Clarissa Orrico

Bioactive glasses (BGs) have been thoroughly investigated for their osteogenic and antibacterial potential, which may be modulated through their composition. In this study, two different poly(ε-caprolactone) (PCL)/BG composites, SBA3 and SBA3_Cu, were fabricated using fused deposition modeling to obtain planar samples for antibacterial assays and scaffolds with a bone-like tridimensional architecture, while neat PCL constructs were kept as controls. Escherichia coli was selected as prototypical Gram-negative model for investigating implant-associated bone infections. The scaffolds were seeded with adipose derived stem cells (ASCs), a model of osteoblastic precursors, to evaluate their osteogenic properties. Both the composites containing SBA3 and SBA3_Cu relevantly reduced the bacterial metabolic activity and viability after 5 h, while the gene expression analysis of RUNX2, OSX and ALP did not reveal any significant difference among the tested biomaterials. MicroCT analysis demonstrated consistent mineral matrix formation in osteogenic medium, with the exception of PCL/SBA3_Cu. These findings suggest that copper addition could not produce any beneficial osteogenic or antibacterial effect, within the experimental setting adopted. Hence, optimizing the elemental composition and dissolution kinetics of copper-doped BGs is mandatory to provide appropriate differentiative cues to mesenchymal stromal cells, maintaining, at the same time, the antibacterial features required for bone alloplastic materials.

Background/Objectives: An ideal bone scaffold should promote bone cell growth and functional vascularization, hence the importance of imbuing biomaterials with pro-angiogenic cues. In this work, silica-based bioactive glasses, either pristine (SBA3) or doped with copper (SBA3_Cu), were embedded in poly(ε-caprolactone) (PCL), which was also used as a control. Methods: In vitro co-cultures of adipose-derived mesenchymal stem/stromal cells (ASCs) and human microvascular endothelial cells (HMEC-1s) were kept in α-MEM, MCDB131, and EndoGRO media to test the biomaterials. The co-cultures were visualized by immunofluorescence and SEM, while flow cytometry was performed to characterize cellular immunophenotype. The angiogenic potential was evaluated using conditioned media of co-cultures to perform a tubulogenesis assay and VEGF-A quantification. Results: Immunophenotypic analysis showed a significant decrease in the endothelial CD31+ cellular subset, whereas the OB-like cellular subset expressing CD105, CD73, CD90, and ALP increased in all culture media over time. In α-MEM, HMEC-1s were unable to form a capillary network independent of the substrates. A more organized network was visible when co-cultures were plated on PCL, in MCDB131 and EndoGRO, or if they were kept in EndoGRO on PCL/SBA3_Cu. The VEGF-A concentrations were similar in the conditioned media from co-cultures grown on PCL/SBA_Cu, in EndoGRO, and on PCL and PCL/SBA3, in MCDB131. Conclusions: The presence of copper did not promote the angiogenic potential of HMEC-1, likely due to the low concentration of released copper ions and the predominant osteoinductive effect of the other ions released by the bioglass. A re-evaluation of formulation and structure of bioglass scaffold could enhance the angiogenic potential.

The increasing demand for smart bone substitutes has boosted the implementation of biomaterials possibly endowed with both pro-osteogenic and pro-angiogenic capabilities, among which bioactive glasses hold great potential. Hence, two Poly(ε-caprolactone) (PCL)-based composites were loaded at 10 wt.%, with either pristine (SBA3) or copper-doped (SBA3_Cu) silica-based bioactive glasses, through a solvent casting method with chloroform. Neat PCL was used as a control. Samples produced by 3D printing underwent SEM and EDX analyses, and the following were measured: tensile strength and hardness, surface roughness, ion release through ICP-OES, surface free energy, and optical contact angle. Adipose-derived mesenchymal stem cells (ASCs) and human microvascular endothelial cells (HMEC-1) were used to test the biocompatibility of the materials through cell adhesion, spreading, and viability assays. A significant improvement in tensile strength and hardness was observed especially with Cu-doped composites. Both SBA3 and SBA3_Cu added to the PCL favored the early adhesion and the proliferation of HMEC-1 after 3 and 7 days, while ASCs proliferated significantly the most on the SBA-containing composite, at all the time points. Cellular morphology analysis highlighted interesting adaptation patterns to the samples. Further biological characterizations are needed to understand thoroughly how specific bioactive glasses may interact with different cellular types.

