Osteoblastic Cells Research Articles

The Viability of Osteoblasts against SHED Metabolites and EGCG for Biomaterial Osteogenesis

Background: EGCG, the principal flavonoid found in green tea, exhibits numerous advantageous properties, notably promoting bone regeneration by enhancing the activity of osteoblasts and osteogenic differentiation. Cell-free therapy is an alternative to avoiding the side effects of cell-based therapy. By harnessing the potential of metabolites, SHED combined with EGCG can be a biomaterial to increase osteogenesis. Objectives: This study aims to assess the viability of osteoblast cells when exposed to the combination of SHED metabolites and two concentrations of EGCG, namely 10μM and 50μM. Methods: Osteoblast viability is examined with the 3-(4.5-dimethylthiazole-2-yl)2.5-diphenyl tetrazolium bromide (MTT) assays using an ELISA reader 570nm, and the absorbance value is converted to per cent form. CD50 is a parameter that indicates non-toxicity when the percentage value of living cells is more than 50%. Results: The percentage of living cells exceeded 50%, and statistically significant distinctions were observed among the control media, control cell groups, and the groups exposed to the combination of SHED metabolites and EGCG (p = 0.031). Conclusions: The viability of osteoblast cells exposed to the combination of SHED metabolites and EGCG 10µM, as well as the combination of SHED metabolites and EGCG 50µM, showed no toxicity. The combination of 10µM SHED metabolites and EGCG showed a higher osteoblast cell viability value than the combination of SHED metabolites and EGCG 50 µM.

Novel Antimicrobial Agents Based on Zinc-Doped Hydroxyapatite Loaded with Tetracycline.

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.

Gingerol-zinc complex loaded 3D-printed calcium phosphate for controlled release application.

The therapeutic potential of natural medicines in treating bone disorders is well-established. Modifications in formulation or molecular structure can enhance their efficacy. Gingerol, an osteogenic active compound derived from ginger roots (Zingiber officinale), can form metal ion complexes. Zinc (Zn), a trace element that combats bacterial infections and promotes osteoblast proliferation, can be complexed with gingerol to form a G-Zn+2 complex. This study investigates a porous 3D-printed (3DP) calcium phosphate (CaP) scaffold loaded with the G-Zn+2 complex for drug release and cellular interactions. The scaffold is coated with polycaprolactone (PCL) to control the drug release. Diffusion-mediated kinetics results in 50% releaseof the G-Zn+2 complex over 6weeks. The G-Zn+2 complex demonstrates cytotoxicity against MG-63 osteosarcoma cells, indicated by the formation of apoptotic bodies and ruptured cell morphology on the scaffolds. G-Zn+2 PCL-coated scaffolds show a 1.2 ± 0.1-fold increase in osteoblast cell viability, and an 11.6 ± 0.5% increase in alkaline phosphatase compared to untreated scaffolds. Treated scaffolds also exhibit reduced bacterial colonization against Staphylococcus aureus bacteria, highlighting the antibacterial potential of the G-Zn+2 complex. The functionalized 3DP CaP scaffold with the G-Zn+2 complex shows significant potential for enhancing bone regeneration and preventing infections in low-load-bearing applications.

Comparison of the efficacy of Er,Cr:YSGG laser on oral biofilm removal from implant surfaces with various application times for the treatment of peri-implantitis defects: ex vivo study

