Exploring the potential of chitosan/aragonite biocomposite derived from cuttlebone waste: Elaboration, physicochemical properties and in vitro bioactivity.

Cuttlefish bone biowaste is a potential source of a composite matrix based on chitin and aragonite. In the present work, we propose for the first time the elaboration of biocomposites based on chitosan and aragonite through the valorization of bone waste. The composition of the ventral and dorsal surfaces of bone is well studied by ICP-OES. An extraction process has been applied to the dorsal surface to extract ß-chitin and chitosan with controlled physico-chemical characteristics. In parallel, aragonite isolation was carried out on the ventral side. The freeze-drying method was used to incorporate aragonite into the chitosan polymer to form CHS/ArgS biocomposites. Physicochemical characterizations were performed by FT-IR, SEM, XRD, 1H NMR, TGA/DSC, potentiometry and viscometry. The ICP-OES method was used to evaluate in vitro the bioactivity level of biocomposite in simulated human plasma (SBF), enabling analysis of the interactions between the material and SBF. The results obtained indicate that the CHS/ArgS biocomposite derived from cuttlefish bone exhibits bioactivity, and that chitosan enhances the bioactivity of aragonite. The CHS/ArgS biocomposite showed excellent ability to form an apatite layer on its surface. After three days' immersion, FTIR and SEM analyses confirmed the formation of this layer.

Investigation on Akis granulifera (Coleoptera, Sahlberg, 1823) as a potential source of chitin and chitosan: Extraction, characterization and hydrogel formation.

The majority of studies have focused on the industrial exploitation of marine fisheries waste through the production of natural bioactive bioploymeres such as chitin and chitosan. However, in recent years, beetles are increasingly attracting the interest of scientists as a source of chitin and chitosan for the preparation of hydrogels for sustainable engineering development. In the present work, we focus on the study for the first time a new Moroccan species of beetle (Akis granulifera Sahlberg, 1823), for the extraction of chitin and the elaboration of chitosan. A chemical extraction process was used. Then, physicochemical characterizations by FT-IR, SEM, XRD, 1H NMR, TGA/DSC, Potentiometry, Viscosimetry, and elemental analysis were performed. In addition, to evaluate its physicochemical quality, the elaborated chitosan is combined with alginate to form a hydrogel. This hydrogel was effectively characterized by SEM, DRX and FTIR to show the potential of chitosan from Akis granulifera in biomaterial applications.

High Boron Content Enhances Bioactive Glass Biodegradation.

Derived Hench bioactive glass (BaG) containing boron (B) is explored in this work as it plays an important role in bone development and regeneration. B was also found to enhance BaG dissociation. However, it is only possible to incorporate a limited amount of B. To increase the amount of B in BaG, bioactive borosilicate glasses (BaG-Bx) were fabricated based on the use of the solution-gelation process (sol-gel). In this work, a high B content (20 wt.%) in BaG, respecting the conditions of bioactivity and biodegradability required by Hench, was achieved for the first time. The capability of BaG-Bx to form an apatite phase was assessed in vitro by immersion in simulated body fluid (SBF). Then, the chemical structure and the morphological changes in the fabricated BaG-Bx (x = 0, 5, 10 and 20) were studied. The formation of hydroxyapatite (HAp) layer was observed with X-ray diffraction (XRD) and infrared (IR) spectroscopy. The presence of HAp layer was confirmed using scanning electron microscopy (SEM) and transmission electron microscopy (TEM). Enhanced bioactivity and chemical stability of BaG-Bx were evaluated with an ion exchange study based on Inductively Coupled Plasma-Optical Emission Spectrometry (ICP-OES) and energy dispersive spectroscopy (EDS). Results indicate that by increasing the concentration of B in BaG-Bx, the crystallization rate and the quality of the newly formed HAp layer on BaG-Bx surfaces can be improved. The presence of B also leads to enhanced degradation of BaGs in SBF. Accordingly, BAG-Bx can be used for bone regeneration, especially in children, because of its faster degradation as compared to B-free glass.