The role of periodontal ligament stem cells (PDLSCs) and dental pulp stem cells (DPSCs) in stimulating angiogenesis has been reported, but their angiogenetic potential has not been directly compared. In this work, paired PDLSCs and DPSCs, i.e., derived from the same donor, were tested for their immunophenotype and multi-differentiation capabilities, with particular emphasis on their pro-angiogenic activity. Flow cytometry was utilized to study the expression of mesenchymal stem cell, pericyte, and endothelial markers, while gene expression was evaluated through real-time PCR. The angiogenic potential was assessed recurring to tubulogenesis assay, co-cultures with Human Microvascular Endothelial Cell (HMEC-1), and VEGF-A quantification. The immunophenotype of DPSCs and PDLSCs was different in CD146+ and CD31+ cell subsets, but both cell types promoted HMEC-1 tubulogenesis in vitro. Consistently, VEGF-A gene expression level and its quantification in cell-conditioned media of PDLSCs and DPSCs was comparable between them, and both promoted the formation of vessel-like structures, when co-cultured with HMEC-1 cells. All together, these results showed the heterogeneity of PDLSCs and DPSCs, which are constituted of different cellular subsets, likely modulated by the microenvironmental cues. PDLSCs and DPSCs showed comparable pro-angiogenic activity, enhanced by the contemporary expression of angiogenic and chemotactic factors.

Bone graft substitutes combining the mechanical features of poly-ε-caprolactone (PCL) and the bioactivity of β-tricalcium phosphate (β-TCP) have been widely reported in the literature. Surprisingly, however, very little is known about the incorporation of carbonate at a biomimicking level. The authors studied β-TCP/PCL composites at 20 wt.% and 40 wt.%, either enriched or not with sodium bicarbonate (at 2 wt.% and 4 wt.%), through SEM and EDX analyses; surface free energy estimation; pH measurement after 1, 2, and 3 days of incubation in cell media; nanoindentation; and a protein adsorption test with bovine serum albumin. The early biological response was assessed using adipose mesenchymal stem cells, as an established in vitro model, via cellular adhesion (20 min), spreading (24 h), and viability assays (1, 3, 7 days). By increasing the β-TCP content, the composites’ hardnesses and Young’s moduli (EiT) were improved, as well as their protein adsorption compared to neat PCL. Sodium bicarbonate increased the polar component of the surface energy, alkalinized the composite with a higher β-TCP content, and attenuated its early negative cell response. Further investigation is needed to deepen the knowledge of the mechanisms underpinning the mechanical features and long-term biological behavior.

The improvement of the mucosal sealing around the implant represents a challenge, one that prompted research into novel materials. To this purpose, a printable poly(ε-caprolactone) (PCL)-based composite loaded with alumina-toughened zirconia (ATZ) at increasing rates of 10, 20, and 40 wt.% was prepared, using a solvent casting method with chloroform. Disks were produced by 3D printing; surface roughness, free energy and optical contact angle were measured. Oral fibroblasts (PF) and epithelial cell (SG) tests were utilized to determine the biocompatibility of the materials through cell viability assay and adhesion and spreading evaluations. The highest level of ATZ resulted in an increase in the average roughness (S_a), while the maximum height (S_z) was higher for all composites than that of the unmixed PCL, regardless of their ATZ content. Surface free energy was significantly lower on PCL/ATZ 80/20 and PCL/ATZ 60/40, compared to PCL and PCL/ATZ 90/10. The contact angle was inversely related to the quantity of ATZ in the material. PF grew without variations among the different specimens at 1 and 3 days. After 7 days, PF grew significantly less on PCL/ATZ 60/40 and PCL/ATZ 80/20 compared to unmixed PCL and PCL 90/10. Conversely, ATZ affected and improved the growth of SG. By increasing the filler amount, PF cell adhesion and spreading augmented, while PCL/ATZ 80/20 was the best for SG adhesion. Overall, PCL/ATZ 80/20 emerged as the best composite for both cell types; hence, it is a promising candidate for the manufacture of custom made transmucosal dental implant components.

This in vitro study assessed the efficacy of a solution containing 33% trichloroacetic acid (CCl₃COOH; TCA) and hydrogen peroxide (H₂O₂) in decontaminating machined (MAC) and sand-blasted acid-etched (SBAE) titanium surfaces. A total of 80 titanium disks were prepared (40 MAC and 40 SBAE). Streptococcus sanguinis and Enterococcus faecalis strains were incubated on 36 samples, while the remaining 44 were kept as controls. Roughness analysis and scanning electron microscopy were used to evaluate the surface features before and after TCAH₂O₂ treatment. The viability of human adipose-derived mesenchymal stem cells (ASCs) after TCAH₂O₂ decontamination was assessed with a chemiluminescent assay along with cell morphology through fluorescent staining. TCAH₂O₂ preserved the surface topography of MAC and SBAE specimens. It also effectively eradicated bacteria on both types of specimens without altering the surface roughness (p > 0.05). Also, no significant differences in protein adsorption between the pristine and TCAH₂O₂-treated surfaces were found (p = 0.71 and p = 0.94). While ASC proliferation remained unchanged on MAC surfaces, a decrease was observed on the decontaminated SBAE specimens at 24 and 48 h (p < 0.05), with no difference at 72 h (p > 0.05). Cell morphology showed no significant changes after 72 h on both surface types even after decontamination. This study suggests TCAH₂O₂ as a promising decontamination agent for titanium surfaces, with potential implications for peri-implant health and treatment outcomes.