Cultures Of Osteoblast-like Cells Research Articles

Effects of Daidzein, Tempeh, and a Probiotic Digested in an Artificial Gastrointestinal Tract on Calcium Deposition in Human Osteoblast-like Saos-2 Cells.

Adequate calcium intake is crucial for the prevention and treatment of bone-related issues. Developing a nutritional source of readily bioavailable calcium is particularly significant for individuals deficient in this essential element and at risk of developing osteoporosis. This research aimed to evaluate the impact of tempeh (T), daidzein (D), and Lactobacillus acidophilus (LA) within a simulated intestinal environment consisting of Caco-2 epithelial and Saos-2 cells, focusing on their implications for bone mineralization mechanisms. In the initial phase, calcium bioaccessibility from calcium citrate (CaCt), LA, D, the daidzein combination D-CaCt-LA (D1:1:1), and the tempeh combination T-CaCt-LA (T1:1:1) was assessed through digestion simulation. The calcium content of both untreated and digested samples was determined using atomic absorption spectrometry (AAS). In the subsequent stage, the digested samples were used to induce intestinal absorption in differentiated enterocyte-like Caco-2 cells. The permeable fractions were then evaluated in a culture of osteoblast-like Saos-2 cells. Preliminary cellular experiments employed the MTT assay to assess cytotoxicity. The results indicated that the analyzed products did not influence the deposition of extracellular calcium in Saos-2 cells cultured without mineralization stimulators. The combined formulations of permeable fractions of digested CaCt, LA, D, and T demonstrated the capacity to enhance the proliferation of Saos-2 cells. In Saos-2 cells, D, D1:1:1, and LA showed no discernible impact on intracellular calcium accumulation, whereas T and T1:1:1 reduced the calcium deposits. Additionally, mRNA transcripts and alkaline phosphatase (ALP) activity levels in Saos-2 cells cultured without mineralization induction were unaffected by the analyzed products. An examination of the products revealed no discernible effect on ALP activity or mRNA expression during Saos-2 cell differentiation. Our findings suggest that tempeh, daidzein, and L. acidophilus did not positively impact cellular calcium deposition in Saos-2 cells. However, tempeh, daidzein and its combination, and L. acidophilus might enhance the process of osteogenic differentiation in Saos-2 cells. Nevertheless, this study did not identify any synergistic impact on calcium deposition and the process of osteogenic differentiation in Saos-2 cells of isoflavones and probiotics.

Osteoblast-Derived Matrix Vesicles Exhibit Exosomal Traits and a Unique Subset of microRNA: Their Caveolae-Dependent Endocytosis Results in Reduced Osteogenic Differentiation.

Matrix vesicles (MVs) are nano-sized extracellular vesicles that are anchored in the extracellular matrix (ECM). In addition to playing a role in biomineralization, osteoblast-derived MVs were recently suggested to have regulatory duties. The aims of this study were to establish the characteristics of osteoblast-derived MVs in the context of extracellular vesicles like exosomes, assess their role in modulating osteoblast differentiation, and examine their mechanism of uptake. MVs were isolated from the ECM of MG63 human osteoblast-like cell cultures and characterized via enzyme activity, transmission electron microscopy, nanoparticle tracking analysis, Western blot, and small RNA sequencing. Osteoblasts were treated with MVs from two different culture conditions (growth media [GM]; osteogenic media [OM]) to evaluate their effects on the differentiation and production of inflammatory markers and on macrophage polarization. MV endocytosis was assessed using a lipophilic, fluorescent dye and confocal microscopy with the role of caveolae determined using methyl-β-cyclodextrin. MVs exhibited a four-fold enrichment in alkaline phosphatase specific activity compared to plasma membranes; were 50-150 nm in diameter; possessed exosomal markers CD63, CD81, and CD9 and endosomal markers ALIX, TSG101, and HSP70; and were selectively enriched in microRNA linked to an anti-osteogenic effect and to M2 macrophage polarization. Treatment with GM or OM MVs decreased osteoblast differentiation. Osteoblasts endocytosed MVs using a mechanism that involves caveolae. These results support the hypothesis that osteoblasts produce MVs that participate in the regulation of osteogenesis.

