Efficacy and safety of a new heterologous fibrin biopolymer on socket bone healing after tooth extraction: An experimental pre-clinical study.

AIM: To assess the efficacy of heterologous fibrin biopolymer (HFB) in promoting alveolar bone healing after tooth extraction in rats. MATERIALS AND METHODS: The upper right incisors of 48 Wistar rats were extracted. Toothless sockets were filled with HFB (HFBG, n = 24) or blood clot (BCG, n = 24). The tooth extraction sites were subjected to micro-computed tomography (micro-CT), histological, histomorphometric and immunohistochemical (for Runt-related transcription factor 2/Runx2 and tartrate-resistant acid phosphatase/TRAP) analyses on days 0, 7, 14 and 42 after extraction. RESULTS: Socket volume remained similar between days 0 and 14 (69 ± 5.4 mm3), except in the BCG on day 14, when it was 10% lower (p = .043). Although the number of Runx2+ osteoblasts was high and similar in both groups (34 × 102 cells/mm2), the HFBG showed lower inflammatory process and osteoclast activity than BCG at 7 days. On day 14, the number of Runx2+ osteoblasts remained high and similar to the previous period in both groups. However, osteoclast activity increased. This increase was 55% lower in the HFBG than BCG. In the BCG, the presence of an inflammatory process and larger and numerous osteoclasts on day 14 led to resorption of the alveolar bone ridge and newly formed bone. On day 42, numbers of Runx2+ osteoblast and TRAP+ osteoclasts decreased dramatically in both groups. Although the BCG exhibited a more mature cortical bone formation, it exhibited a higher socket reduction (28.3 ± 6.67%) and smaller bone volume (37 ± 5.8 mm3) compared with HFBG (socket reduction of 14.8 ± 7.14% and total bone volume of 46 ± 5.4 mm3). CONCLUSIONS: HFB effectively suppresses osteoclast activity and reduces alveolar bone resorption compared with blood clot, thus preventing three-dimensional bone loss, particularly during the early healing period. HFB emerges as a promising biopharmaceutical material for enhancing healing processes after tooth extraction.

Tissue Bioengineering with Fibrin Scaffolds and Deproteinized Bone Matrix Associated or Not with the Transoperative Laser Photobiomodulation Protocol.

Extending the range of use of the heterologous fibrin biopolymer, this pre-clinical study showed a new proportionality of its components directed to the formation of scaffold with a lower density of the resulting mesh to facilitate the infiltration of bone cells, and combined with therapy by laser photobiomodulation, in order to accelerate the repair process and decrease the morphofunctional recovery time. Thus, a transoperative protocol of laser photobiomodulation (L) was evaluated in critical bone defects filled with deproteinized bovine bone particles (P) associated with heterologous fibrin biopolymer (HF). The groups were: BCL (blood clot + laser); HF; HFL; PHF (P+HF); PHFL (P+HF+L). Microtomographically, bone volume (BV) at 14 days, was higher in the PHF and PHFL groups (10.45 ± 3.31 mm3 and 9.94 ± 1.51 mm3), significantly increasing in the BCL, HFL and PHFL groups. Histologically, in all experimental groups, the defects were not reestablished either in the external cortical bone or in the epidural, occurring only in partial bone repair. At 42 days, the bone area (BA) increased in all groups, being significantly higher in the laser-treated groups. The quantification of bone collagen fibers showed that the percentage of collagen fibers in the bone tissue was similar between the groups for each experimental period, but significantly higher at 42 days (35.71 ± 6.89%) compared to 14 days (18.94 ± 6.86%). It can be concluded that the results of the present study denote potential effects of laser radiation capable of inducing functional bone regeneration, through the synergistic combination of biomaterials and the new ratio of heterologous fibrin biopolymer components (1:1:1) was able to make the resulting fibrin mesh less dense and susceptible to cellular permeability. Thus, the best fibrinogen concentration should be evaluated to find the ideal heterologous fibrin scaffold.

Use of Photobiomodulation Combined with Fibrin Sealant and Bone Substitute Improving the Bone Repair of Critical Defects.

In this preclinical protocol, an adjunct method is used in an attempt to overcome the limitations of conventional therapeutic approaches applied to bone repair of large bone defects filled with scaffolds. Thus, we evaluate the effects of photobiomodulation therapy (PBMT) on the bone repair process on defects filled with demineralized bovine bone (B) and fibrin sealant (T). The groups were BC (blood clot), BT (B + T), BCP (BC + PBMT), and BTP (B + T + PBMT). Microtomographically, BC and BCP presented a hypodense cavity with hyperdense regions adjacent to the border of the wound, with a slight increase at 42 days. BT and BTP presented discrete hyperdensing areas at the border and around the B particles. Quantitatively, BCP and BTP (16.96 ± 4.38; 17.37 ± 4.38) showed higher mean bone density volume in relation to BC and BT (14.42 ± 3.66; 13.44 ± 3.88). Histologically, BC and BCP presented deposition of immature bone at the periphery and at 42 days new bone tissue became lamellar with organized total collagen fibers. BT and BTP showed inflammatory infiltrate along the particles, but at 42 days, it was resolved, mainly in BTP. In the birefringence analysis, BT and BTP, the percentage of red birefringence increased (9.14% to 20.98% and 7.21% to 27.57%, respectively), but green birefringence was similar in relation to 14 days (3.3% to 3.5% and 3.5% to 4.2%, respectively). The number of osteocytes in the neoformed bone matrix proportionally reduced in all evaluated groups. Immunostaining of bone morphogenetic protein (BMP­2/4), osteocalcin (OCN), and vascular endothelial growth factor (VEGF) were higher in BCP and BTP when compared to the BC and BT groups (p < 0.05). An increased number of TRAP positive cells (tartrate resistant acid phosphatase) was observed in BT and BTP. We conclude that PBMT positively influenced the repair of bone defects filled with B and T.

