A comparative in vivo evaluation of bioactive glasses and bioactive glass-based composites for bone tissue repair

In this work a set of novel materials for bone tissue regeneration have been tested in vivo in an animal model. In fact, despite many studies have been devoted to amorphous 45S5 Bioglass®, there is lack in the literature of works aimed to study the in vivo performance of heat-treated – and thus partially crystallized – 45S5. As widely reported, crystallization limits the bioactivity of 45S5 and is the main reason that prevents a broader use of this material. Thus, in the present work, a recently developed bioactive glass (BG_Ca/Mix) is tested, since previous investigations demonstrated that BG_Ca/Mix is particularly promising by virtue of both its high bioactivity and lower tendency to crystallize with respect to 45S5. BG_Ca/Mix sintered powders and two composites, which contain BG_Ca/Mix and an increasing percentage (20 wt% or 70 wt%) of hydroxyapatite (HA), were considered. As a term of comparison, 45S5 sintered powders were also studied. The samples were implanted in rabbits' femurs and harvested after 8 weeks. The histological analysis demonstrated that BG_Ca/Mix has an osteoconductive ability slightly higher than that of 45S5 glass-ceramics, followed by that of the composites, which may represent the starting point for obtaining systems with degradation rate tailored for a given clinical application. Moreover, the 45S5 samples were locally cracked, probably because of a non-uniform dissolution in the physiological environment. On the contrary such cracks, which could lead to implant instability and unsuitable mechanical performance, were not observed in BG_Ca/Mix

Cell Proliferation to Evaluate Preliminarily the Presence of Enduring Self-Regenerative Antioxidant Activity in Cerium Doped Bioactive Glasses

(1) Background: a cell evaluation focused to verify the self-regenerative antioxidant activity is performed on cerium doped bioactive glasses. (2) Methods: the glasses based on 45S5 Bioglass®, are doped with 1.2 mol%, 3.6 mol% and 5.3 mol% of CeO2 and possess a polyhedral shape (~500 µm2). Glasses with this composition inhibit oxidative stress by mimicking catalase enzyme (CAT) and superoxide dismutase (SOD) activities; moreover, our previous cytocompatibility tests (neutral red (NR), 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) and Bromo-2-deoxyUridine (BrdU)) reveal that the presence of cerium promotes the absorption and vitality of the cells. The same cytocompatibility tests were performed and repeated, in two different periods (named first and second use), separated from each other by four months. (3) Results: in the first and second use, NR tests indicate that the presence of cerium promotes once again cell uptake and viability, especially after 72 h. A decrease in cell proliferation it is observed after MTT and BrdU tests only in the second use. These findings are supported by statistically significant results (4) Conclusions: these glasses show enhanced proliferation, both in the short and in the long term, and for the first time such large dimensions are studied for this kind of study. A future prospective is the implantation of these bioactive glasses as bone substitute in animal models

PRELIMINARY FINDINGS OF A POTENZIATED PIEZOSURGERGICAL DEVICE AT THE RABBIT SKULL

The number of available ultrasonic osteotomes has remarkably increased. In vitro and in vivo studies have revealed differences between conventional osteotomes, such as rotating or sawing devices, and ultrasound-supported osteotomes (Piezosurgery®) regarding the micromorphology and roughness values of osteotomized bone surfaces. Objective: the present study compares the micro-morphologies and roughness values of osteotomized bone surfaces after the application of rotating and sawing devices, Piezosurgery Medical® and Piezosurgery Medical New Generation Powerful Handpiece. Methods: Fresh, standard-sized bony samples were taken from a rabbit skull using the following osteotomes: rotating and sawing devices, Piezosurgery Medical® and a Piezosurgery Medical New Generation Powerful Handpiece. The required duration of time for each osteotomy was recorded. Micromorphologies and roughness values to characterize the bone surfaces following the different osteotomy methods were described. The prepared surfaces were examined via light microscopy, environmental surface electron microscopy (ESEM), transmission electron microscopy (TEM), confocal laser scanning microscopy (CLSM) and atomic force microscopy. The selective cutting of mineralized tissues while preserving adjacent soft tissue (dura mater and nervous tissue) was studied. Bone necrosis of the osteotomy sites and the vitality of the osteocytes near the sectional plane were investigated, as well as the proportion of apoptosis or cell degeneration. Results and Conclusions: The potential positive effects on bone healing and reossification associated with different devices were evaluated and the comparative analysis among the different devices used was performed, in order to determine the best osteotomes to be employed during cranio-facial surgery

Sol-gel derived bioactive glasses with low tendency to crystallize: synthesis, post-sintering bioactivity and possible application for the production of porous scaffolds.