PD03-01 FGF-23 PRODUCTION COUPLING WITH ALDOSTERONE AND PREDICTING CLINICAL OUTCOMES IN PRIMARY ALDOSTERONISM

You have accessJournal of UrologyCME1 Apr 2023PD03-01 FGF-23 PRODUCTION COUPLING WITH ALDOSTERONE AND PREDICTING CLINICAL OUTCOMES IN PRIMARY ALDOSTERONISM Jeff Chueh, Shuo-Meng Wang, VinCent Wu, and Kang-Yung Peng Jeff ChuehJeff Chueh More articles by this author , Shuo-Meng WangShuo-Meng Wang More articles by this author , VinCent WuVinCent Wu More articles by this author , and Kang-Yung PengKang-Yung Peng More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000003220.01AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: Higher fibroblast growth factor-23 (FGF-23) levels are associated with activating renin-angiotensin-aldosterone system (RAAS), more volume overload, and hypertension. The roles of FGF-23 in primary aldosteronism (PA) haven't been studied. METHODS: Unilateral PA (uPA) patients who underwent adrenalectomy were enrolled and followed for their clinical outcomes from Aug 15 to Jan 19. The FGF-23 expression in a rat osteoblast-like cell culture was evaluated after the addition of aldosterone. RESULTS: A total of 121 uPA patients & 69 matched essential hypertension (EH) controls were enrolled. The log [carboxyl-terminal FGF-23 (cFGF-23)] level was higher in uPA patients (p=0.001).] In Cox proportional hazard model, the preoperative Log [plasma cFGF-23]>1.98 was a risk factor for either all-cause mortality and cardiovascular or kidney events (HR, 4.01, p=0.038). Plasma cFGF-23 levels were increased in a nonlinear manner re: plasma aldosterone concentration (PAC). Log [plasma cFGF-23]>1.75 independently predicted post-op remission of hypertension (odds ratio (OR), 3.8, p=0.004). The plasma total FGF-23 level was significantly higher in PA patients (p<0.001), and it was significantly reduced post-op (p<0.001). When osteoblast-like UMR-106 cells were cultured with aldosterone, the cFGF-23 concentration in the culture medium was elevated (p=0.028), but it was abrogated after family with sequence similarity 20, member C (FAM20c) short hairpin RNA was added (p= 0.659). Docking experiments showed that aldosterone may bind to the potential drug binding pocket of FAM20c (Pdb: 5YH3; Figure 1A, B). CONCLUSIONS: Compared with EH controls, uPA patients had higher cFGF-23 levels--associated with a higher risk of cardiovascular composite outcomes, though such higher cFGF-23 levels predicted hypertension remission after adrenalectomy. Their total FGF-23 levels were significantly attenuated after adrenalectomy. An increase in cFGF-23 production among the uPA patients was secondary to increased cleavage of intact FGF-23 evidenced by higher FAM20c activity modulated by aldosterone. Source of Funding: Taiwan (NSC 101-2314-B-002-132-MY3, NSC100-2314-B-002-119, NSC 101-2314-B-002-085-MY3, MOST 104-2314-B-002 -125 -MY3; 111-2314-B-002-232-MY3) and NTUH 100-N1776, 101-M1953, 102-S2097 © 2023 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 209Issue Supplement 4April 2023Page: e74 Advertisement Copyright & Permissions© 2023 by American Urological Association Education and Research, Inc.MetricsAuthor Information Jeff Chueh More articles by this author Shuo-Meng Wang More articles by this author VinCent Wu More articles by this author Kang-Yung Peng More articles by this author Expand All Advertisement PDF downloadLoading ...

