaterials meeting public health needs.
This biological analysis platform will lead to the sequential association of the elaboration processes with the physico-chemical properties of three-dimensional structured CPCs (e.g. chemical composition, porosity, roughness...) and their biological performance. Micro-macroporous CPCs based on stoichiometric HA or substituted by chemical elements of interest (CO3, Si, Cu), fully characterized from a physico-chemical point of view, will be produced. The variables will be modified step by step and associated with the changes in the characteristics of the final ceramics. Their dissolution properties in biological environment will first be evaluated under dynamic conditions through the identification and dosage of the chemical species released over time. The effects of these dissolution products will be tested on different cell lines involved in triggering the repair process via recruitment of progenitors (innate immunity cells), vascularization (endothelial cells), an essential process, and bone formation (bone cells). In a second step, the living/material interface will be studied in detail. After validating the biocompatibility of the materials tested, cell proliferation, activation and/or differentiation will be studied in three-dimensional flow perfusion bioreactor cultures. The impact of biomaterials on cellular communication will also be taken into account through the analysis of secreted soluble factors. This will lead, at the end of the project, to the implementation of a reliable and robust 3D cellular model of bone microenvironment, based on co-cultures of cells from the three physiological systems mentioned above. Finally, the use of multiparametric statistical analysis will help to identify direct connections between all collected data, whatever their origin. The final objective is to link biological and chemical physical data in a rational way to identify pertinent levers for the optimization and the enhancement of the ceramic biomaterials applicative performances .