Alejandro A. Gorustovich

Osteoconductivity of strontium-doped bioactive glass particles: A histomorphometric study in rats

There is accumulating evidence that strontium (Sr)-containing bioceramics have positive effects on bone tissue repair. The aims of the present study were to evaluate the osteoconductivity of Sr-doped bioactive glass (BG) particles implanted in rat tibia bone marrow, and characterize the neoformed bone tissue by SEM-energy-dispersive X-ray microanalysis. Melt-derived BGs were prepared from a base 45S5 BG. Sr-doped glass (45S5.6Sr) was prepared using 6 wt % SrO as a substitute for the CaO. Histological analysis using undecalcified sections showed that new lamellar bone had formed along the surface of both 45S5 and 45S5.6Sr BG particles within 4 weeks. To evaluate osteoconductivity, affinity indices were calculated. At 30 days after implantation, 45S5 and 45S5.6Sr BGs had almost identical affinity indices (88% +/- 7% and 87% +/- 9%; p > 0.05). Strontium was not detected in the neoformed bone tissue surrounding 45S5.6Sr BG particles. These results indicate that 45S5.6Sr BG particles are osteoconductive when implanted inside the intramedullary canal of rat tibiae, and no alterations in bone mineralization, in terms of Ca/P ratio, were observed in the neoformed bone tissue around 45S5.6Sr BG particles.

Biological performance of boron-modified bioactive glass particles implanted in rat tibia bone marrow.

The aim of the present study was to characterize the neoformed bone tissue around boron-modified bioactive glass particles implanted in rat tibia bone marrow by histologic, histomorphometric and microchemical evaluation. Melt-derived glasses were prepared from a base 45S5 bioactive glass of nominal composition (45% SiO(2), 24.5% CaO, 24.5% Na(2)O and 6% P(2)O(5) in wt%). The glass composition was modified by adding 2% wt of boron oxide (45S5.2B). Histological and histomorphometric analyses using undecalcified sections showed that at 15 days post-implantation the area of neoformed bone tissue around the 45S5.2B particles was significantly higher than control 45S5 glass. No statistically significant differences were observed at 30 days post-implantation. The thickness of osseointegrated tissue on 45S5.2B BG particles was significantly greater than on the control at all experimental time-points evaluated. A statistically significant increase in the Ca:P ratio was observed in the neoformed bone around 45S5.2B particles 15 days post-implantation. The results of the present study provide evidence that particles of boron-modified 45S5 BG (45S5.2B) enhance bone formation more than 45S5 glass when implanted into the intramedullary canal of rat tibiae.

Biocompatibility and bone mineralization potential of 45S5 Bioglass®-derived glass–ceramic scaffolds in chick embryos

The aim of the present study was to evaluate the biocompatibility and bone mineralization potential of 45S5 Bioglass-derived glass-ceramic scaffolds using a chick embryo shell-less (ex ovo) culture system. Chick embryos were divided into two groups: control (C) and experimental (E). Scaffolds were placed on the chorioallantoic membrane (CAM) in embryos of group E at 10 days of total incubation. The 45S5 Bioglass-derived glass-ceramic scaffolds proved to be biocompatible in terms of the absence of inflammatory response at the implant site (CAM). Moreover, no alterations in the other end-points assessed, i.e. survival, stage of embryonic development and body weight, were detected. However, body length was greater in group E embryos than in group C embryos (p0.05). A marked reduction (93%) in Ca content in the scaffolds was evidenced by energy-dispersive X-ray analysis at 5 days post-implantation. Calcium release from the scaffold implanted on the CAM might have been responsible for the restoration of the bone-like phenotype in chick embryonic skeleton of group E as detected by Alcian blue-Alizarin red double staining, as well as by histological and microchemical analyses. Conversely, the control embryos exhibited a chondrogenic phenotype.