Effect of tantalum interlayer on hydroxyapatite biointerface for orthopedic applications

Various attempts have been made during recent decades to improve the functionality, biocompatibility and osseointegration of metallic implants. With technological advancement, modifying superficial properties of orthopedic implants has undergone great evolution. Current efforts of scientists and the orthopedic industry have focused on modifying surface properties to achieve a specific biointerface that can reduce patient's healing time and prevent aseptic loosening. This study investigates the contribution of a protective tantalum interlayer deposited on titanium substrates on the growth of hydroxyapatite (HAp) crystals with hydrothermal synthesis method. We investigated the morphology and structure of the produced coatings, as well as in vitro bioactivity in simulated body fluid and biocompatibility in osteoblast-like cell culture. The hydrothermal method allows to synthesize coating with micro-sized HAp crystallites that exhibit rod-like hexagonal shape. The diameter of HAp crystallites may be controlled and driven by nature of the interlayer (tantalum, tantalum oxide or titanium oxide) previously formed on the titanium substrate. Coatings with smaller diameters of HAp crystals (between 0,3 μm and 0,6 μm) synthesized on tantalum-based films were found to be more favorable for cell viability and attachment. High bioactivity and positive effect on the osteoblast-like cells of the HAp biointerface with a tantalum intermediate layer was observed.

Bioactivity evaluation of printable calcium polyphosphate/alginoplast cement for bone tissue engineering; In vitro study

By developing the three-dimensional (3D) printing technology and patient-specific medicine, 3D calcium phosphate cement, designed for every complex defect, remains a new challenge. This study developed a novel printable calcium polyphosphate (CPP) powder by adding alginoplast (ALG) to fabricated 3D cement using low-temperature binder jetting-based 3D printing without any sintering. The powder flowability and 3D cement biological properties were analyzed using Freeman technology, immersion in simulated body fluid (SBF) solution, and MG63 human osteoblast-like cells culture. According to the Freeman technology, adding 10 wt% of ALG in CPP showed better powder flowability. Together with the SEM, XRD, and FTIR results, the high bioactivity potential of 3D cement containing ALG was recorded by the cauliflower-like carbonate apatite formation on the 3D cement surface after 21 days of immersion. Furthermore, the 3D cement porous microstructure provided a cell attachment. The number of cells showed the 3D cement biocompatibility increased by increasing the cell culture time in 3D cement extraction through the MTT result.

Wettability, Corrosion Resistance, and Osteoblast Response to Reduced Graphene Oxide on CoCr Functionalized with Hyaluronic Acid

The durability of metal–metal prostheses depends on achieving a higher degree of lubrication. The beneficial effect of hyaluronic acid (HA) on the friction and wear of both natural and artificial joints has been reported. For this purpose, graphene oxide layers have been electrochemically reduced on CoCr surfaces (CoCrErGO) and subsequently functionalized with HA (CoCrErGOHA). These layers have been evaluated from the point of view of wettability and corrosion resistance in a physiological medium containing HA. The wettability was analyzed by contact angle measurements in phosphate buffer saline-hyaluronic acid (PBS-HA) solution. The corrosion behavior of functionalized CoCr surfaces was studied with electrochemical measurements. Biocompatibility, cytotoxicity, and expression of proteins related to wound healing and repair were studied in osteoblast-like MC3T3-E1 cell cultures. All of the reported results suggest that HA-functionalized CoCr surfaces, through ErGO layers in HA-containing media, exhibit higher hydrophilicity and better corrosion resistance. Related to this increase in wettability was the increase in the expressions of vimentin and ICAM-1, which favored the growth and adhesion of osteoblasts. Therefore, it is a promising material for consideration in trauma applications, with improved properties in terms of wettability for promoting the adhesion and growth of osteoblasts, which is desirable in implanted materials used for bone repair.