Design of antibacterial apatitic composite cement loaded with Ciprofloxacin: Investigations on the physicochemical Properties, release Kinetics, and antibacterial activity.

This work aims to develop an injectable and antibacterial composite cement for bone substitution and prevention/treatment of bone infections. This cement is composed of calcium phosphate, calcium carbonate, bioactive glass, sodium alginate, and ciprofloxacin. The effect of ciprofloxacin on the microstructure, chemical composition, setting properties, cohesion, injectability, and compressive strength was investigated. The in vitro drug release kinetics and the antibacterial activity of ciprofloxacin-loaded composites against staphylococcus aureus and Escherichia coli pathogens were investigated. XRD and FTIR analysis demonstrated that the formulated cements are composed of a nanocrystalline carbonated apatite analogous to the mineral part of the bone. The evaluation of the composite cement's properties revealed that the incorporation of 3 and 9 wt% of ciprofloxacin affects the microstructural and physicochemical properties of the cement, resulting in a prolonged setting time, and a slight decrease in injectability and compressive strength. The in vitro drug release study revealed sustained release profiles over 18 days. The amounts of ciprofloxacin released per day (0.2 -15.2 mg/L) depend on the cement composition and the amount of ciprofloxacin incorporated. The antibacterial activity of ciprofloxacin-loaded cement composites attested to their effectiveness to inhibit the growth of Staphylococcus aureus and Escherichia coli.

The evolution of the global COVID-19 epidemic in Morocco and understanding the different therapeutic approaches of chitosan in the control of the pandemic.

In 2020, Coronavirus disease (COVID-19), a new viral respiratory disease caused by a virus that belongs to Coronaviridae family, has been identified. It is a very severe flu that negatively affects the functions of the lung and other respiratory organs. COVID-19 virus can be transmitted between people either by touching an infected person or by direct contact with their respiratory droplets. Therefore, the COVID-19 virus has become a global concern due to its rapid spread and severity. Based on the World Health Organization report from 2 March 2020 to 24 October 2022, the total infected cases and deaths in Morocco are around 1,265,389 (3.46%) and 16,280 (0.04%), respectively. Recently, some scientists have found that chitosan, a polymer existed in nature, can inhibit COVID-19 infection and repair damaged tissue. Therefore, understanding chitosan mechanisms in controlling COVID-19, might lead to innovative strategies in the medical field, such as developing drugs against SARS-CoV-2, and replacing vaccines, which have negative side effects. This review aims to show the evolution of the COVID-19 pandemic worldwide, specifically in Morocco, its pathophysiology, and its ability to silence the immune system. This review also provides an overview of the treatments and measures applied to protect human beings and how chitosan acts and controls COVID-19.

Formulation and characterization of hydroxyapatite-based composite with enhanced compressive strength and controlled antibiotic release.

A composite based on hydroxyapatite (HA) and chitosan (CS) combined with ciprofloxacin (CIP) was formulated by the solid-liquid mixing method. The optimization of the solid to the liquid ratio and the use of chitosan in a small amount (≤5 wt%) promoted the preparation of stable and rigid monoliths. A synergistic effect of CS and CIP contents on the compressive strength of the CIP-loaded composite was evidenced. The compressive strength of the fabricated biocomposite ranged in values from 1 to 6 MPa, comparable to those reported for cancellous bone. The improvement of the mechanical properties with the increase of the rate of organic components was correlated with the diminution of the surface area and the reduction in the pore volume of the specimens. On the other hand, the in vitro release experiments of the antibiotic indicated a sustained and controlled release of CIP over 10 days. Moreover, in vitro antibacterial tests performed on the biocomposite HA-CS5-CIP showed significant inhibition of Staphylococcus aureus and Escherichia coli pathogens. According to the showed results, the formulated composite with three-phase components could be a promising material for bone repair and local antibiotic release for the treatment of bone infections.

Bioactive glass doped with noble metal nanoparticles for bone regeneration: in vitro kinetics and proliferative impact on human bone cell line.