Concentration-dependent effects of latex F1-protein fraction incorporated into deproteinized bovine bone and biphasic calcium phosphate on the repair of critical-size bone defects.

F1-protein fraction (F1) is a natural bioactive compound extracted from the rubber tree, Hevea brasiliensis, and has been recently studied for its therapeutic potential in wound healing. In this study, we investigated the concentration-dependent effects of F1 (0.01%, 0.025%, 0.05%, and 0.1%) incorporated into deproteinized bovine bone (DBB) and porous biphasic calcium phosphate (pBCP), on the repair of rat calvarial critical-size bone defects (CSBD). The defects were analyzed by 3D-microtomography and 2D-histomorphometry at 12 weeks postsurgery. The binding efficiency of F1 to pBCP (96.3 ± 1.4%) was higher than that to DBB (67.7 ± 3.3%). In vivo analysis showed a higher bone volume (BV) gain in all defects treated with DBB (except in 0.1% of F1) and pBCP (except in 0.05% and 0.1% of F1) compared to the CSBD without treatment/control group (9.96 ± 2.8 mm3 ). DBB plus 0.025% F1 promoted the highest BV gain (29.7 ± 2.2 mm3 , p < .0001) compared to DBB without F1 and DBB plus 0.01% and 0.1% of F1. In the pBCP group, incorporation of F1 did not promote bone gain when compared to pBCP without F1 (15.9 ± 4.2 mm3 , p > .05). Additionally, a small BV occurred in defects treated with pBCP plus 0.1% F1 (10.4 ± 1.4 mm3, p < .05). In conclusion, F1 showed a higher bone formation potential in combination with DBB than with pBCP, in a concentration-dependent manner. Incorporation of 0.25% F1 into DBB showed the best results with respect to bone formation/repair in CSBD. These results suggest that DBB plus 0.25% F1 can be used as a promising bioactive material for application in bone tissue engineering.

Evaluation of the Use of an Inorganic Bone Matrix in the Repair of Bone Defects in Rats Submitted to Experimental Alcoholism.

To assess the effects of chronic alcoholism on the repair of bone defects associated with xenograft. Forty male rats were distributed in: control group (CG, n = 20) and experimental group (EG, n = 20), which received 25% ethanol ad libitum after a period of adaptation. After 90 days of liquid diet, the rats were submitted to 5.0-mm bilateral craniotomy on the parietal bones, subdividing into groups: CCG (control group that received only water with liquid diet and the defect was filled with blood clot), BCG (control group that received only water with liquid diet and the defect was filled with biomaterial), CEG (alcoholic group that received only ethanol solution 25% v/v with liquid diet and the defect was filled with blood clot), and BEG (alcoholic group that received only ethanol solution 25% v/v with liquid diet and the defect was filled with biomaterial). In the analysis of body mass, the drunk animals presented the lowest averages in relation to non-drunk animals during the experimental period. Histomorphologically all groups presented bone formation restricted to the defect margins at 60 days, with bone islets adjacent to the BCG biomaterial particles. CEG showed significant difference compared to BEG only at 40 days (17.42 ± 2.78 vs. 9.59 ± 4.59, respectively). In the birefringence analysis, in early periods all groups showed red-orange birefringence turning greenish-yellow at the end of the experiment. The results provided that, regardless of clinical condition, i.e., alcoholic or non-alcoholic, in the final period of the experiment, the process of bone defect recomposition was similar with the use of xenograft or only clot.

In vitro and in vivo assessment of CaP materials for bone regenerative therapy. The role of multinucleated giant cells/osteoclasts in bone regeneration.

In this work, bone formation/remodeling/maturation was correlated with the presence of multinucleated giant cells (MGCs)/osteoclasts (tartrate-resistant acid phosphatase [TRAP]-positive cells) on the surface of beta-tricalcium phosphate (ß-TCP), sintered deproteinized bovine bone (sDBB), and carbonated deproteinized bovine bone (cDBB) using a maxillary sinus augmentation (MSA) in a New Zealand rabbit model. Microtomographic, histomorphometric, and immunolabeling for TRAP-cells analyses were made at 15, 30, and 60 days after surgery. In all treatments, a faster bone formation/remodeling/maturation and TRAP-positive cells activity occurred in the osteotomy region of the MSA than in the middle and submucosa regions. In the ß-TCP, the granules were rapidly reabsorbed by TRAP-positive cells and replaced by bone tissue. ß-TCP enabled quick bone regeneration/remodeling and full bone and marrow restoration until 60 days, but with a significant reduction in MSA volume. In cDBB and sDBB, the quantity of TRAP-positive cells was smaller than in ß-TCP, and these cells were associated with granule surface preparation for osteoblast-mediated bone formation. After 30 days, more than 80% of granule surfaces were surrounded and integrated by bone tissue without signs of degradation, preserving the MSA volume. Overall, the materials tested in a standardized preclinical model led to different bone formation/remodeling/maturation within the same repair process influenced by different microenvironments and MGCs/osteoclasts. © 2019 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 108B:282-297, 2020.