Mechanical and biological properties of atmospheric plasma-sprayed carbon nanotube-reinforced tantalum pentoxide composite coatings on Ti6Al4V alloy

Ta2O5 coatings containing 0, 3, 5 and 7 vol% multi-walled carbon nanotubes (CNTs) were deposited on Ti6Al4V substrates by atmospheric plasma spraying (APS). The addition of CNTs had no obvious effect on the surface morphology and roughness of the coatings, but increased the coating porosity. The coatings consisted primarily of β-Ta2O5 phase, a small amount of α-Ta2O5, and minor C peaks corresponding to the CNTs. As the CNT content increased, the elastic modulus (E) and indentation fracture toughness also increased. However, the higher CNT content increased the coating porosity and reduced the microhardness. Following immersion in simulated body fluid (SBF) for 14 days, the surfaces of all the coatings were completely covered with hemispherical bone-like apatite. Furthermore, after incubation in osteoblast-like osteosarcoma MG-63 cell culture for 7 days, all of the coatings showed excellent cell attachment, growth and spreading. Overall, the present results show that the addition of CNTs to Ta2O5 raw powder improves the indentation fracture toughness of the resulting coatings without degrading their biological properties.

Mechanical-Stress-Related Epigenetic Regulation of ZIC1 Transcription Factor in the Etiology of Postmenopausal Osteoporosis.

Mechanical loading exerts a profound influence on bone density and architecture, but the exact mechanism is unknown. Our study shows that expression of the neurological transcriptional factor zinc finger of the cerebellum 1 (ZIC1) is markedly increased in trabecular bone biopsies in the lumbar spine compared with the iliac crest, skeletal sites of high and low mechanical stress, respectively. Human trabecular bone transcriptome analyses revealed a strong association between ZIC1 mRNA levels and gene transcripts characteristically associated with osteoblasts, osteocytes and osteoclasts. This supposition is supported by higher ZIC1 expression in iliac bone biopsies from postmenopausal women with osteoporosis compared with age-matched control subjects, as well as strongly significant inverse correlation between ZIC1 mRNA levels and BMI-adjusted bone mineral density (BMD) (Z-score). ZIC1 promoter methylation was decreased in mechanically loaded vertebral bone compared to unloaded normal iliac bone, and its mRNA levels correlated inversely with ZIC1 promoter methylation, thus linking mechanical stress to epigenetic control of gene expression. The findings were corroborated in cultures of rat osteoblast progenitors and osteoblast-like cells. This study demonstrates for the first time how skeletal epigenetic changes that are affected by mechanical forces give rise to marked alteration in bone cell transcriptional activity and translate to human bone pathophysiology.

Low concentrations of caffeic acid phenethyl ester stimulate osteogenesis in vitro

ObjectiveThe aim of this study was to evaluate the effects of caffeic acid phenethyl ester (CAPE) on osteoblast-like cell cultures (SAOS-2). MethodsSAOS-2 were exposed to CAPE at 1 nM, 10 nM, 100 nM, 1 μM, and 10 μM. Non-exposed cultures were used as control. The following parameters were assayed: 1) cell viability at 1, 3, and 7 days; 2) alkaline phosphatase (ALP) activity at 5 and 10 days; 3) matrix mineralization at 14 days; and 4) Runt-related transcription factor 2 (RUNX2), ALP, osteopontin (SPP1), and osteocalcin (BGLAP) gene expression at 5 and 10 days. The data were analyzed by ANOVA two-way or Kruskal-Wallis (α = 5%). ResultsAt day 1, cell viability was similar among all groups (p > 0.05). At days 3 and 7, cultures exposed to CAPE at 10 μM exhibited a significant reduction in cell viability compared with the others groups (p < 0.05). At day 5, ALP activity was similar among all experimental groups; at day 10, however, the stain intensity was higher in cultures exposed to CAPE at 100 nM and 10 nM in comparison with the other groups (p < 0.05). At days 5 and 10, RUNX2, ALP, SPP1, and BGLAP gene expression was greater in cultures exposed to CAPE in comparison with the control (p < 0.05). At day 14, matrix mineralization was similar in cultures exposed to CAPE at 1 nM and 10 nM (p > 0.05), but superior to those ones observed in the other experimental groups (p < 0.05). ConclusionCAPE at low concentrations can positively module the osteogenesis in vitro.

