Search Results (25)

This study examines the cytotoxicity of two silver nanoparticle formulations—AgNPs conjugated with chlorhexidine (AgNPs-CHL) and AgNPs conjugated with polyethylene glycol and metronidazole (AgNPs-PEG-MET)—as examples of the surface functionalization of silver nanoparticles with drugs via sulfur–silver bonds and nitrogen–silver interactions. We previously reported the synthesis of these NPs and their efficiency in periodontitis treatment. Here, we analyze the relationships between the cytotoxic mechanisms of AgNPs and their surface chemistry. UV–Vis spectroscopy, dynamic light scattering (DLS), and scanning electron microscopy (SEM) with energy-dispersive X-ray spectroscopy (EDX) were used for physicochemical studies of the conjugates in two environments: aqueous solutions and commonly used cell culture media. Cytotoxicity was assessed in human fetal osteoblasts (hFOB 1.19) and human gingival fibroblasts (HGF-1) through BrdU and LDH assays, ROS detection, cell cycle analysis, apoptosis assays, and protein expression studies. AgNPs-CHL showed aggregation and increased hydrodynamic diameters in the culture medium, while AgNPs-PEG-MET remained stable. Both exhibited concentration-dependent cytotoxicity: AgNPs-CHL at 0.4–10 μg/mL and AgNPs-PEG-MET at 0.75–10 μg/mL. AgNPs-CHL, in which silver surface functionalization was realized via nitrogen–silver interactions, induced significant ROS generation, LDH release, and necroptosis, marked by increased RIP1, RIP3, and MLKL proteins. In the case of AgNPs-PEG-MET, where sulfur–silver bonds combined the drug via a PEG linker, they triggered apoptosis, as evidenced by elevated caspase-2 levels and flow cytometry. These findings highlight that the type of surface functionalization of silver nanoparticles significantly influences their physicochemical behavior and biological effects. Understanding these mechanisms is crucial in designing safer, more effective nanoparticle-based therapies for periodontal and other inflammatory conditions. Full article

This research aims to study the antibacterial coatings of invasive surgical medical devices, including dental implants, to reduce superficial and deep local infections over the long term. To obtain the coating without altering the initial properties of the substrate (dental implant made of TiZr bioalloy), simple, cost-effective, and efficient methods were employed, such as chemical deposition of silver (Ag). The deposition characteristics were analyzed using scanning electron microscopy (SEM), EDX analysis, and FT-IR infrared analysis. The in vitro testing of antimicrobial activity was conducted using the diffusion method by cultivating the bacterial strains Escherichia coli (E. coli) ATCC25922 and Staphylococcus aureus (S. aureus) ATCC25923 and measuring the diameter of the bacterial inhibition zone. Investigations and biocompatibility evaluations were performed on both uncoated and silver-coated (Ag) samples by analyzing cell viability and morphology in the presence of human fetal osteoblasts (hFOB cell line) and human gingival fibroblasts (HFIB-G cells) after 8 days of incubation. The research results confirm the biocompatibility of the coating, demonstrated by the lack of significant differences in cell density between the Ag-coated samples and the control group, as well as by the fact that the silver-coated surface effectively supports actin cytoskeleton organization, adhesion, and migration of both human osteoblasts and gingival fibroblasts. The results regarding the antibacterial efficiency of the silver implant coating indicated that the E. coli bacterial strain is more resistant than S. aureus. The resistance difference between the two bacterial strains was attributed to differences in the structure of their cell envelopes. Full article