A new sol-gel (SG) method is proposed to produce special bioactive glasses (BG_Ca family) characterized by a low tendency to devitrify. These formulations, derived from 45S5 Bioglass®, are characterized by a high content of CaO (45.6 mol%) and by a partial or complete substitution of sodium oxide with potassium oxide (total amount of alkaline oxides: 4.6 mol%), which increases the crystallization temperature up to 900°C. In this way, it is possible to produce them by SG preserving their amorphous nature, in spite of the calcination at 850°C. The sintering behavior of the obtained SG powders is thoroughly investigated and the properties of the sintered bodies are compared to those of the melt-derived (M) counterparts. Furthermore, the SG glass powders are successfully used to produce scaffolds by means of a modified replication technique based on the combined use of polyurethane sponges and polyethylene particles. Finally, in the view of a potential application for bone tissue engineering, the cytotoxicity of the produced materials is evaluated in vitro

Bone regeneration by novel bioactive glasses containing strontium and/or magnesium: A preliminary in-vivo study

In this work, a set of novel bioactive glasses have been tested in vivo in an animal model. The new compositions, characterized by an exceptional thermal stability and high in vitro bioactivity, contain strontium and/or magnesium, whose biological benefits are well documented in the literature. To simulate a long-term implant and to study the effect of the complete dissolution of glasses, samples were implanted in the mid-shaft of rabbits' femur and analyzed 60 days after the surgery; such samples were in undersized powder form. The statistical significance with respect to the type of bioactive glass was analyzed by Kruskal-Wallis test. The results show high levels of bone remodeling, several new bone formations containing granules of calcium phosphate (sometimes with amounts of strontium and/or magnesium), and the absence of adverse effects on bone processes due to the almost complete glass dissolution. In vivo results confirming the cell culture outcomes of a previous study highlighted that these novel bioglasses had osteostimulative effect without adverse skeletal reaction, thus indicating possible beneficial effects on bone formation processes. The presence of strontium in the glasses seems to be particularly interesting

Further Quinolizidine Derivatives as Antiarrhythmic Agents- 3

Fourteen quinolizidine derivatives, structurally related to the alkaloids lupinine and cytisine and previously studied for other pharmacological purposes, were presently tested for antiarrhythmic, and other cardiovascular effects on isolated guinea pig heart tissues in comparison to well-established reference drugs. According to their structures, the tested compounds are assembled into three subsets: (a) N-(quinolizidinyl-alkyl)-benzamides; (b) 2-(benzotriazol-2-yl)methyl-1- (quinolizidinyl)alkyl-benzimidazoles; (c) N-substituted cytisines. All compounds but two displayed antiarrhythmic activity that was potent for compounds 4, 1, 6, and 5 (in ascending order). The last compound (N-(3,4,5-trimethoxybenzoyl)aminohomolupinane) was outstanding, exhibiting a nanomolar potency (EC50 = 0.017 µM) for the increase in the threshold of ac-arrhythmia. The tested compounds shared strong negative inotropic activity; however, this does not compromise the value of their antiarrhythmic action. On the other hand, only moderate or modest negative chronotropic and vasorelaxant activities were commonly observed. Compound 5, which has high antiarrhythmic potency, a favorable cardiovascular profile, and is devoid of antihypertensive activity in spontaneously hypertensive rats, represents a lead worthy of further investigation

Pulsed Electro-Magnetic Field (PEMF) Effect on Bone Healing in Animal Models: A Review of Its Efficacy Related to Different Type of Damage

Biophysical energies are a versatile tool to stimulate tissues by generating biopotentials. In particular, pulsed electromagnetic field (PEMF) stimulation has intrigued researchers since the 1970s. To date, many investigations have been carried out in vivo, but a gold standard treatment protocol has not yet been defined. The main obstacles are represented by the complex setting of PEMF characteristics, the variety of animal models (including direct and indirect bone damage) and the lack of a complete understanding of the molecular pathways involved. In the present review the main studies about PEMF stimulation in animal models with bone impairment were reviewed. PEMF signal characteristics were investigated, as well as their effect on molecular pathways and osseous morphological features. We believe that this review might be a useful starting point for a prospective study in a clinical setting. Consistent evidence from the literature suggests a potential beneficial role of PEMF in clinical practice. Nevertheless, the wide variability of selected parameters (frequency, duration, and amplitude) and the heterogeneity of applied protocols make it difficult to draw certain conclusions about PEMF effectiveness in clinical implementation to promote bone healing. Deepening the knowledge regarding the most consistent results reported in literature to date, we believe that this review may be a useful starting point to propose standardized experimental guidelines. This might provide a solid base for further controlled trials, to investigate PEMF efficacy in bone damage conditions during routine clinical practic

Bone Healing Evaluation Following Different Osteotomic Techniques in Animal Models: A Suitable Method for Clinical Insights