Magnetic 3D scaffolds for tissue engineering applications: bioactive glass (45S5) coated with iron-loaded hydroxyapatite nanoparticles

Magnetic 45S5 bioactive glass (BG) based scaffolds covered with iron-loaded hydroxyapatite (Fe-HA-BG) nanoparticles were obtained and its cytotoxicity investigated. Fe-HA nanoparticles were synthesized by a wet chemical method involving the simultaneous addition of Fe2+/Fe3+ ions. BG based scaffolds were prepared by the foam replica procedure and covered with Fe-HA by dip-coating. Fe-HA-BG magnetic saturation values of 0.049 emu g−1 and a very low remanent magnetization of 0.01 emu g−1 were observed. The mineralization assay in simulated body fluid following Kokubo’s protocol indicated that Fe-HA-BG scaffolds exhibited improved hydroxyapatite formation in comparison to uncoated scaffolds at shorter immersion times. The biocompatibility of the material in vitro was assessed using human osteoblast-like MG-63 cell cultures and mouse bone marrow-derived stroma cell line ST-2. Overall, the results herein discussed suggest that magnetic Fe-HA coatings seem to enhance the biological performance of 45S5 BG based scaffolds. Thus, this magnetic Fe-HA coated scaffold is an interesting system for bone tissue engineering applications and warrant further investigation.

3D Bioprinted Bacteriostatic Hyperelastic Bone Scaffold for Damage-Specific Bone Regeneration.

Current strategies for regeneration of large bone fractures yield limited clinical success mainly due to poor integration and healing. Multidisciplinary approaches in design and development of functional tissue engineered scaffolds are required to overcome these translational challenges. Here, a new generation of hyperelastic bone (HB) implants, loaded with superparamagnetic iron oxide nanoparticles (SPIONs), are 3D bioprinted and their regenerative effect on large non-healing bone fractures is studied. Scaffolds are bioprinted with the geometry that closely correspond to that of the bone defect, using an osteoconductive, highly elastic, surgically friendly bioink mainly composed of hydroxyapatite. Incorporation of SPIONs into HB bioink results in enhanced bacteriostatic properties of bone grafts while exhibiting no cytotoxicity. In vitro culture of mouse embryonic cells and human osteoblast-like cells remain viable and functional up to 14 days on printed HB scaffolds. Implantation of damage-specific bioprinted constructs into a rat model of femoral bone defect demonstrates significant regenerative effect over the 2-week time course. While no infection, immune rejection, or fibrotic encapsulation is observed, HB grafts show rapid integration with host tissue, ossification, and growth of new bone. These results suggest a great translational potential for 3D bioprinted HB scaffolds, laden with functional nanoparticles, for hard tissue engineering applications.

Dose- and time-dependent effects of taxifolin on viability and mineralization markers of osteoblast-like cells

The current study evaluated the effects of taxifolin treatments on the viability of osteoblast-like cells, and on the expression of early mineralization markers, as part of the ongoing search for new endodontic materials able to induce periapical healing without causing cytotoxicity. Saos-2 osteoblast-like cells were exposed to different concentrations of taxifolin (5 and 10 µM), applied as pretreatments either for 24h and 72h, or continuously throughout the experimental protocol. Cell viability using the methylthiazole tetrazolium (MTT) assay, alkaline phosphatase activity using thymolphthalein monophosphate assays, deposition of mineralized nodules using alizarin red staining, and expression of ALP and COL-1 by qPCR were determined after 6 and 13 days of treatment. The data were analyzed statistically (p<0.05). Taxifolin was not cytotoxic in the concentrations tested. Pretreatments with taxifolin for 24h and 72h at 10 µM stimulated ALP activity, and increased mineralized nodule deposition by Saos-2 cells. Continuous treatment with taxifolin was not effective in stimulating ALP activity and mineralization. ALP and COL-1 gene expression increased with taxifolin pretreatments, since the highest mRNA levels were observed after 72h of pretreatment with taxifolin at 10 µM on day 13. In conclusion, taxifolin was cytocompatible, and induced mineralization markers when applied for short periods in osteoblast-like cell culture.

