## 4 Discussion  
Research into shark tooth-derived calcium phosphate compounds with traces of F, Na and Mg ions marks a recent development in the treatment of bone defects [17]. The crystalline structure of this bone graft is a combination of an apatite phase (65-70%), including hydroxyapatite and fluorapatite, and a non-aquitic phase (35-30%), mainly whilockite. Preliminary in vitro studies have shown that this biphasic marine bone graft promotes the proliferation of MC3T3 preosteoblasts after 21 days of incubation and results in a significant increase in osteogenic activity (\(p<0.01\)) [17]. Moreover, in vivo assays in a rodent model demonstrated an excellent osseointegration grade 3 weeks after implantation, with significantly higher bone mineral density (\(p<0.05\)) than commercial synthetic HA/TCP bone graft (60%/40%) [18], and potential osteeconductive properties, due to the porous morphology and the formation of intergranular cavities which allowed the ingrowth of new bone tissue cells. Biomaterials for clinical applications are subject to the strict requirements set out in ISO 10993-6:2007 (E), which specifies \"test methods for the assessment of the local effects after implantation of biomaterials intended for use in medical devices\". The purpose of such tests is to establish the safety and efficacy of the biomaterial in a biological environment, to ensure that it does not generate adverse reactions or release toxic substances. Our study demonstrated the biocompatibility of a marine-origin bone graft obtained from shark teeth. The preclinical trial carried out on critical femoral defects in rabbits showed a non-irritant behavior and revealed that the test group (marine-origin bone graft) presented greater osseointegration at 12 weeks than the control group (bovine-origin bone graft). Results from the histological analysis showed a higher area of occupancy by bone  
Figure 5: Boxplot and table of the microtomographic results. Percentage of the bone graft + bone tissue (BIOMAT + BONE), the bone tissue, and the intersection in percentage for the marine bone graft (test) and the Bio-Oss® (control). tissue in the test material (38 \(\pm\) 5%) than in the control (30 \(\pm\) 6%). Additional evaluation by micro-CT analysis confirmed this finding, given that values for bone mineral density were statistically higher in the test group (54 \(\pm\) 6%) than in the control group (27 \(\pm\) 8%). Moreover, the measurement of the bone graft-bone tissue intersection was much higher for the shark tooth-derived bone graft (86 \(\pm\) 8%) than the control (30 \(\pm\) 1%). While these figures confirm the superiority of the test bone graft in terms of bone formation and intersection measurements, the actual microtomographic values are likely to be overestimated, probably due to the similar radiological characteristics of the control sample and the bone tissue. This fact would also explain the large differences in the percentage of intersection. Nevertheless, taken as a whole, the results of the histometric and microtomographic analysis support the conclusion that the specific composition and structure of this marine-origin bone graft plays a key role in the promotion of bone regeneration. The presence of fluor in the form of fluorine in the marine bone graft composition participates in the biochemical processes that enhance the synthesis of growth factors in bone cells, contributing to the differentiation of osteoprogenitor cells [22] and also to bone consolidation and regeneration [23]. In the form of fluorapatite, this element promotes a more crystalline and therefore more stable apatite [24], improving the mechanical properties with respect to hydroxyapatite. The non-apatite phase, on the other hand, is less crystalline and therefore less stable, allowing some degree of resorption. The incorporation of Mg in the form of whitlockite is also of interest because of its influence on the synthesis of parathyroid hormone, which regulates bone homeostasis [25].