Background and aims: Enhanced cell proliferation is crucial for reducing production time and cost in cell therapy, and human platelet lysate (HPL) is often used to boost cell proliferation due to its favorable safety profile. Understanding the roles of different HPL components and their effects on cell culture can lead to more informed choices in medium formulation, which in turn can influence cell behavior and outcomes. Therefore, this study aimed to investigate the effects of two types of HPL, i.e., heparin-supplemented HPL (He-HPL) and fibrinogen-depleted HPL without heparin (Fd-HPL), on human osteoblasts. Materials and Methods: He-HPL and Fd-HPL were prepared from expired platelet concentrates. The presence of growth factors, i.e., brain-derived neurotrophic factor (BDNF) and vascular endothelial growth factor (VEGF), and cytokines, i.e., interleukin-6 (IL-6) and tumor necrosis factor alpha (TNF-α), in HPL was evaluated. Human fetal osteoblast (hFOB) cells were cultured in Dulbecco’s Modified Eagle Medium supplemented with either He-HPL or Fd-HPL. The cell morphology, viability, calcium deposition, and expression of osteogenic genes were assessed. Results: Comparable levels of BDNF (p > 0.05), VEGF (p > 0.05), and IL-6 (p > 0.05) were detected in both types of HPL, whereas He-HPL exhibited significantly higher levels of TNF-α (p < 0.05). However, there were no notable differences in cell morphology, viability, population doubling time, or total cell yield between the two HPL types. Similarly, no differences were observed in the mineralization of cells treated with He-HPL compared to Fd-HPL. Nonetheless, hFOB cells cultured with He-HPL demonstrated significantly higher expression of osteogenic markers Runx2 and ALP (p < 0.05) compared to those cultured with Fd-HPL. Conclusions: He-HPL and Fd-HPL demonstrate comparable performance in promoting osteoblast proliferation and mineralization, making both usable for bone tissue engineering. However, He-HPL might have a slight edge as it enhances osteogenic gene expression. Full article

Background: The burden of prosthetic joint infection in combination with antibiotic-resistant bacterial strains is a rising dilemma for patients experiencing total joint replacements. Around 0.8–2% of patients experience prosthetic joint infections, while up to 21% of patients are considered fatal cases after 5 years. Staphylococcus aureus is one of the main reasons for prosthetic joint infections. Its capability of forming biofilms and developing mechanisms against antibiotics is one of the most dangerous clinical topics being currently discussed. Previous studies have shown the promising results of omega-3 fatty acids as an antimicrobial agent against Staphylococcus aureus. Though an antimicrobial effect has been examined, the influence of polyunsaturated fatty acids on Staphylococcus aureus in the presence of human osteoblasts has not been reported yet. In this study, we aimed to investigate the influence of omega-3 fatty acids on the biofilm formation of Staphylococcus aureus ATCC 29213 in the presence of hFOB 1.19 cells. The co-culture setup helped to examine the influence of omega-3 fatty acids on the race for surface to simulate prosthetic joint infections. Methods: In this study, we tested Staphylococcus aureus ATCC 29213 co-cultured with human fetal osteoblasts hFOB 1.19 in the presence of sub-MIC and MIC concentrations of docosahexaenoic acid (1.25 mg/L, 2.5 mg/L) and eicosapentaenoic acid (0.15 mg/L, 0.3 mg/L) after 1, 6 and 24 h of incubation. After establishing the co-culture, cell culture and biofilm, we performed colony-forming unit counting and cell counting to examine cell survivability. In addition, we carried out scanning electron microscopy to study the race for surface behaviour of the cells. Results: We found a protective influence of omega-3 fatty acids on osteoblasts when present in co-culture with Staphylococcus aureus after 6 h of incubation. Omega-3 fatty acids increase the cell survival of osteoblasts after 6 h in a co-culture with bacteria and are able to influence the race for surface. In this study, the strain of Staphylcoccus aureus ATCC 29213 showed signs of growth inhibition within the first 6 h. Conclusions: Omega-3 fatty acids can be a valuable antimicrobial agent in terms of decreasing the risk of on-site infection during surgery. Omega-3 fatty acids were shown to decrease the bacterial load within the first 6 h of incubation and increase the survivability of osteoblasts. Full article