Histomorphometric Study of Alveolar Bone Healing in Rats Fed a Boron-Deficient Diet

Bone healing after tooth extraction in rats is a suitable experimental model to study bone formation. Thus, we performed a study to determine the effects of boron (B) deficiency on bone healing by using this model. The first lower right molar of weanling Wistar rats was extracted under anesthesia. The animals were divided into two groups: +B (adequate; 3 mg B/kg diet), and −B (boron‐deficient; 0.07 mg/kg diet). The animals in both groups were killed in groups of 10 at 7 and 14 days after surgery. The guidelines of the NIH for the care and use of laboratory animals were observed. The mandibles were resected, fixed, decalcified, and embedded in paraffin. Buccolingually oriented sections were obtained at the level of the mesial alveolus and used for histometric evaluations. Total alveolar volume (TAV) and trabecular bone volume per total volume (BV/TV) in the apical third of the alveolus were determined. Percentages of osteoblast surface (ObS), eroded surface (ES), and quiescent surface (QS) were determined. No statistical significant differences in food intake and body weight were observed. Histomorphometric evaluation found −B rats had 36% and 63% reductions in BV/TV at 7 and 14 days, respectively. When compared with +B rats, −B rats had significant reductions (57% and 87%) in ObS concomitantly with increases (120% and 126%) in QS at 7 and 14 days, respectively. The findings show that boron deficiency results in altered bone healing because of a marked reduction in osteogenesis. Anat Rec, 291:441–447, 2008.

A histomorphometric study of alveolar bone modelling and remodelling in mice fed a boron-deficient diet

OBJECTIVE Emerging evidence indicates that boron (B) plays a role in bone formation and maintenance. Thus, a study was performed to determine whether dietary B-deficiency affects periodontal alveolar bone modelling and remodelling. DESIGN Weanling Swiss mice (n=30) were divided into three groups: control diet (GI, 3mg B/kg); B-deficient diet (GII, 0.07 mg B/kg); and pair-fed with GII (GIII). The animals were maintained on their respective diets for 9 weeks and then sacrificed. The guidelines of the NIH for the care and use of laboratory animals were observed. The mandibles were resected, fixed, decalcified in 10% EDTA and embedded in paraffin. Buccolingually oriented sections were obtained at the level of the mesial root of the first lower molar and stained with H-E. Histomorphometric studies were performed separately on the buccal and lingual sides of the periodontal alveolar bone. Percentages of osteoblast surfaces (ObSs), eroded surfaces (ESs), and quiescent surfaces (QSs) were determined. RESULTS No statistically significant differences in food intake and body weight were observed between the groups. When compared with GI and GIII mice, GII mice (B-deficient) had 63% and 48% reductions in ObS and 58% and 73% increases in QS in buccal and lingual plates, respectively. ES were not affected by B nutriture. CONCLUSION The results are evidence that dietary boron deprivation in mice alters periodontal alveolar bone modelling and remodelling by inhibiting bone formation.

Histomorphometric study of peri-implant bone healing in the case of nerve injury: an experimental model in rats.

Causes of dental implant failure are of more than passing interest. Within the group of failures caused by iatrogenic factors, injury to the epineurium has been reported to cause the formation of peri-implant fibrous tissue (fibrointegration). The aim of this study was to perform a histomorphometric evaluation of the percentage of osseointegration of implants in contact with the epineurium. Twenty Wistar rats were used. The first lower molars were extracted under xylazine-ketamine anesthesia. A titanium screw implant (diameter, 0.75 mm; length, 2.26 mm) was placed. In the control group (n = 10), apical anchorage of the implant was performed. In the experimental group (n = 10), the apical portion of the implant was placed in contact with the epineurium of the lower mandibular nerve. All animals were killed by ether overdose 30 days after implantation. Radiographs were taken, and the samples were processed for embedding in acrylic resin. Ground sections were obtained along the vestibulo-lingual axis of the mesial alveolus that contained the implant and were stained with toluidine blue. The histologic analysis revealed the presence of bone tissue in the apical portion of the control group samples. In the experimental group, the implant was in contact with the epineurium. There were no statistically significant differences in the percentage of osseointegration between both groups (control group, 39% ± 9%; experimental group, 38% ± 10%). The results obtained with this experimental model show that the contact of the implant with the epineurium would not impair the process of osseointegration.