PurposeThe major struggle in peri-implantitis therapy is the availability of successful decontamination of the infected implant surface. The main hypothesis of this study was the Er,Cr: YSGG laser decontamination efficacy investigation on the infected implant surfaces with various peri-implantitis defects. The primary objective of this study was to decide the efficacy of Er,Cr:YSGG laser as a decontamination tool at various peri-implantitis simulating defects. The secondary objective was to compare the efficacy of the Er,Cr: YSGG laser on oral biofilm removal between two protocols the first protocol (4 cycles at 2.5 min) and the second protocol (5 cycles at 5 min) at various peri-implantitis simulating defects.Materials and methodsA total of 3 subjects whose plaque biofilms formed in-vivo on twenty-four tested implants were divided into four tested groups. Two native implants were tested as controls.The in vitro defect model was computer‐aided designed and printed into a 3D-printed model with various anulations in peri-implant infrabony defects, which were 15,30,60,and 90 degrees.ResultsBoth Er, Cr: YSGG decontamination protocols at 50 mJ (1.5 W/30 Hz), 50% air, and 40% water were effective at reducing the total implant surface area/ biofilm ratio (%), but the second protocol had a markedly greater reduction in the duration of application (5 cycles at 5 min) than did the first protocol (4 cycles at 2.5 min).ConclusionThe Er, Cr: YSGG laser is an effective decontamination device in various peri-implantitis defects. The second protocol(5 cycles at 5 min) with greater application time and circles is more effective than the first one. The defect angulation influence the decontamination capability in peri-implantitis therapy.Clinical relevance (Scientific rationale for study)Clinicians anticipate that the exploration of suitable therapeutic modalities for peri-implantitis therapy is limited by the obvious heterogeneity of the available evidence in the literature and need for a pre-clinical theoretical basis setup. The major challenges associated with peri-implantitis therapy include the successful decontamination of the infected implant surface, the absence of any damage to the treated implant surface with adequate surface roughness, and the biocompatibility of the implant surface, which allows osteoblastic cells to grow on the treated surface and is the key for successful re-osseointegration. Therefore, these are the expected empirical triads that need to be respected for successful peri-implantitis therapy. Failure of one of the triads represents a peri-implantitis therapeutic failure. The Er, Cr: YSGG laser is regarded as one of the expected devices for achieving the required triad.Trial registration"Efficacy of Er,Cr YSGG Laser in Treatment of Peri-implantitis". ClinicalTrials.gov ID NCT05137821. First Posted date: 30 -11–2021.

Zinc-doped phosphate coatings for enhanced corrosion resistance, antibacterial properties, and biocompatibility of AZ91D Mg alloy

Magnesium and its alloys have been foreseen as promising bone-implant owing to good biocompatibility, high strength, mechanical properties, and biodegradability to replace the conventional bio-metals (Titanium, stainless-steel, Co-Cr alloys) in orthopedic applications. However, high corrosion rate and high degradation rates adversely effects like abrupt evolution of H2 gas and localized alkalization in a physiological environment limit its medical applications. To overcome these issues, phosphate-based surface coating is developed on Mg-alloy to resist the high biodegradation and corrosion rate utilizing magnesium substrate as source of magnesium ion through a single-step hydrothermal process. Controlling the fast corrosion rate and localized alkalization would reduce the antibacterial character of magnesium-based implants therefore, the simple phosphate-based coatings has been induced by doping it with zinc ions due to its physiological tolerance and excellent antibacterial efficacy. The doping of zinc ions may provide a sustained drug-free antibacterial characteristic for potential application in orthopedics. The synthesized phosphate-based coating was characterized using advanced techniques like Scanning Electron Microscopy, X-ray Diffraction, and wettability test, followed by comprehensive electrochemical testing to assess its corrosion behavior. The in vitro immersion test in simulated body fluid (SBF) indicates that deposition of phosphate and zinc doped phosphate coatings reduces the degradation rate consequently minimized the fast dissolution of Mg2+ ions and OH- in physiological environment. Additionally, cytotoxicity and biocompatibility were evaluated using Alamar Blue and live/dead assays on the MC3T3-E1 mouse osteoblast cell line. These assays demonstrated good cell viability, indicating the coatings possess favorable cytocompatibility and support cellular metabolic activity.

Reinforcement of hydroxyapatite bone cement via thin glycerol-citrate polyester infiltration: Microstructural, mechanical and in-vitro evaluation

The development of calcium phosphate cements (CPCs) incorporating biopolymers, such as thermoplastic polylactides, is known as an efficient strategy to enhance the physico-chemical, biological, and mechanical properties of CPCs. However, the use of thermosetting polyol citrates as fillers in CPCs has rarely been reported. Hence, the main goal of the present work was to produce composite CPCs based on tetracalcium phosphate–monetite cement matrix (C) and glycerol-citrate polyester (GCA) and to thoroughly study the effect of GCA addition on microstructural, micro, and nano-mechanical as well as in-vitro cellular properties of final cement composites. The GCA polymer was synthesized by esterification of glycerol with citric acid and coated on the C cement powder matrix at a concentration of 2.5 wt% by infiltration in ethanol solution. Two systems were prepared, by drying the composite powder at 105 and 170 °C (denoted as GCA/C105 and GCA/C170), while the respective composite cements were produced by mixing the powders with the 2 wt% of NaH2PO4 liquid phase. The results showed that the GCA polyester had no significant effect on the hydrolysis and transformation of the original cement matrix, and the nanocrystalline hydroxyapatite (Hap) was found as the main hydration product in all types of cement. The cement setting time decreased slightly with the GCA addition. On the other hand, a significant increase in mechanical properties was reached by introducing the GCA polyester into the CPC matrix, achieving more than a 70 % increase in the compressive strength, and about 50 % and 20 % in micro and nanohardness values compared to that obtained for the original C cement sample. The nanoindentation analysis revealed that the improved compressive strength and hardness were due to the stronger boundaries between the platelet-like hydroxyapatite particles in GCA-doped cements and partially caused by the denser, less porous, and more compact microstructures. The results of in-vitro cytotoxicity testing revealed high proliferation activity of the osteoblastic cells in all cement extracts, providing useful information for designing novel composite cements with the potential to be applied as bone substitute materials.