Objectives: Dental bone formation involves various cellular and molecular mechanisms, and phytoestrogens such as formononetin (FORM) are promising because of their estrogenic, anti-inflammatory, and antioxidant effects. This study investigated the effect of FORM on osteoblast proliferation, differentiation, and mineralization in combination with spongiosa granulates (BO) in vitro. Materials and Methods: Human fetal osteoblast cells (hFOB1.19) were treated with increasing concentrations of FORM (1, 10, and 100 µg/mL), BO, or their combination. Cell proliferation was assessed using a MTT assay. Alkaline phosphatase (ALP) activity, intracellular Ca²⁺, and Pi levels were measured using ELISA. Vascular endothelial growth factor (VEGF) and osteocalcin expression levels were analyzed by western blotting. Results: Cell proliferation increased with FORM, with or without BO, after 6 days (p < 0.001). FORM and BO had a synergistic effect on ALP activity (p < 0.001). Intracellular Ca²⁺ and Pi levels were highest in the BO-FORM group, suggesting superior mineralization (p < 0.05). VEGF and osteocalcin expression was significantly upregulated with FORM, alone and with BO (p < 0.05), indicating improved angiogenesis and bone maturation over 9 days. Conclusions: FORM enhances osteoblast proliferation, differentiation, and mineralization potential, particularly in BO spongiosa granulates. These data support the in vitro potential of formononetin-phytoestrogen in promoting osteoblast differentiation and mineralization potential with BO. These findings suggest that FORM, combined with BO, could improve bone augmentation in clinical applications such as maxillofacial surgery. FORM shows valuable potential for clinical applications, such as maxillofacial surgery, by promoting faster and more effective healing. Full article

Background/Objectives: A biocomposite based on magnesium-doped hydroxyapatite and enriched with amoxicillin (MgHApOx) was synthesized using the coprecipitation method and is presented here for the first time. Methods: The stability of MgHAp and MgHApOx suspensions was evaluated by ultrasound measurements. The structure of the synthesized MgHAp and MgHApOx was examined with X-ray diffraction (XRD), Fourier transform infrared (FT-IR) spectroscopy and X-ray photoelectron spectroscopy (XPS). The crystalline structure was determined by X-ray diffraction. The FTIR data were collected in the range of 4000–400 cm⁻¹. The morphology of the nanoparticles was evaluated by scanning electron microscopy (SEM). Furthermore, the biocompatible properties of MgHAp, MgHApOx and amoxicillin (Ox) suspensions were assessed using human fetal osteoblastic cells (hFOB 1.19 cell line). The antimicrobial properties of the MgHAp, MgHApOx and Ox suspension nanoparticles were assessed using the standard reference microbial strains Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922 and Candida albicans ATCC 10231. Results: X-ray studies have shown that the biocomposite retains the characteristics of HAp and amoxicillin. The SEM assessment exhibited that the apatite contains particles at nanometric scale with acicular flakes morphology. The XRD and SEM results exhibited crystalline nanoparticles. The average crystallite size calculated from XRD analysis increased from 15.31 nm for MgHAp to 17.79 nm in the case of the MgHApOx sample. The energy-dispersive X-ray spectroscopy (EDS) and X-ray photoelectron spectroscopy (XPS) analysis highlighted the presence of the constituent elements of MgHAp and amoxicillin. Moreover, XPS confirmed the substitution of Ca²⁺ ions with Mg²⁺ and the presence of amoxicillin constituents in the MgHAp lattice. The results of the in vitro antimicrobial assay demonstrated that MgHAp, MgHApOx and Ox suspensions exhibited good antimicrobial activity against the tested microbial strains. The results showed that the antimicrobial activity of the samples was influenced by the presence of the antibiotic and also by the incubation time. Conclusions: The findings from the biological assays indicate that MgHAp and MgHApOx are promising candidates for the development of new biocompatible and antimicrobial agents for biomedical applications. Full article