Nanocomposite scaffolds with tunable mechanical and degradation capabilities: co-delivery of bioactive agents for bone tissue engineering

Novel multifunctional nanocomposite scaffolds made of nanobioactive glass and alginate crosslinked with therapeutic ions such as calcium and copper were developed for delivering therapeutic agents, in a highly controlled and sustainable manner, for bone tissue engineering. Alendronate, a well-known antiresorptive agent, was formulated into microspheres under optimized conditions and effectively loaded within the novel multifunctional scaffolds with a high encapsulation percentage. The size of the cation used for the alginate crosslinking impacted directly on porosity and viscoelastic properties, and thus, on the degradation rate and the release profile of copper, calcium and alendronate. According to this, even though highly porous structures were created with suitable pore sizes for cell ingrowth and vascularization in both cases, copper-crosslinked scaffolds showed higher values of porosity, elastic modulus, degradation rate and the amount of copper and alendronate released, when compared with calcium-crosslinked scaffolds. In addition, in all cases, the scaffolds showed bioactivity and mechanical properties close to the endogenous trabecular bone tissue in terms of viscoelasticity. Furthermore, the scaffolds showed osteogenic and angiogenic properties on bone and endothelial cells, respectively, and the extracts of the biomaterials used promoted the formation of blood vessels in an ex vivo model. These new bioactive nanocomposite scaffolds represent an exciting new class of therapeutic cell delivery carrier with tunable mechanical and degradation properties; potentially useful in the controlled and sustainable delivery of therapeutic agents with active roles in bone formation and angiogenesis, as well as in the support of cell proliferation and osteogenesis for bone tissue engineering.

In Vitro Human Umbilical Vein Endothelial Cells Response to Ionic Dissolution Products from Lithium-Containing 45S5 Bioactive Glass

Since lithium (Li+) plays roles in angiogenesis, the localized and controlled release of Li+ ions from bioactive glasses (BGs) represents a promising alternative therapy for the regeneration and repair of tissues with a high degree of vascularization. Here, microparticles from a base 45S5 BG composition containing (wt %) 45% SiO2, 24.5% Na2O, 24.5% CaO, and 6% P2O5, in which Na2O was partially substituted by 5% Li2O (45S5.5Li), were obtained. The results demonstrate that human umbilical vein endothelial cells (HUVECs) have greater migratory and proliferative response and ability to form tubules in vitro after stimulation with the ionic dissolution products (IDPs) of the 45S5.5Li BG. The results also show the activation of the canonical Wnt/β-catenin pathway and the increase in expression of proangiogenic cytokines insulin like growth factor 1 (IGF1) and transforming growth factor beta (TGFβ). We conclude that the IDPs of 45S5.5Li BG would act as useful inorganic agents to improve tissue repair and regeneration, ultimately stimulating HUVECs behavior in the absence of exogenous growth factors.

Chapter 15:Bioactive Glasses for Soft Tissue Engineering Applications

In the last few years the usage of bioactive glasses as scaffolds for soft tissue engineering has been investigated more thoroughly. The reason for the boost in interest is the attractive properties bioactive glasses offer, including bioactivity as well as antibacterial, angiogenic and hemostatic properties. So far, most research efforts have focused on applications for repairing skin and nerve tissue although there have been interesting developments in other fields including lung and intestine, which could potentially benefit a large group of patients. Three review articles on this topic have been published, so this chapter will mainly focus on the latest relevant findings. A great number of patents have been registered for the use of bioactive glass for hard tissue engineering. Recently, however, patents have been filed detailing the use of bioactive glass for soft tissue engineering applications which open the way to marketing bioactive glasses for soft tissue repair. The angiogenic effect of bioactive glasses and their dissolution products is of great interest for tissue engineering applications in general and in particular for soft tissue regeneration and repair. The third part of this chapter will detail the latest research on these angiogenic properties of bioactive glasses.