Can Novel Bioceramics Withstand Osteoclastic Resorption or Act as a Vehicle to Deliver Local Therapies for Treatment of Fractures and Limit Bone Destruction of the Active Charcot Foot in Diabetes?

Background and hypothesis: The active Charcot foot in diabetes is characterised with excessive osteoclastic activity and reduced osteoblastic repair leading to non-healing fractures, bone collapse/ fragmentation and ultimately to significant foot deformity. Despite prolonged immobilisation of the lower limb, many fractures do not fully heal, collapsed bone does not regenerate and fracture non-union is common. We hypothesise that targeted therapy with suitable synthetic bone substitutes could augment healing, repair Charcot bones, and lessen deformity. However, it is unknown whether novel bone substitutes can withstand biological degradation of the highly resorptive Charcot osteoclasts.
Aims: The aim was to investigate how newly formed osteoclasts generated from peripheral blood monocytes derived from people with active Charcot foot degrade different types of bioceramics and compare it with bone resorption in the presence /absence of activators and inhibitors. Next, we assessed the chemical composition of resorbed surfaces before and after the addition of the cathepsin K inhibitor, Odanacatib (ODN).
Methods: Osteoclast precursors isolated from whole blood from people with active Charcot foot were cultured in the presence of macrophage-colony stimulating factor (M-CSF) and receptor activator of nuclear factor-κβ ligand (RANKL) on β-tri-calcium phosphate, hydroxyapatite and bovine bone discs. Osteoclast formation and resorption were assessed after 21 days in vitro with confocal image analysis, contact and optical surface analysis methods. The compositional variation of randomly selected resorbed areas were examined with Raman spectroscopy.
Results: We observed that osteoclast biodegradation of β-TCP discs resembled resorption of bovine bone discs (extracellular resorption with formation of pits), whereas osteoclast biodegradation of HA-discs included intracellular digestion of the material (endocytosis) which was carried out by giant osteoclast-like cells. Initial assessment with contact surface profilometry indicated differences between resorption depth of bone discs vs resorption depth of biomaterials. Optical profilometry facilitated analysis of trenches and pits on bovine bone discs and on biomaterials. This method showed differences between bone resorption (organic and inorganic) and dissolution of biomaterials (inorganic). Pilot experiments with the resorption inhibitor ODN, added in vitro reversed the deep trenches (bulldozer type of resorption) to a less aggressive resorption of pits (drilling resorption).
Next, resorbed bovine bone discs were analysed with Raman spectroscopy. Peaks were normalised to the highest peak at 960 cm-1, which primarily corresponds to phosphate. In comparison to samples in MCSF+RANKL treatment, the addition to ODN led to an average reduction of 20% on peak 853 cm-1 (collagen proline), 14% on 872 cm-1 (collagen hydroxyproline), 33% on peak 1003 cm-1 (phenylalanine), 24% on peak 1340 cm-1 (amide III region) and 52% on peak 1660 cm-1 (amide III region).
Conclusion: These observations have translational potential as they may inform novel therapies with bioceramics coated with an osteoclast inhibitor as targeted therapy to modulate bone destruction of the active Charcot foot.

Acknowledgement: This work was supported by the MRC IAA 2021 Kings College London (MR/X502923/1)