Photobiomodulation of human osteoblast-like cells invitro by low-intensity-pulsed LED light.

Visible light irradiation is an emerging area in regenerative medicine research. We hypothesized that low‐intensity‐pulsed LED light irradiance may exert photobiomodulatory effects on cultured osteoblast‐like cells. To test this hypothesis, we investigated cell proliferation and markers of cell maturation and metabolic activity following pulsed LED irradiance. Monolayer explant cultures of human osteoblast‐like cells were exposed four times in 24‐h intervals to 2 min of pulsed white LED irradiance of 2.4–2.5 mW·cm−2 and its different spectra of 0.2–0.5 mW·cm−2 (frequency range of 10–40 Hz). Cell proliferation was estimated from microscopic cell counting and cell death by lactate dehydrogenase activity in culture media (measured by a colorimetric method). The early markers of osteoblast maturation and metabolic activity, that is, cellular alkaline phosphatase activity and osteocalcin content, were measured using a colorimetric method and ELISA, respectively. Irradiance of 40 Hz caused the highest increase in cell number (P < 0.01). Osteocalcin content in cells decreased following 40 Hz and 10 Hz irradiance (P < 0.05). The 40 Hz blue range irradiance (diffuse transmittance 420–580 nm, maximal cell irradiance 0.5 mW·cm−2) caused a decrease in alkaline phosphatase cellular activity (P < 0.001) and an increase in media osteocalcin content (P < 0.05). The 40 Hz green range (diffuse transmittance 560–650 nm, maximal cell irradiance 0.4 mW·cm−2) irradiance caused an increase in the number of cells and in cell death. In summary, pulsed (40 Hz) white light irradiance has photomodulatory effects, with its green range spectrum affecting cell proliferation and cell death, and its blue range spectrum affecting cellular maturation and metabolism. The results indicate a low‐intensity threshold of photobiomodulation of osteoblast‐like cells in vitro.

Baghdadite/Polycaprolactone nanocomposite scaffolds: preparation, characterisation, and in vitro biological responses of human osteoblast-like cells (Saos-2 cell line)

ABSTRACT In the present research, porous composite scaffolds consisting of baghdadite scaffolds with polycaprolactone (PCL) coating, were fabricated via combining polyurethane sponge replication method and dip-coating processes and characterised by x-ray diffraction (XRD), scanning electron microscope (SEM), and porosity and compressive strength measurements. MTT assay and cell attachment were carried out via human osteoblast-like cell culture on the scaffolds. Results showed that the scaffolds possessed porous structures with porosity about 70-83%, and compressive strengths 0.18–1.31 MPa. Polycaprolactone can affect a significant role in improving the mechanical properties of baghdadite scaffold. It could enhance the compressive strength of baghdadite scaffold from 0.18 to 0.41 MPa while its porous structure is maintained. The evaluation of cell viability and attachment, verified no cytotoxicity, and good attachment and proliferation of cells on the scaffolds. To summarise, baghdadite–PCL composite scaffold with a porous structure, great mechanical properties and excellent biological responses is a promising candidate for bone tissue engineering.

Bioactive glass (45S5)-based 3D scaffolds coated with magnesium and zinc-loaded hydroxyapatite nanoparticles for tissue engineering applications

