Marine collagen scaffolds and photobiomodulation on bone healing process in a model of calvaria defects.

INTRODUCTION: Collagen from marine esponges has been used as a promising material for tissue engineering proposals. Similarly, photobiomodulation (PBM) is able of modulating inflammatory processes after an injury, accelerating soft and hard tissue healing and stimulating neoangiogenesis. However, the effects of the associated treatments on bone tissue healing have not been studied yet. In this context, the present study aimed to evaluate the biological temporal modifications (using two experimental periods) of marine sponge collagen or sponging (SPG) based scaffold and PBM on newly formed bone using a calvaria bone defect model. MATERIAL AND METHODS: Wistar rats were distributed into two groups: SPG or SPG/PBM and euthanized into two different experimental periods (15 and 45 days post-surgery). A cranial critical bone defect was used to evaluate the effects of the treatments. Histology, histomorfometry and immunohistological analysis were performed. RESULTS: Histological findings demonstrated that SPG/PBM-treated animals, 45 days post-surgery, demonstrated a higher amount of connective and newly formed bone tissue at the region of the defect compared to CG. Notwithstanding, no difference among groups were observed in the histomorphometry. Interestingly, for both anti-transforming growth factor-beta (TGF-ß) and anti-vascular endothelial growth factor (VEGF) immunostaining, higher values for SPG/PBM, at 45 days post-surgery could be observed. CONCLUSION: It can be concluded that the associated treatment can be considered as a promising therapeutical intervention.

Influence of the incorporation of marine spongin into a Biosilicate®: an in vitro study.

The combination of different biomaterials can be a promising intervention for the composites manufacture, mainly by adding functional and structural characteristics of each material and guarantee the advantages of the use of these composites. In this context, the aim of this study was to develop and evaluated the influence of the incorporation of marine spongin (SPG) into Biosilicate® (BS) in different proportions be used during bone repair. For this purpose, it was to develop and investigate different BS/SPG formulations for physico-chemical and morphological characteristics by pH, loss mass, Fourier transform infrared spectrometer (FTIR) and scanning electron microscope (SEM) analysis. Additionally, the influence of these composites on cell viability, proliferation, and alkaline phosphatase (ALP) activity were investigated. The results revealed that the pH values of all BS groups (with or without SPG) increased over time. A significant mass loss was observed in all composites, mainly with higher SPG percentages. Additionaly, SEM micrographies demonstrated fibers of SPG into BS and material degradation over time. Moreover, FTIR spectral analysis revealed characteristic peaks of PMMA, BS, and SPG in BS/SPG composites. BS/SPG groups demonstrated a positive effect for fibroblast proliferation after 3 and 7 days of culture. Additionally, BS and BS/SPG formulations (at 10% and 20% of SPG) presented similar values of osteoblasts viability and proliferation after 7 days of culture. Furthermore, ALP activity demonstrated no significant difference between BS and BS/SPG scaffolds, at any composition. Based on the present in vitro results, it can be concluded that the incorporation of SPG into BS was possible and produced an improvement in the physical-chemical characteristics and in the biological performance of the graft especially the formulation with 80/20 and 90/10. Future research should focus on in vivo evaluations of this novel composite.

Bioglass/collagen scaffolds combined with bone marrow stromal cells on bone healing in an experimental model in cranial defects in rats.

This study aimed to develop bone regenerative therapeutic strategies, based on the addition of bone marrow stromal cells (BMSC) on bioglass/collagen (BG/COL) scaffolds. For this purpose, an in vivo study was conducted using tissue response of the BG/COL scaffolds combined with BMSC in a critical-size defects. Wistar rats were submitted to the surgical procedure to perform the cranial critical size bone defects and distributed in four groups (20 animals per group): Control Group (CG) (rats submitted to the cranial bone defect surgery without treatment), Bioglass Group (BG) (rats treated with BG), BG/COL Group (rats treated with BG/COL) and Bioglass/Collagen and BMSC Group (BG/COL/BMSC) (rats treated with BG/COL scaffolds enriched with BMSCs). Animals were euthanized 15 and 30 days after surgery. Scanning electron microscopy, histopathological and immunohistochemistry analysis were used. SEM analysis demonstrated that porous scaffolds were obtained, and Col fibers were successfully impregnated to BG matrices. The implantation of the BMSC on BG/COL based scaffolds was effective in stimulating newly bone formation and produced an increased immunoexpression of markers related to the bone repair. These results highlight the potential of BG/COL scaffolds and BMSCs to be used as a therapeutic approach for bone regeneration.

Incorporation of Collagen from Marine Sponges (Spongin) into Hydroxyapatite Samples: Characterization and In Vitro Biological Evaluation.

Biomaterial-based bone grafts have an important role in the field of bone tissue engineering. One of the most promising classes of biomaterials is collagen, including the ones from marine biodiversity (in general, called spongin (SPG)). Also, hydroxyapatite (HA) has an important role in stimulating bone metabolism. Therefore, this work investigated the association of HA and SPG composites in order to evaluate their physico-chemical and morphological characteristics and their in vitro biological performance. For this, pre-set composite disks were evaluated by means of mass loss after incubation, pH, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), and "in vitro" cell viability. pH measurements showed no statistical difference between groups. Moreover, a higher mass loss was observed for HA/SPG70/30 compared to the other groups for all experimental periods. Moreover, SEM representative micrographs showed the degradation of the samples with and without immersion. FTIR analysis demonstrated the absorption peaks for poly(methyl methacrylate) (PMMA), HA, and SPG. A higher L292 cell viability for control and PMMA was observed compared to HA and HA/SPG 90/10. Also, HA/SPG 70/30 showed higher cell viability compared to HA and HA/SPG 90/10 on days 3 and 7 days of culture. Furthermore, HA showed a significant lower MC3T3 cell viability compared to control and HA/SPG 70/30 on day 3 and no significant difference was observed between the composites in the last experimental period. Based on our investigations, it can be concluded that the mentioned composites were successfully obtained, presenting improved biological properties, especially the one mimicking the composition of bone (with 70% of HA and 30% of SPG). Consequently, these data highlight the potential of the introduction of SPG into HA to improve the performance of the graft for bone regeneration applications. Further long-term studies should be carried out to provide additional information concerning the late stages of material degradation and bone healing in the presence of HA/SPG.

Biosilicate/PLGA osteogenic effects modulated by laser therapy: In vitro and in vivo studies.

The main purpose of the present work was to evaluate if low laser level therapy (LLLT) can improve the effects of Biosilicate®/PLGA (BS/PLGA) composites on cell viability and bone consolidation using a tibial defects of rats. The composites were characterized by scanning electron microscope (SEM) and reflection Fourier transform infrared spectrometer (FTIR). For the in vitro study, fibroblast and osteoblast cells were seeded in the extract of the composites irradiated or not with LLLT (Ga-Al-As, 808nm, 10J/cm2) to assess cell viability after 24, 48 and 72h. For the in vivo study, 80 Wistar rats with tibial bone defects were distributed into 4 groups (BS; BS+LLLT; BS/PLGA and BS/PLGA+LLLT) and euthanized after 2 and 6weeks. Laser irradiation Ga-Al-As (808nm, 30J/cm2) in the rats was performed 3 times a week. The SEM and FTIR results revealed that PLGA were successfully inserted into BS and the microparticles degraded over time. The in vitro findings demonstrated higher fibroblast viability in both BS/PLGA groups after 24h and higher osteoblast viability in BS/PLGA+LLLT in all periods. As a conclusion, animals treated with BS/PLGA+LLLT demonstrated an improved material degradation and an increased amount of granulation tissue and newly formed bone.