This work investigates the bioactivity of novel silver-doped (BG-Ag) and gold-doped (BG-Au) quaternary 46S6 bioactive glasses synthesized via a semi-solid-state technique. A pseudo-second-order kinetic model successfully predicted the in vitro uptake kinetic profiles of the initial ion-exchange release of Ca in simulated body fluid, the subsequent Si release, and finally, the adsorption of Ca and P onto the bioactive glasses. Doping with silver nanoparticles enhanced the rate of P uptake by up to approximately 90%; whereas doping with gold nanoparticles improved Ca and P uptake rates by up to about 7 and 2 times, respectively; as well as Ca uptake capacity by up to about 19%. The results revealed that the combined effect of Ca and Si release, and possibly the release of silver and gold ions into solution, influenced apatite formation due to their effect on Ca and P uptake rate and capacity. In general, gold-doped bioactive glasses are favoured for enhancing Ca and P uptake rates in addition to Ca uptake capacity. However, silver-doped bioactive glasses being less expensive can be utilized for applications targeting rapid healing. In vitro studies showed that BG, BG-Ag and BG-Au had no cytotoxic effects on osteosarcoma MG-63 cells, while they exhibited a remarkable cell proliferation even at low concentration. The prepared bioactive glass doped with noble metal nanoparticles could be potentially used in bone regeneration applications.

Development of hybrid scaffold: Bioactive glass nanoparticles/chitosan for tissue engineering applications.

Bone tissue engineering is gaining popularity as an alternative method for the treatment of osseous defects. A number of biodegradable polymers have been explored for tissue engineering purposes. A new family of biodegradable polymer/bioactive glass composite materials has been designed to be used in bone regeneration approaches. In this work, a hybrid scaffold of chitosan (CH) and bioactive glass nanoparticles (BGN) was prepared by the freeze-gelation method. This method has been studied by adjusting the concentration of acetic acid; this process can influence the structure properties of the scaffold. In this work, several BGN/CH composites have been prepared by varying the proportion of BGN in the hybrid scaffold (20, 40, 60, and 80%). Brunauer-Emmett-Teller results showed the increased surface area and porosity volume of our composite with decreasing BGN proportion. BGN/CH hybrid scaffold was characterized by using physicochemical techniques. Obtained results showed a macroporous morphology of the scaffold with a pore size of about 200 µm, and a homogeneous distribution of the BGN in the CH matrix. X-ray diffraction study confirmed the amorphous state of the BGN/CH hybrid scaffold. Interaction between CH and BGNs in the composite was confirmed. The in vitro assays showed adequate degradation properties, which is essential for the potential replacement by the new tissue. The in vitro bioactivity studies confirmed the formation of an apatite layer on the surface of the hybrid scaffold, which results in a direct bone bonding of the implant. These results indicate that BGN/CH hybrid scaffold developed is a potential candidate for bone tissue engineering.

Sub-chronic exposure to Kalach 360 SL, Glyphosate-based Herbicide, induced bone rarefaction in female Wistar rats.

We investigated the effects of Kalach 360 SL (KL), Glyphosate (G)-based herbicide, on bone tissue in different groups of female Wistar rats. Group 1 (n = 6) received a standard diet and served as a control, groups 2 and 3 (n = 6 each) received 0.07 ml (D1: 126 mg/Kg) and 0.175 ml (D2: 315 mg/Kg) of KL dissolved in the water for 60 days. The plasma was used to examine the metabolic balance markers (calcium, phosphorus, phosphatase alkaline (PAL), and vitamin D (vit D) and hormonal status (oestrogen and thyroid hormones). As a result, sub-chronic exposure to KL induced a perturbation of bone metabolism (calcium and phosphorus) and hormonal status disturbance. The histological and immunohistochemical study of the thyroid gland revealed a disturbance in morphological structure and thyroid cells function. Moreover, the KL disrupting eff ;ect on thyroid function was investigated by measuring changes in plasma levels of thyroid hormones. Free triiodothyronine (FT3) and thyroxine (FT4) were decreased in female rats breast-fed from rats treated with D and D2 of KL. This eff ;ect was associated with an increase in the plasma level of thyroid-stimulating hormone (TSH). Thus, that KL leads to hypothyroidism. Decrease in levels of oestrogen and thyroid dysfunction led to a disruption in the skeletal bone. The histological study and SEM in bone results allowed us to observe, in rats exposed to KL, the thinning and discontinuity of bone trabecular with a significant decrease in the number of nodes (intertrabecular links).In conclusion, KL sub-chronic exposure caused an aspect of osteoporosis.