The suppressive effect of bisphosphonates (BPs) on bone metabolism is considered to be a major cause of medication-related osteonecrosis of the jaw (MRONJ). Enamel matrix derivative (EMD) stimulates and activates growth factors, leading to the regeneration of periodontal tissues. In this study, we aimed to explore the potential of EMD in reversing the detrimental effects of BPs on human fetal osteoblasts (hFOBs) and osteosarcoma-derived immature osteoblasts (MG63s) by assessing cell viability, apoptosis, migration, gene expression, and protein synthesis. While the suppressive effect of zoledronate (Zol) on cell viability and migration was observed, the addition of EMD significantly mitigated this effect and enhanced cell viability and migration. Furthermore, an increased apoptosis rate induced by Zol was decreased with the addition of EMD. The decreased gene expression of alkaline phosphatase (ALP), osteocalcin (OC), and the receptor activator of nuclear factors kappa-B ligand (RANKL) caused by BP treatment was reversed by the co-addition of EMD to hFOB cells. This trend was also observed for ALP and bone sialoprotein (BSP) levels in MG63 cells. Furthermore, suppressed protein levels of OC, macrophage colony-stimulating factor (M-CSF), BSP, and type 1 collagen (COL1) were recovered following the addition of EMD. This finding suggests that EMD could mitigate the effects of BPs, resulting in the recovery of cell survival, migration, and gene and protein expression. However, the behavior of the osteoblasts was not fully restored, and further studies are necessary to confirm their effects at the cellular level and to assess their clinical usefulness in vivo for the prevention and treatment of MRONJ. Full article

In this paper, we present for the first time the development of zinc-doped hydroxyapatite enriched with tetracycline (ZnHApTe) powders and provide a comprehensive evaluation of their physico-chemical and biological properties. Various techniques such as X-ray diffraction (XRD), X-ray photoelectron spectroscopy (XPS), scanning electron microscopy (SEM), and Fourier transform infrared spectroscopy (FTIR) were used for the sample’s complex evaluation. Moreover, the biocompatibility of zinc-doped hydroxyapatite (ZnHAp) and ZnHApTe nanoparticles was evaluated with the aid of human fetal osteoblastic cells (hFOB 1.19 cell line). The results of the biological assays suggested that these nanoparticles hold great promise as potential candidates for the future development of novel biocompatible and antimicrobial agents for biomedical applications. The antimicrobial properties of the ZnHAp and ZnHApTe nanoparticles were assessed using the standard reference microbial strains Staphylococcus aureus ATCC 25923, Escherichia coli ATCC 25922, and Candida albicans ATCC 10231. The results of the in vitro antimicrobial assay demonstrated that both tested materials exhibited good antimicrobial activity. Additionally, these data also indicated that the antimicrobial effects of the ZnHAp nanoparticles were intensified by the presence of tetracycline (Te). Furthermore, the results also suggested that the antimicrobial activity of the samples increased with the incubation time. Full article

Natural bone is a complex material that has been carefully designed. To prepare a successful bone substitute, two challenging conditions need to be met: biocompatible and bioactive materials for cell proliferation and differentiation, and appropriate mechanical stability after implantation. Therefore, a hybrid Poly ε-caprolactone/Poly(lactic-co-glycolide)/β-tricalcium phosphate (PCL/PLGA/β-TCP) scaffold has been introduced as a suitable composition that satisfies the above two conditions. The blended PCL and PLGA can improve the scaffold’s mechanical properties and biocompatibility compared to single PCL or PLGA scaffolds. In addition, the incorporated β-TCP increases the mechanical strength and osteogenic potential of PCL/PLGA scaffolds, while the polymer improves the mechanical stability of ceramic scaffolds. The PCL/PLGA/β-TCP scaffold is designed using spiral structures to provide a much better transport system through the gaps between spiral walls than conventional cylindrical scaffolds. Human fetal osteoblasts (hFOBs) were cultured on spiral PCL/PLGA/β-TCP (PPBS), cylindrical PCL/PLGA/β-TCP (PPBC), and cylindrical PCL scaffolds for a total of 28 days. The cell proliferation, viability, and osteogenic differentiation capabilities were analyzed. Compared with PCL and PPBC scaffolds, the PPBS scaffold exhibits great biocompatibility and potential to stimulate cell proliferation and differentiation and, therefore, can serve as a bone substitute for bone tissue regeneration. Full article