The comparison of extracellular vesicles with matrix vesicles derived from human vascular smooth muscle versus bone osteoblastic and osteosarcoma cells

The comparison of extracellular vesicles with matrix vesicles derived from human vascular smooth muscle versus bone osteoblastic and osteosarcoma cells

Effects of Different No-Ozone Cold Plasma Treatment Methods on Mouse Osteoblast Proliferation and Differentiation.

Background and Objectives: Enhanced osteoblast differentiation may be leveraged to prevent and treat bone-related diseases such as osteoporosis. No-ozone cold plasma (NCP) treatment is a promising and safe strategy to enhance osteoblast differentiation. Therefore, this study aimed to determine the effectiveness of direct and indirect NCP treatment methods on osteoblast differentiation. Mouse osteoblastic cells (MC3T3-E1) were treated with NCP using different methods, i.e., no NCP treatment (NT group; control), direct NCP treatment (DT group), direct NCP treatment followed by media replacement (MC group), and indirect treatment with NCP-treated media only (PAM group). Materials and Methods: The MC3T3-E1 cells were subsequently assessed for cell proliferation, alkaline phosphatase (ALP) activity, calcium deposition, and ALP and osteocalcin mRNA expression using real-time polymerase chain reaction. Results: Cell proliferation significantly increased in the NCP-treated groups (DT and PAM; MC and PAM) compared to the NT group after 24 h (p < 0.038) and 48 h (p < 0.000). ALP activity was increased in the DT and PAM groups at 1 week (p < 0.115) and in the DT, MC, and PAM groups at 2 weeks (p < 0.000) compared to the NT group. Calcium deposition was higher in the NCP-treated groups than in NT group at 2 and 3 weeks (p < 0.000). ALP mRNA expression peaked in the MC group at 2 weeks compared to the NP group (p < 0.014). Osteocalcin mRNA expression increased in the MC group at 2 weeks (p < 0.000) and was the highest in the PAM group at 3 weeks (p < 0.000). Thus, the effects of direct (DT and MC) and indirect (PAM) treatment varied, with MC direct treatment showing the most significant impact on osteoblast activity. Conclusions: The MC group exhibited enhanced osteoblast differentiation, indicating that direct NCP treatment followed by media replacement is the most effective method for promoting bone formation.

Mn2+ vs Co2+ substitution into β-TCP: Structural details and bone cells response

This work investigated the range of substitution of two biologically relevant ions, namely Mn2+ and Co2+, into the structure of β-tricalcium phosphate, as well as their influence on bone cells response. To this aim, β-TCP was synthesized by solid state reaction in the presence of increasing amount of the substituent ions. The results of the X-ray diffraction analysis reveal that just limited amounts of these ions can enter into the β-TCP structure: 15 at% and 20 at% for cobalt and manganese, respectively. Substitution provokes aggregation of the micrometric particles and reduction of the lattice constants. In particular, the dimension of the c-parameter exhibits a discontinuity at about 10 at% for both cations, although with different trend. Moreover, Rietveld refinement demonstrates a clear preference of both manganese and cobalt for the octahedral site (V). The influence of these ions on cell response was tested on osteoblast, osteoclast and endothelial cells. The results indicate that the presence of manganese promotes a good osteoblast viability, significantly enhances the expression of osteoblast key genes and the angiogenic process of endothelial cells, while inhibiting osteoclast resorption. At variance, osteoblast viability appears reduced in the presence of Co samples, on which osteoblast genes reach higher expression than on β-TCP just in a few cases. On the other hand, the results clearly show that cobalt significantly stimulates the angiogenic process and inhibits osteoclast resorption.