Risedronate Effects on the In Vivo Bioactive Glass Behavior: Nuclear Magnetic Resonance and Histopathological Studies.

The present study aimed to enhance the anti-osteoporotic performance of bioactive glass (46S6) through its association with bisphosphonate such as risedronate with amounts of 8, 12, and 20%. Obtained composites have been called 46S6-8RIS, 46S6-12RIS, and 46S6-20RIS, respectively. In vitro and in vivo explorations have been carried out. Bioactive glass and risedronate association has been performed by adsorption process. Structure analyses have been carried out to evaluate and to understand their chemical interactions. Solid Nuclear Magnetic Resonance (NMR) has been employed to study the structural properties of obtained biocomposite. The spectra deconvolution showed the appearance of a species (Q 4) in the biocomposites 46S6-8RIS, 46S6-12RIS, and 46S6-20RIS indicating their successful chemical association. In vitro experiments showed the enhancement of the chemical reactivity of the composites 46S6-xRIS compared to the pure bioactive glass. In fact, the silicon liberation after 30 days of immersion was 50 ppm for pure bioactive glass 46S6, and 41, 64, and 62 from 46S6-8RIS, 46S6-12RIS, and 46S6-20RIS, respectively. Based on the in vitro results, 46S6-8RIS was implanted in the femoral condyle of an ovariectomized rat and compared with implanted pure glass in the goal to highlight its anti-osteoporotic performance. After 60 days, implanted group with 46S6-8RIS showed the increase in bone mineral density (BMD with 10%) and bone volume fraction (BV/TV with 80%) and the decrease in trabecular separation (Tb/Sp with 74%) when compared to that of 46S6 group. These results are confirmed by the histopathological analyses, which showed the bone trabeculae reconnection after the 46S6-8RIS implantation. Chemical analyses showed the reduction in silicon (Si) and sodium (Na) ion concentrations, and the rise in calcium (Ca) and phosphorus (P) ion levels, which was explained by the dissolution of biocomposite matrix and the deposition of hydroxyapatite layer. Histomorphometric results highlighted the risedronate effect on the antiosteoporotic phenomenon. Obtained results showed good behavior with only 8% of introduced risedronate in the glass matrix.

Antioxidative/oxidative effects and retarding osteoconductivity of ciprofloxacin-loaded porous polyvinyl alcohol/bioactive glass hybrid.

This study investigated the effect of bioglass (melting)-polyvinyl alcohol (BG (M)-PVA) and bioglass (melting)-polyvinyl alcohol-20 %ciprofloxacin (BG(M)-PVA-20Cip) in improving antioxidant activity and regenerating bone capacity. These composites were implanted in femoral condyles of ovariectomized Wistar rats and compared to that of controls groups. After the different period of implantation (15, 30, 60 and 90 days), the treatment of ovariectomized rats with BG(M)-PVA-20Cip showed a significantly higher malondialdehyde concentration when compared to that of BG(M)-PVA group. The superoxide dismutase, glutathione peroxidase and catalase in BG(M)-PVA-20Cip group showed significantly lower activities when compared to those in BG(M)-PVA group. So, BG(M)-PVA is more tolerated by organism than BG(M)-PVA-20Cip. Moreover, the alkaline phosphatase and acid phosphatase activities showed an excellent osteoinductive property of BG (M)-PVA. This property decreased with the presence of ciprofloxacin which is confirmed by histopathological analysis. Several physicochemical techniques showed a rapid reduction in Si and Na in one hand and an accelerator rise in Ca and P ions concentrations in other hand in BG(M)-PVA than in the BG(M)-PVA-20Cip. Therefore, the incorporation of ciprofloxacin in BG(M)-PVA is characterized by a prooxidant effect in oxidant-antioxidant balance at the beginning of treatment and a retard effect of formation of apatitic phase.