Infections related to orthopedic/stomatology surgery are widely recognized as a significant health concern. Therefore, the development of new materials with superior biological properties and good stability could represent a valuable alternative to the classical treatments. In this paper, the fluorine-substituted hydroxyapatite (FHAp) suspension, with the chemical formula Ca₁₀(PO₄)₆(OH)_2−2xF_2x (where x = 0.05), was prepared using a modified coprecipitation technique. Stability studies were conducted by zeta potential and ultrasound measurements for the first time. The X-ray diffraction (XRD) patterns of FHAp powders displayed a hexagonal structure akin to that of pure hydroxyapatite (HAp). The XPS general spectrum revealed peaks corresponding to the constituent elements of fluorine-substituted hydroxyapatite such as calcium, phosphorus, oxygen, and fluorine. The purity of the obtained FHAp samples was confirmed by energy-dispersive X-ray spectroscopy (EDS) studies. The FHAp morphology was evaluated by scanning electron microscopy (SEM) measurements. Fourier-transform infrared spectroscopy (FTIR) studies were performed in order to study the vibrational properties of the FHAp samples. The FHAp suspensions were tested for antibacterial activity against reference strains such as Staphylococcus aureus 25923 ATCC, Escherichia coli ATCC 25922, and Candida albicans ATCC 10231. Additionally, the biocompatibility of the FHAp suspensions was assessed using human fetal osteoblastic cells (hFOB 1.19 cell line). The results of our biological tests suggest that FHAp suspensions are promising candidates for the future development of new biocompatible and antimicrobial agents for use in the biomedical field. Full article

In vitro testing is the first important step in the development of new biomaterials. The human fetal osteoblast cell line hFOB 1.19 is a very promising cell model; however, there are vast discrepancies in cultivation protocols, especially in the cultivation temperature and the presence of the selection reagent, geneticin (G418). We intended to use hFOB 1.19 for the testing of Zn-based degradable metallic materials. However, the sensitivity of hFOB 1.19 to zinc ions has not yet been studied. Therefore, we compared the toxicity of zinc towards hFOB 1.19 under different conditions and compared it with that of the L929 mouse fibroblast cell line. We also tested the cytotoxicity of three types of Zn-based biomaterials in two types of media. The presence of G418 used as a selection reagent decreased the sensitivity of hFOB 1.19 to Zn²⁺. hFOB 1.19 cell line was more sensitive to Zn²⁺ at elevated (restrictive) temperatures. hFOB 1.19 cell line was less sensitive to Zn²⁺ than L929 cell line (both as ZnCl₂ and extracts of alloys). Therefore, the appropriate cultivation conditions of hFOB 1.19 during biomaterial testing should be chosen with caution. Full article

High-throughput RNA-sequencing can determine the impact of nutrients and their combinations on gene transcription levels in osteocytes, and clarify the biological pathways associated with their impact on bone tissues. Previously, we reported that resveratrol (RES) and peonidin-3-O-glucoside (POG) increased osteoblastogenesis, as well as reduced osteoclastogenesis in transgenic teleost fish models. Here, we perform whole-genome transcriptomic profiling of osteoblasts treated with POG or RES to provide a comprehensive understanding of alterations in gene expression and the molecular mechanisms involved. Cultured human fetal osteoblastic hFOB 1.19 cells were treated with the test compounds, and then RNA was used to prepare RNA-seq libraries, that were sequenced using a NovaSeq 6000. Treatment with POG or RES increased osteoblast proliferation and reduced apoptosis. Transcriptomic profiling showed that of the 29,762 genes investigated, 3177 were differentially expressed (1481 upregulated, 1696 downregulated, FDR ≤ 0.05) in POG-treated osteoblasts. In the RES-treated osteoblasts, 2288 genes were differentially expressed (DGEs, 1068 upregulated, 1220 downregulated, FDR ≤ 0.05). Ingenuity^® Pathway Analysis (IPA) of DGEs from RES or POG-treated osteoblasts revealed significant downregulation of the apoptosis, osteoarthritis and HIF1α canonical pathways, and a significant reduction in Rankl mRNA expression. The data suggest that RES and POG have both anabolic and anticlastogenic effects. Full article