Bioactive glass (BG)-based scaffolds of 45S5 composition covered with hydroxyapatite nanoparticles loaded with Mg2+, Zn2+ and, both Mg2+ and Zn2+ ions, were developed and tested as materials for tissue engineering applications. The scaffolds were prepared by the foam replica technique and mono- and bi-metal loaded and unloaded hydroxyapatite nanoparticles (HA, Zn-HA, Mg-HA and Mg-Zn-HA) were obtained by an adaptation of the wet chemical deposition method. Coating of BG with these nanoparticles was performed by dip-coating to obtain HA-BG, Zn-HA-BG, Mg-HA-BG and Mg-Zn-HA-BG scaffolds.As predictor of the bone bonding ability of the produced scaffolds, in this study we investigated the formation of an apatite layer on the scaffold surfaces in the presence of simulated body fluid. The cytotoxicity and osteogenic properties of the materials in vitro was evaluated using human osteoblast-like MG-63 cell cultures. The mineralization assay following Kokubo’s protocol indicated that bi-metal loaded Mg-Zn-HA-BG scaffolds exhibited higher/faster bioactivity than mono-metal loaded scaffolds while mineralization of HA-BG, Zn-HA-BG and Mg-HA-BG was similar to that of uncoated scaffolds. Moreover, an increase of proliferation of MG-63 cells after 48 h and 7 days was measured by BrdU assays for Mg-Zn-HA-BG scaffolds. In agreement with these results, SEM images confirmed increased interaction between these scaffolds and cells, in comparison to that observed for mono-metal-loaded HA-coated scaffolds. Altogether, the obtained results suggest that nanocrystalline Mg-Zn-HA coatings enhance the biological performance of standard scaffolds of 45S5 BG composition. Thus these novel ion doped HA coated scaffolds are attractive systems for bone tissue engineering.

Surface-functionalization of mesoporous silica films for effective osteoblast-like cell culture

Abstract The transparent mesoporous silica (MPS) films prepared by a supramolecular templating method was immersed in simulated body fluid (SBF), and subsequently was sterilized to evaluate the film structural stability. With immersing in SBF, the calcium and phosphate ion adsorption on the film surfaces was promoted and the mesopore structures underwent the expansion, collapse and reconstruction processes for the SBF immersion time of 6 h. The MPS films immersed in SBF for 3 h exhibited the mesostructural stability with the sterilization processes. We also investigated the Fgn adsorption states on the sterilized MPS films, which were clearly changed with the immersion time in SBF because of the different adsorption states of calcium and phosphate ions at the mesopores. Furthermore, the proliferation and differentiation behaviors of the osteoblast-like cells on the sterilized MPS films were investigated to evaluate the cell culture ability. The osteoblast–like cells cultured on the sterilized MPS films had the fibrous morphologies while maintaining the high adhesion density and were promoted to be the bone tissue formation for 14 days, indicating the preferential film environment for the cells. Therefore, it was suggested that the MPS films of this study were useful for the cell culture substrate based on their high transparency, stability against sterilization treatment, and preferential cytocompatibility.

Development of hydroxyapatite from eggshell waste and a chitosan-based composite: In vitro behavior of human osteoblast-like cell (Saos-2) cultures

Hydroxyapatite was successfully synthesized using eggshell waste as a raw material. Eggshell waste and orthophosphoric acid were co-precipitated for 2 h at an ambient temperature. The pH of the solution was adjusted to 10 using ammonium hydroxide. Then, 10–30 wt% of hydroxyapatite was loaded into the chitosan film. Scanning electron microscopy and energy dispersive analyses confirmed the morphological properties and dispersion. A thermogravimetric analysis showed a significant enhancement in the thermal stability of the composite. The existence of hydroxyapatite resulted in a higher thermal stability. Furthermore, atomic force microscopy was used to investigate the roughness of the surface. With the addition of hydroxyapatite, the roughness significantly increased. The swelling behavior of the composite was observed in phosphate buffer saline solution. The hydroxyapatite offered the inferiority on the swelling behavior. A preliminary investigation on the in vitro behavior of Saos-2 was also performed. The composite presented good cytotoxicity, and thus, excellent properties as a tissue engineering material.

Prototype Orthopedic Bone Plates 3D Printed by Laser Melting Deposition.