Prospect of Stem Cells in Bone Tissue Engineering: A Review.

Mesenchymal stem cells (MSCs) have been the subject of many studies in recent years, ranging from basic science that looks into MSCs properties to studies that aim for developing bioengineered tissues and organs. Adult bone marrow-derived mesenchymal stem cells (BM-MSCs) have been the focus of most studies due to the inherent potential of these cells to differentiate into various cell types. Although, the discovery of induced pluripotent stem cells (iPSCs) represents a paradigm shift in our understanding of cellular differentiation. These cells are another attractive stem cell source because of their ability to be reprogramed, allowing the generation of multiple cell types from a single cell. This paper briefly covers various types of stem cell sources that have been used for tissue engineering applications, with a focus on bone regeneration. Then, an overview of some recent studies making use of MSC-seeded 3D scaffold systems for bone tissue engineering has been presented. The emphasis has been placed on the reported scaffold properties that tend to improve MSCs adhesion, proliferation, and osteogenic differentiation outcomes.

A quantitative approach for studying the bioactivity of nanohydroxyapatite/gold composites.

This work describes a quantitative kinetic approach to assess the in vitro bioactivity of gold-doped hydroxyapatite-polyvinyl alcohol nanocomposites. The surface morphology of the in situ prepared nanocomposites as characterized by transmission electron microscopy (TEM) revealed a rod-like shape. Differential thermal analysis-thermogravimetric (DTA-TG), and fourier transformed infrared spectroscopy (FTIR) as well as zeta potential measurements of the prepared nanocomposites were carried out. Uptake profiles of Ca and P were studied onto nanocomposites of different gold concentrations after their soaking in simulated body fluid and they best followed the pseudo second-order kinetic model. The highest uptakes of both Ca and P were obtained using the nanocomposite with the lowest concentration of gold. Furthermore, sorption mechanism was described by the intraparticle diffusion model where pore diffusion was found to be the rate limiting step. The prepared nanocomposites have promising potential in orthopedic and tissue engineering applications because of their high capacity and fast uptake for Ca and P, which form apatite.

Substitution effects of a carbonated hydroxyapatite biomaterial against intoxication chloride nickel-exposed rats.

AIMS: This study aimed to investigate the potential effects of a synthetic apatite (carbonated hydroxyapatite) on the detoxification of a group of male "Wistar" rats exposed to nickel chloride. METHODS: Toxicity was evaluated by rats' bioassay of nickel chloride. Wistar rats received this metal daily by gavage for seven days (4 mg/ml nickel chloride/200 g body weight, BW). To detoxify this organism, a subcutaneous implantation of the apatite is made. RESULTS: The results revealed that exposure to nickel induced oxidative stress, disorders in the balances of ferric phosphocalcic, renal failures, liver toxicity and significant increase in nickel rates in the bones of intoxicated rats. The application of the carbonated hydroxyapatite presented in this study restored those disorders back to normal. The synthetic apatite protected the rats against the toxic effects of nickel by lowering the levels of lipid peroxidation markers and improving the activities of defense enzymes. It also amended ferric and phosphocalcic equilibriums, protected liver and kidney functions and reduced the nickel rate in the bones of the rats. Overall, the results provided strong support for the protective role of carbonated hydroxyapatite in the detoxification of rats exposed to nickel. Those beneficial effects were further confirmed by physico-chemical characterization (X-ray diffraction and infrared spectroscopy), which revealed its property of anionic and cationic substitution, thus supporting its promising candidacy for future biomedical application. CONCLUSION: The hydroxyapatite is an effective biomaterial to solve health problems, particularly detoxification against metals (nickel).

Genotoxicity effect, antioxidant and biomechanical correlation: experimental study of agarose-chitosan bone graft substitute in New Zealand white rabbit model.