Osteotomy is a common step in oncological, reconstructive, and trauma surgery. Drilling and elevated temperature during osteotomy produce thermal osteonecrosis. Heat and associated mechanical damage during osteotomy can impair bone healing, with consequent failure of fracture fixation or dental implants. Several ex vivo studies on animal bone were recently focused on heating production during osteotomy with conventional drill and piezoelectric devices, particularly in endosseous dental implant sites. The current literature on bone drilling and osteotomic surface analysis is here reviewed and the dynamics of bone healing after osteotomy with traditional and piezoelectric devices are discussed. Moreover, the methodologies involved in the experimental osteotomy and clinical studies are compared, focusing on ex vivo and in vivo findings

Synergistic Proapoptotic Activity of Recombinant Trail Plus the AKT Inhibitor Perifosine in Acute Myelogenous Leukemia Cells

To potentiate the response of acute myelogenous leukemia (AML) cells to TNF-Related Apoptosis- Inducing Ligand (TRAIL) cytotoxicity, we have examined the efficacy of a combination with perifosine, a novel phosphatidylinositol 3-kinase (PI3K)/Akt signaling inhibitor. The rationale for using such a combination is that perifosine was recently described to increase TRAIL-R2 receptor expression and decrease the cellular FLICE-Inhibitory Protein (cFLIP) in human lung cancer cell lines. Perifosine and TRAIL both induced cell death by apoptosis in the THP-1 AML cell line, which is characterized by constitutive PI3K/Akt activation, but lacks functional p53. Perifosine, at concentrations below IC50, dephosphorylated Akt and increased TRAIL-R2 levels, as demonstrated by western blot, RT-PCR, and flow cytometric analysis. Perifosine also decreased the long isoform of cFLIP (cFLIP-L) and the X-linked Inhibitor of Apoptosis Protein (XIAP) expression. Perifosine and TRAIL synergized to activate caspase-8 and induce apoptosis, which was blocked by a caspase- 8 selective inhibitor. Upregulation of TRAIL-R2 expression was dependent on a protein kinase Cα/ c-Jun-NH2-kinase 2/c-Jun signaling pathway activated by perifosine through reactive oxygen species production. Perifosine synergized with TRAIL also in primary AML cells displaying constitutive activation of the Akt pathway, by inducing apoptosis, Akt dephosphorylation, TRAIL-R2 upregulation, cFLIP-L and XIAP downregulation, and c-Jun phosphorylation. The combined treatment negatively affected the clonogenic activity of CD34+ cells from AML patients. In contrast, CD34+ cells from healthy donors were resistant to perifosine and TRAIL treatment. Our findings suggest that the combination perifosine and TRAIL might offer a novel therapeutic strategy for AML. Originally published Cancer Research, Vol. 68, No. 22, Nov 200

Comparative morphological study of bone regeneration in different rabbit cranial osteotomies: traditional versus new generation osteotomes

The aim of this study was to compare the bone regeneration after skull surgery by means of different recent osteotomy devices, piezosurgery medical (PM) and piezosurgery medical plus (PMP), versus a conventional rotary osteotomy device (OT) in 16 adult white New Zealand rabbits. After 14 days from surgery, the recovery of different resulting bone gaps were observed under structural and TEM-ultrastructural analyses. Our preliminary observations showed that the minimum distance from the osteotomy edge, at which it is possible to observe viable osteocytes in the pre-existent bone, was lower in PMP samples (average 38.2µm) with respect to PM (65.7µm) and OT (83.4µm) ones. Moreover, the size of osteotomy gap performed with OT was about twice in thickness with respect to those obtained by PM and PMP. In relation to our previous investigations (1) on two different types of bone formation occurring in sequence (static and dynamic osteogenesis), we observed in the present study that in PM and PMP samples the osteotomy gap is in a more advanced step of recovery with respect to OT ones: in OT samples numerous cords of stationary osteoblasts, forming preliminary bony trabeculae, were observed in the regenerating newly-formed bone (step of static osteogenesis); whereas, in PM and PMP samples bony trabeculae appeared mostly covered by typical prismatic osteoblasts, arranged in movable laminae (advanced step of dynamic osteogenesis). By histomorphometry, in PM and PMP samples with respect to OT ones: i) values of the regenerated bone area with respect to the total osteotomy area (BV/TV%) are about twice; ii) the number of TRAP positive osteoclasts per linear surface showed a significant increase, suggesting higher bone remodelling. Concerning SEM analysis, in all samples the regenerated bone displays, as expected, higher cell density and less mineralized matrix with respect to the pre-existent bone, independently from the device used. In conclusion, our results indicate that osteotomies performed with PM and PMP can be considered equivalent and show more advanced stages of healing compared to OT, in part due to the lower osteotomy thickness in PM and PMP with respect to OT. This work was supported by grants from MECTRON 2013-15