Laser melting deposition is a 3D printing method usually studied for the manufacturing of machine parts in the industry. However, for the medical sector, although feasible, applications and actual products taking advantage of this technique are only scarcely reported. Therefore, in this study, Ti6Al4V orthopedic implants in the form of plates were 3D printed by laser melting deposition. Tuning of the laser power, scanning speed and powder feed rate was conducted, in order to obtain a continuous deposition after a single laser pass and to diminish unwanted blown powder, stuck in the vicinity of the printed elements. The fabrication of bone plates is presented in detail, putting emphasis on the scanning direction, which had a decisive role in the 3D printing resolution. The printed material was investigated by optical microscopy and was found to be dense, with no visible pores or cracks. The metallographic investigations and X-ray diffraction data exposed an unusual biphasic α+β structure. The energy dispersive X-ray spectroscopy revealed a composition very similar to the one of the starting powder material. The mapping of the surface showed a uniform distribution of elements, with no segregations or areas with deficient elemental distribution. The in vitro tests performed on the 3D printed Ti6Al4V samples in osteoblast-like cell cultures up to 7 days showed that the material deposited by laser melting is cytocompatible.

Graphene Oxide Finely Tunes the Bioactivity and Drug Delivery of Mesoporous ZnO Scaffolds.

Mesoporous zinc oxide (ZnO) scaffolds coated with drop-cast graphene oxide (GO) flakes are proposed to be a novel bilayer system featuring bioactivity, biocompatibility, and promising loading/release properties for controlled drug-delivery systems. The high-surface-area ZnO scaffolds show clear apatite deposition, but their particular surface chemistry and topography prevent the formation of a continuous coating, resulting in micrometric crystalline apatite aggregates after 28 days in simulated body fluid (SBF). When gentamicin sulfate (GS) is considered as a model molecule, pure ZnO scaffolds also show functional GS loading efficiency, with fast in vitro release kinetics driven by a simple diffusion mechanism. Strikingly, the bioactivity and GS delivery properties of mesoporous ZnO are efficiently triggered by drop-casting GO flakes atop the mesoporous scaffold surface. The resulting ZnO@GO bilayer scaffolds show the formation of a uniform apatite coating after 28 days in SBF and demonstrate a biocompatible behavior, supporting the culture of SaOS-2 osteoblast-like cells. Moreover, the GO coating also leads to a barrier-layer effect, preventing fast GS release, particularly in the short time range. This barrier effect, coupled with the existence of nanopores within the GO structure, sieves drug molecules from the mesoporous ZnO matrix and allows for a delayed release of the GS molecule. We, thus, demonstrated a new-generation ZnO@GO bilayer system as effective multifunctional and biocompatible scaffold for bone tissue engineering.

Bone Environment Influences Irreversible Adhesion of a Methicillin-Susceptible Staphylococcus aureus Strain.

Prosthesis and joint infections are an important threat in public health, especially due to the development of bacterial biofilms and their high resistance to antimicrobials. Biofilm-associated infections increase mortality and morbidity rates as well as hospitalization costs. Prevention is the best strategy for this serious issue, so there is an urgent need to understand the signals that could induce irreversible bacterial adhesion on a prosthesis. In this context, we investigated the influence of the bone environment on surface adhesion by a methicillin-susceptible Staphylococcus aureus strain. Using static and dynamic biofilm models, we tested various bone environment factors and showed that the presence of Mg2+, lack of oxygen, and starvation each increased bacterial adhesion. It was observed that human osteoblast-like cell culture supernatants, which contain secreted components that would be found in the bone environment, increased bacterial adhesion capacity by 2-fold (p = 0.015) compared to the medium control. Moreover, supernatants from osteoblast-like cells stimulated with TNF-α to mimic inflammatory conditions increased bacterial adhesion by almost 5-fold (p = 0.003) without impacting on the overall biomass. Interestingly, the effect of osteoblast-like cell supernatants on bacterial adhesion could be counteracted by the activity of synthetic antibiofilm peptides. Overall, the results of this study demonstrate that factors within the bone environment and products of osteoblast-like cells directly influence S. aureus adhesion and could contribute to biofilm initiation on bone and/or prosthetics implants.