Bone loss associated with skeletal trauma or metabolic diseases often requires bone grafting. In such situations, a biomaterial is necessary for migrated cells to produce new tissue. In this study, agarose-chitosan was implanted in the femoral condyle of New Zealand White rabbits that were divided into three groups: Group I was used as control; Groups II and III were used as implanted tissue with agarose-chitosan and presenting empty defects, respectively. This study evaluated the agarose-chitosan biocompatibility by determining the in vivo genotoxicity, oxidative stress balance that correlated with the hardness mechanical property. Moreover, the histopathological and quantitative elements analyzed by using inductively coupled plasma optical emission spectrometry were determined. After 30 days of implantation, the in vivo analysis of genotoxicity showed that agarose-chitosan did not induce chromosome aberration or micronucleus damage. A significant decrease in thiobarbituric and acid-reactive substance was observed after agarose-chitosan implantation in the bone tissue. Superoxide dismutase, catalase and glutathione peroxidase were significantly enhanced in agarose-chitosan-treated group compared with that of control group. A negative correlation coefficient of the mechanical property with malonyldialdehyde level was detected (R = -0.998). The histological study exhibited a significantly increased angiogenesis and newly formed tissue. No presence of inflammatory process, necrotic or fibrous tissue was detected. Major and trace elements such as Ca, P, Zn, Mg and Fe were increased significantly in the newly formed bone. These findings show that agarose-chitosan biomaterial implantation might be effective for treating trauma and bone regeneration.

Kinetic evaluation study on the bioactivity of silver doped hydroxyapatite-polyvinyl alcohol nanocomposites.

This work investigates the effect of adding silver nanoparticles (NPs) in ppm on the bioactivity of hydroxyapatite/polyvinyl alcohol nanocomposites (HAV). HAV prepared by an in situ biomimetic approach was doped with different concentrations of silver NPs (HAV-Ag), and the formed powder samples were characterized by different techniques such as Inductively Coupled Plasma-Optical Emission Spectroscopy (ICP-EOS), X-ray diffraction, transmission electron microscope, and Fourier Transform Infrared Spectroscopy. Bioactivity was evaluated in simulated body fluid through studying the kinetics of Ca and P uptake onto the different HAV-Ag nanocomposites. Uptake profiles of Ca and P were well described by a pseudo-second order kinetic model, and the obtained kinetic parameters confirmed that the highest uptake capacities were achieved by adding less than 0.001 ppm of silver NPs which is an amount not detectable by ICP. Furthermore, HAV-Ag nanocomposites were shown to be non-toxic as well as have a strong antibacterial effect. Silver NPs significantly enhanced the bioactivity of HAV nanocomposites and thus the developed nanocomposites promise to be excellent biomaterials for bone and reconstructive surgery applications.

The impact of orthopedic device associated with carbonated hydroxyapatite on the oxidative balance: experimental study of bone healing rabbit model.

Orthopedic devices are used in pathologic disorder as an adjunct to bone grafts to provide immediate structural stability. Unfortunately, the use of metallic devices has some complications. This study aimed to characterize the oxidative stress biomarker and the antioxidant enzyme profiles during bone regeneration. New Zealand White rabbits were divided into 4 groups: Group (I) was used as control (T), Groups II, III, and IV were used, respectively, as implanted tissue with carbonated hydroxyapatite (CHA), carbonated hydroxyapatite associated with external fixator (CHA + EF), and presenting empty defects (ED). Grafted bone tissues were carefully removed to measure malondialdehyde (MDA) concentration, superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase activities (GPx). Our results showed that 4 weeks after operation, treatment of rabbits with CHA + EF showed a significantly higher malondialdehyde (MDA) concentration when compared to that of control group. The SOD, CAT, and GPx in CHA + EF group showed significantly lower activities when compared to those in control group. Eight weeks after surgery, the CHA + EF group presented a lower concentration of MDA as compared to those seen after the first 4 weeks after surgery. On the other hand, the SOD, CAT, and GPx showed a higher activity when compared with the same group. Consequently, MDA concentration and the antioxidant enzyme activities were not significant (p > 0.05) when compared to those in control group rabbits. Histologic sampling has demonstrated successful time-patterned resorption accompanied by bone replacement and remodeling. These results suggest that there was a temporary increase in the oxidative marker level in CHA + EF healing bone and the 8-week period was sufficient to re-establish oxidant-antioxidant balance accompanied by bone repair in the tibia rabbit model.