Evolutionary Design of Self-Templated Supramolecular Fibrils Using M13 Bacteriophage for Tissue Engineering.

Biomaterials in nature form hierarchical structures and functions across various length scales through binding and assembly processes. Inspired by nature, we developed hierarchically organized tissue engineering materials through evolutionary screening and self-templating assembly. Leveraging the M13 bacteriophage (phage), we employed an evolutionary selection process against hydroxyapatite (HA) to isolate HA-binding phage (HAPh). The newly discovered phage exhibits a bimodal length, comprising 950 nm and 240 nm, where the synergistic effect of these dual lengths promotes the formation of supramolecular fibrils with periodic banded structures. The assembled HAPh fibrils show the capability of HA mineralization and the directional growth of osteoblast cells. When applied to a dentin surface, it induces the regeneration of dentin-like tissue structures, showcasing its potential applications as a scaffold in tissue engineering. The integration of evolutionary screening and self-templating assembly holds promise for the future development of hierarchically organized tissue engineering materials.

Pro-Osteogenic Effect of the Nutraceutical BlastiMin Complex® in Women with Osteoporosis or Osteopenia: An Open Intervention Clinical Trial.

Osteoporosis is a chronic disease that affects millions of patients worldwide and is characterized by low bone mineral density (BMD) and increased risk of fractures. Notably, natural molecules can increase BMD and exert pro-osteogenic effects. Noteworthily, the nutraceutical BlastiMin Complex® (Mivell, Italy, European Patent Application EP4205733A1) can induce differentiation of human bone marrow mesenchymal stem cells (BM-MSCs) in osteoblasts and can exert in vitro pro-osteogenic and anti-inflammatory effects. Thus, the purpose of this study was to verify the effects of BlastiMin Complex® on bone turnover markers (BTMs) and BMD in patients with senile and postmenopausal osteopenia or osteoporosis. The efficacy of BlastiMin Complex® on BTMs in serum was evaluated through biochemical assays. BMD values were analyzed by dual-energy X-ray absorptiometry (DXA) and Radiofrequency Echographic Multi Spectrometry (R.E.M.S.) techniques, and the SNPs with a role in osteoporosis development were evaluated by PCR. Clinical data obtained after 12 months of treatment showed an increase in bone turnover index, a decrease in C-reactive protein levels, and a remarkable increase in P1NP levels, indicating the induction of osteoblast proliferation and activity in the cohort of 100% female patients recruited for the study. These findings show that the nutraceutical BlastiMin Complex® could be used as an adjuvant in combination with synthetic drugs for the treatment of osteoporosis pathology.

Senescence of skeletal stem cells and their contribution to age-related bone loss

Human aging is linked to bone loss, resulting in bone fragility and an increased risk of fractures. This is primarily due to an age-related decline in the function of bone-forming osteoblastic cells and accelerated cellular senescence within the bone microenvironment. Here, we provide a detailed discussion of the hypothesis that age-related defective bone formation is caused by senescence of skeletal stem cells, as they are the main source of bone forming osteoblastic cells and influence the composition of bone microenvironment. Furthermore, this review discusses potential strategies to target cellular senescence as an emerging approach to treat age-related bone loss.

IL-33 prevents age-related bone loss and memory impairment by suppression of Th17 response: evidence in a d-galactose-induced aging mouse model.

Cytokines are the primary mediators of age-related disorders. The IL-17/IL-10 axis plays a crucial role in bone destruction and neuro-inflammation. Additionally, a new Th2 cytokine-IL-33-has gained attention for its potential implications in aging-associated conditions. However, the involvement of IL-33 in aging-mediated bone loss and memory impairment remains unclear and needs further investigation. This study reveals the impact of IL-33 on various aspects of the immune system, bone health, and neural functions. To induce senescence, we used d-galactose for its convenience and fewer side effects. The experimental design involved treating 20-week-old C57BL/6J mice with d-galactose subcutaneously for 10weeks to induce aging-like effects. Thereafter, IL-33 recombinant protein was administered intraperitoneally for 15days to evaluate its impact on various immune, skeletal, and neural parameters. The results demonstrated that d-galactose-induced aging led to bone loss and compromised osteogenic parameters, accompanied by increased oxidative stress and neurodegeneration in specific brain regions. Behavioral activities were also affected. However, supplementation with IL-33 mitigated these effects, elevating osteogenic parameters and reducing senescence markers in osteoblast cells in an aging mouse model and exerted neuroprotective potential. Notably d-galactose-induced aging was characterized by high bone turnover, reflected by altered serum levels of CTX, PTH, beta-galactosidase, and P1NP. IL-33 treatment attenuated these effects, suggesting its role in regulating bone metabolism. Furthermore, d-galactose-induced aging was associated with increased differentiation of Th17 cells and upregulation of associated markers, such as STAT-3 and ROR-γt, while downregulating Foxp3, which antagonizes Th17 cell differentiation. IL-33 treatment countered these effects by suppressing Th17 cell differentiation and promoting IL-10-producing T-regulatory cells. Overall, these findings provide insights into the potential therapeutic implications of IL-33 in addressing aging-induced bone loss and memory impairment.