A gelatin hydrolysate with a hydrolysis degree of 13.7% was generated using the skin gelatin of chum salmon (Oncorhynchus keta) and papain-catalyzed enzymatic hydrolysis. The results of analysis demonstrated that four amino acids, namely Ala, Gly, Pro, and 4-Hyp, were the most abundant in the obtained gelatin hydrolysate with measured molar percentages ranging from 7.2% to 35.4%; more importantly, the four amino acids accounted for 2/3 of the total measured amino acids. However, two amino acids, Cys and Tyr, were not detected in the generated gelatin hydrolysate. The experimental results indicated that the gelatin hydrolysate at a dose of 50 µg/mL could combat etoposide-induced apoptosis in human fetal osteoblasts (hFOB 1.19 cells), causing a decrease in the total apoptotic cells from 31.6% to 13.6% (via apoptotic prevention) or 13.3% to 11.8% (via apoptotic reversal). Meanwhile, the osteoblasts exposed to the gelatin hydrolysate showed expression changes for 157 genes (expression folds > 1.5-fold), among which JNKK, JNK1, and JNK3 were from the JNK family with a 1.5–2.7-fold downregulated expression. Furthermore, the protein expressions of JNKK, JNK1, JNK3, and Bax in the treated osteoblasts showed a 1.25–1.41 fold down-regulation, whereas JNK2 expression was not detected in the osteoblasts. It is thus suggested that gelatin hydrolysate is rich in the four amino acids and has an in vitro antiapoptotic effect on etoposide-stimulated osteoblasts via mitochondrial-mediated JNKK/JNK(1,3)/Bax downregulation. Full article

Bone defects lead to the structural loss of normal architecture, and those in the field of bone tissue engineering are searching for new alternatives to aid bone regeneration. Dental pulp-mesenchymal stem cells (DP-MSC) could provide a promising alternative to repair bone defects, principally due to their multipotency and capacity to fabricate three-dimensional (3D) spheroids. The present study aimed to characterize the 3D DP-MSC microsphere and the osteogenic differentiation capacity potential cultured by a magnetic levitation system. To achieve this, the 3D DP-MSC microsphere was grown for 7, 14, and 21 days in an osteoinductive medium and compared to 3D human fetal osteoblast (hFOB) microspheres by examining the morphology, proliferation, osteogenesis, and colonization onto PLA fiber spun membrane. Our results showed good cell viability for both 3D microspheres with an average diameter of 350 μm. The osteogenesis examination of the 3D DP-MSC microsphere revealed the lineage commitment, such as the hFOB microsphere, as evidenced by ALP activity, the calcium content, and the expression of osteoblastic markers. Finally, the evaluation of the surface colonization exhibited similar patterns of cell-spreading over the fibrillar membrane. Our study demonstrated the feasibility of forming a 3D DP-MSC microsphere structure and the cell-behavior response as a strategy for the applications of bone tissue guiding. Full article

Prosthetic joint replacement is the most widely used surgical approach to repair large bone defects, although it is often associated with prosthetic joint infection (PJI), caused by biofilm formation. To solve the PJI problem, various approaches have been proposed, including the coating of implantable devices with nanomaterials that exhibit antibacterial activity. Among these, silver nanoparticles (AgNPs) are the most used for biomedical applications, even though their use has been limited by their cytotoxicity. Therefore, several studies have been performed to evaluate the most appropriate AgNPs concentration, size, and shape to avoid cytotoxic effects. Great attention has been focused on Ag nanodendrites, due to their interesting chemical, optical, and biological properties. In this study, we evaluated the biological response of human fetal osteoblastic cells (hFOB) and P. aeruginosa and S. aureus bacteria on fractal silver dendrite substrates produced by silicon-based technology (Si_Ag). In vitro results indicated that hFOB cells cultured for 72 h on the Si_Ag surface display a good cytocompatibility. Investigations using both Gram-positive (S. aureus) and Gram-negative (P. aeruginosa) bacterial strains incubated on Si_Ag for 24 h show a significant decrease in pathogen viability, more evident for P. aeruginosa than for S. aureus. These findings taken together suggest that fractal silver dendrite could represent an eligible nanomaterial for the coating of implantable medical devices. Full article