Biologic response to carbonated hydroxyapatite associated with orthopedic device: experimental study in a rabbit model.

BACKGROUND: Carbonated hydroxyapatite (CHA) and related calcium phosphates have been studied for many years as implant materials due to their similarity with the mineral phase of bone. The main limitation of CHA ceramics as well as other bioactive materials is that they have poor mechanical proprieties. It is thought that the mechanical device can cause an increase in metabolic activity and bone healing. In this study we investigated the reactivity and tissue behaviour of implanted CHA biomaterial reinforced by mini external fixator. METHODS: The evaluation of biomaterial biocompatibility and osteogenesis was performed on a rabbit model over a period of 6 weeks by radiological, histological and scanning electron microscopy (SEM) coupled with energy dispersive X-ray SEM-energy-dispersive X-ray (EDX) analysis. RESULTS: While rabbits treated with CHA exhibited more bone formation, and fibrous tissue was observed when empty bone defects were observed. EDX analysis detected little calcium and phosphorus on the surface of the bone that was not implanted, while high content of calcium (62.7%) and phosphorus (38%) was found on the interface bone cement. CONCLUSIONS: Bone repairing showed that the mini external fixator stimulated the ossification which was pushed when grafted by CHA. This effect may play an important role in the prevention of implant loosening.

Effect of hydroxyapatite and beta-tricalcium phosphate nanoparticles on promonocytic U937 cells.

OBJECTIVES: Nanoparticles from implanted materials are reported as the main cause of implant failure. Monocytes are among the first cells to colonize the inflammatory site. We evaluated the biological effects of bone substitutes presented to U937 cells in vitro as micron- or nanometer-sized particles. METHODS: The HA (550 nm) and beta-TCP (550 nm) nanoparticles were incubated with U937 cells. Cell cycle modification, specific antigens expression, and the extent of cell death were determined. RESULTS: Firstly, by using the sulforhodamine B (SRB) test and the annexin V-FITC analysis by flow cytometry, our results provide evidence of the absence of cytotoxicity, and show that nanoparticles do not induce more apoptosis than microparticles in U937 cells. Secondly, although morphologic evidence of stimulation of U937-cells was found by confocal microscopy, neither bone substitute altered the distribution of the cells into different phases of the cell cycle (Kit Cycle Test Plus DNA). These results suggest that nanoparticles do not cause promonocyte maturation in macrophages. Thirdly, the flow cytometry results showed no differences in the expression of the adherence and activation markers. SIGNIFICANCE: The results suggest that nanoparticles do not promote the differentiation of promonocytic U937 cells into macrophages and do not induce an enhanced inflammatory response.

Development of a three-dimensional model for rapid evaluation of bone substitutes in vitro: effect of the 45S5 bioglass.

The evaluation of innovative bone substitutes requires the development of an optimal model close to physiological conditions. An interesting alternative is the use of an immortalized cell line to construct multicellular spheroids, that is, three-dimensional (3D) cultures. In this study, a modified hanging drops method has resulted in the generation of spheroids with a well-established human fetal osteoblasts line (hFOB 1.19), and tests have been focused on the effect of 45S5 bioglass ionic dissolution products in comparison with two-dimensional (2D) cultures. Depending on cell culture type, quantitative analysis (cell proliferation, viability/cytotoxicity, and cellular cycle) and qualitative analysis (electron microscopy and genes expression) showed a differential effect. Cell proliferation was enhanced in 2D-conditioned cultures in accordance with literature data, but decreased in 3D cultures submitted to the same conditions, without change of gene expression patterns. The decrease of cell proliferation, observed in conditioned spheroids, appears to be in agreement with clinical observations showing the insufficiency of commercially available bioglasses for bone repairing within nonbearing sites, such as periodontal defects or small bone filling, in general. Therefore, we suggest that this model could be adapted to the screening of innovative bioactive materials by laboratory techniques already available and extended monitoring of their bioactivity.