Bromelain-facilitated GNPs: A strategic innate anticancer delivery system for methotrexate into osteosarcoma cells

Osteosarcoma is the most common malignant bone tumor in children and adolescents during rapid bone growth. Traditional chemotherapy has markedly improved patient outcomes but suffers from nonspecific side effects due to nonspecific drug distribution, highlighting the need for targeted therapies. Methotrexate, although effective, poses toxicity concerns and prompts exploration into advanced drug delivery systems like nanoparticles. Here, we developed a novel method of synthesizing gold nanoparticles using bromelain (G-BNPs) as a reducing and capping agent, and we conjugated G-BNPs with methotrexate (G-B-MNPs) via 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide/N-hydroxysuccinimide (EDC/NHS) as crosslinker. Our meticulous characterization, which included UV–visible spectroscopy, Transmission electron microscopy (TEM), Dynamic light scattering (DLS), Zeta potential, and Fourier-transform infrared spectroscopy (FTIR) analyses confirmed the successful fabrication and drug conjugation of the G-BNPs. We then evaluated the cytotoxicity of the G-BNPs (sized ∼ 21 nm) and G-B-MNPs (sized ∼ 24 nm) on the osteosarcoma cell lines, Saos-2 and MG-63, as well as the non-cancerous primary osteoblast cells. Our findings revealed that G-B-MNPs significantly increased the cytotoxic impact on osteosarcoma cells compared to methotrexate alone while showing minimal toxicity toward primary osteoblast cells. Importantly, using 4`,6-diamidino-2-phenylindole(DAPI), we demonstrated that G-B-MNPs were delivered directly into the nucleus of the cells without encountering lysosomes. G-B-MNPs can be considered a potential drug delivery method for osteosarcoma treatment by improving effectiveness and reducing adverse effects, thereby enhancing patient prognosis.

Bone cells influence the degradation interface of pure Mg and WE43 materials: Insights from multimodal in vitro analysis

In this study, the interaction of pure Mg and WE43 alloy under the presence of osteoblast (OB) and osteoclast (OC) cells and their influence on the degradation of materials have been deeply analyzed. Since OB and OC interaction has an important role in bone remodeling, we examined the surface morphology and dynamic changes in the chemical composition and thickness of the corrosion layers formed on pure Mg and WE43 alloy by direct monoculture and coculture of pre-differentiated OB and OC cells in vitro. Electrochemical techniques examined the corrosion performance. The corrosion products were characterized using a combination of the focused ion beam (FIB), scanning electron microscopy (SEM), and energy-dispersive X-ray spectroscopy (EDX). Cell viability and morphology were assessed by fluorescent microscopy and SEM. Our findings demonstrate cell spread and attachment variations, which differ depending on the Mg substrates. It was clearly shown that cell culture groups delayed degradation processes with the lowest corrosion rate observed in the presence of OBOC coculture for the WE43 substrate. Ca-P enrichment was observed in the outer-middle region of the corrosion layer but only after 7 days of OBOC coculture on WE43 and after 14 days on the pure Mg specimens. Statement of significanceMagnesium metallic materials that can degrade over time provide distinct opportunities for orthopedic application. However, there is still a lack, especially in elucidating cell-material interface characterization. This study investigated the influence of osteoblast-osteoclast coculture in direct Mg-material contact. Our findings demonstrated that pre-differentiated osteoblasts and osteoclasts cocultured on Mg substrates influenced the chemistry of the corrosion layers. The cell spread and attachment were Mg substrate-dependent. The findings of coculturing bone cells directly on Mg materials within an in vitro model provide an effective approach for studying the dynamic degradation processes of Mg alloys while also elucidating cell behavior and their potential contribution to the degradation of these alloys.

Effects of photobiomodulation on different phases of in vitro osteogenesis.

The present study aimed to evaluate the effect of photobiomodulation therapy (PBM) on different stages of osteogenesis in vitro. For this, osteoblastic-like cells (Saos-2 cell lineage) were irradiated in two different periods: during the Proliferation phase (PP; from the second to the fourth day) and during the Differentiation phase (DP; from the seventh to the ninth day). The energy density used in the study was 1.5J/ cm2. The following parameters were evaluated: 1) quantification ofcollagen type 1 (COL 1), osteopontin (OPN), and bone morphogenetic protein 2 (BMP-2); 2) quantification of alkaline phosphatase (ALP) activity; and 3) quantification of extracellular matrix (ECM)mineralization. Non-irradiated cultures were used as controls. The data were analyzed using the Student's t-test or one-way ANOVA, considering a significance level of 5%. The results indicated that COL 1 and BMP-2 quantification was higher in Saos-2 irradiated during the DP in relation to the control group at day 10 (p < 0.05). No differences were observed for other comparisons at this time point(p>0.05).OPN expression was greater in PP compared with the other experimental groups at day 10 (p < 0.05). Irradiation did not affect ALP activity in Saos-2 regardless of the exposure phase and the time point evaluated (p > 0.05). At day 14, ECMmineralization was higher in Saos-2 cultures irradiated during the DP in relation to the PP (p < 0.05). In conclusion, the results suggested that the effects of PBM on osteoblastic cells may be influenced by the stage of cell differentiation.

Biological Effects of Double-Layered Hydroxyapatite and Zirconium Oxide Depositions on Titanium Surfaces.

This study aimed to confirm the synergy effect of these two materials by evaluating osteoblast and antibacterial activity by applying a double-layered hydroxyapatite(HA) zirconium oxide(ZrO2) coating to titanium. The specimens used in this study were divided into four groups: a control group (polished titanium; group T) and three experimental groups: Group TH (RF magnetron sputtered HA deposited titanium), Group Z (ZrO2 ALD deposited titanium), and Group ZH (RF magnetron sputtered HA and ZrO2 ALD deposited titanium). The adhesion of Streptococcus mutans (S.mutans) to the surface was assessed using a crystal violet assay. The adhesion, proliferation, and differentiation of MC3T3-E1 cells, a mouse osteoblastic cell line, were assessed through a WST-8 assay and ALP assay. Group Z showed a decrease in the adhesion of S. mutans (p < 0.05) and an improvement in osteoblastic viability (p < 0.0083). Group TH and ZH showed a decrease in adhesion of S. mutans (p < 0.05) and an increase in osteoblastic cell proliferation and cell differentiation (p < 0.0083). Group ZH exhibited the highest antibacterial and osteoblastic differentiation. In conclusion double-layered HA and ZrO2 deposited on titanium were shown to be more effective in inhibiting the adhesion of S. mutans, which induced biofilm formation, and increasing osteoblastic differentiation involved in osseointegration by the synergistic effect of the two materials.

Production of platelet-rich plasma (PRP)-enriched scaffolds for bone tissue regeneration with 3D printing technology

Bone disorders signify diverse abnormalities in the structure, development, and functions of bone tissue in the human body, with a significant correlation to ageing, insufficient physical activity, and escalating obesity. Recent advancements in bone tissue engineering aim to enhance bone tissue formation through the use of biomaterials, growth factors, and cells. The present study focuses on the fabrication and characterisation of scaffolds with a composition of gelatin (GEL) / sodium alginate (SA) / hydroxyapatite (HA) / platelet-rich plasma (PRP) using the 3D printing process. The inclusion of PRP, derived from blood, is of particular interest due to its potential to enhance bone regeneration through various growth factors. Scanning electron microscope (SEM) analysis revealed average pore sizes ranging from 481.50 ± 7.65 to 623.96 ± 11.54 µm. SEM images also showed that scaffold surfaces became smooth as the concentration of PRP increased. The mechanical test results demonstrated that as the PRP increased, the compressive strength decreased. When the swelling and degradation behaviours of scaffolds were examined, it was observed that GEL/SA/HA/3PRP scaffolds exhibited approximately 200 % swelling capability until the 4th day. GEL/SA/HA scaffolds showed a degradation behaviour about 70 % higher compared to other groups. A controlled release profile of PRP was maintained up to the 144th, 216th, and 240th hours from the scaffolds. According to the highest correlation coefficients (R2) in the release kinetics of scaffolds, GEL/SA/HA/0.5PRP and GEL/SA/HA/1PRP scaffolds were explained by the first-order model. In contrast, the GEL/SA/HA/3PRP scaffold was described using the Korsmeyer–Peppas model. The MTT analysis conducted with osteoblast cells showed that scaffolds did not demonstrate any toxic effects and facilitated cell adhesion by inducing the formation of extensions. These findings underscore the potential of PRP-incorporated GEL/SA/HA composites as a promising approach for bone tissue engineering, offering significant advancements in the treatment of bone disorders. This could lead to more effective treatments for bone disorders and injuries, reducing the need for more invasive procedures and improving patient recovery times.

Zinc phosphate glass microspheres promoted mineralization and expression of BMP2 in MC3T3-E1 cells.

Degradable phosphate glasses have shown favorable properties for tissue engineering. By changing the composition of the glasses, the degradation rate, and ion release are controllable. Zinc oxide can function as a glass network modifier and has been shown to play a positive role in bone formation. Also, phosphate glasses can easily be processed into microspheres, which can be used as microcarriers. This study aims to develop zinc phosphate glasses microspheres and explore the optimized size and composition for applications in bone tissue engineering. Zinc-titanium-calcium-sodium phosphate glasses with 0, 1, 3, 5, or 10 mol % zinc oxide were prepared and processed into microspheres. The smaller microspheres ranged in size from 50 to 106 μm, while the larger ones ranged from 106 to 150 μm. The characteristics of glasses were examined. The osteoblastic cell line MC3T3-E1 was cultured on the surface of microspheres and the cell viability was examined. To evaluate osteogenic differentiation, Alizarin Red S staining, quantitative reverse transcription polymerase chain reaction, and western blot analysis were performed after 14 days. Different sizes of zinc phosphate glass microspheres were successfully made. The glass microspheres with <10 mol % zinc oxide were able to support the adhesion and proliferation of MC3T3-E1 cell lines. The relative gene expression of BMP2 was significantly upregulated in the smaller glass microspheres containing 3 mol % zinc oxide (26-fold, p < .001) and both sizes of microspheres containing 5 mol % zinc oxide (smaller: 27-fold, p < .001; larger: 35-fold, p < .001). Additionally, cluster formation was observed in glass microspheres after 14 days, and the mineralization of MC3T3-E1 cell lines was promoted. Based on these findings, the glass microspheres containing 3-5 mol % of zinc oxide can promote osteogenic differentiation for MC3T3-E1 cells.

Mussel shell-derived biogenic hydroxyapatite as reinforcement on chitosan-loaded gentamicin composite for antibacterial activity and bone regeneration

In this study, hydroxyapatite (HAp) was synthesized from natural biowaste materials, specifically mussel shells, and combined with chitosan (CS) and gentamicin sulfate antibiotic (GA) using an in-situ method. The resulting composite material, designated HAp/CS-GA, has its physicochemical and structural properties characterized by Fourier transform infrared spectroscopy (FTIR) analysis. The structure was confirmed by X-ray diffraction (XRD) analysis. Additionally, field emission scanning electron microscopy (FE-SEM) equipped with the energy dispersive X-ray spectroscopic (EDX) technique was used to determine the surface topography and main components. The composite of HAp/CS-GA was analyzed using a drug release profile by UV–visible spectroscopy (UV–Vis). The fabricated composites antimicrobial behavior was examined against bone infection-causing Gram-positive and Gram-negative bacteria, showing potential activity against Klebsiella pneumoniae, Pseudomonas aeruginosa, and Staphylococcus aureus compared to Escherichia coli, respectively. Simultaneously, the cytotoxicity of the composite was evaluated by MTT assay using an MG-63 osteoblast-like cell line that exhibited no toxicity in the prepared composite. After a 24 h incubation period, the MG-63 cells on the HAp/CS-GA composite showed good proliferation, according to Hoechst 33258 fluorescence staining results. The results suggested that the composite had excellent biocompatibility and antibacterial activity and enhanced the osteoblast cell proliferation. Therefore, the designed HAp/CS-GA composite would be a promising candidate for bone tissue engineering.