Rossella Bedini

Porous gelatin hydrogels by gas-in-liquid foam templating

In the present work, porous gelatin scaffolds were prepared by insufflating an inert gas (argon) inside a concentrated solution of gelatin in the presence of a suitable polymeric surfactant in association with sodium dodecyl sulfate. The implementation of such an approach involved the design and manufacturing of a specially dedicated reactor. Foams were prepared at a temperature of 50 °C and then let gel at 4 °C. After purification, they were auto-cross-linked with EDC and freeze-dried. The scaffolds synthesised with this technique present a morphology characterised by pores of spherical symmetry highly interconnected by a plurality of interconnections and, as a consequence, are particularly suited for cell culturing. The dosage of the volume of the insufflated gas permits to modulate the scaffold pore and interconnect dimensions. In this way matrices characterised by void and interconnect average diameters ranging from 250 to 360 μm and from 80 to 150 μm, respectively, can be successfully obtained.

Ex vivo-transduced autologous skin fibroblasts expressing human Lim mineralization protein-3 efficiently form new bone in animal models.

Local gene transfer of the human Lim mineralization protein (LMP), a novel intracellular positive regulator of the osteoblast differentiation program, can induce efficient bone formation in rodents. To develop a clinically relevant gene therapy approach to facilitate bone healing, we have used primary dermal fibroblasts transduced ex vivo with Ad.LMP-3 and seeded on a hydroxyapatite/collagen matrix prior to autologous implantation. Here, we demonstrate that genetically modified autologous dermal fibroblasts expressing Ad.LMP-3 are able to induce ectopic bone formation following implantation of the matrix into mouse triceps and paravertebral muscles. Moreover, implantation of the Ad.LMP-3-modified dermal fibroblasts into a rat mandibular bone critical size defect model results in efficient healing, as determined by X-rays, histology and three-dimensional microcomputed tomography (3DμCT). These results demonstrate the effectiveness of the non-secreted intracellular osteogenic factor LMP-3 in inducing bone formation in vivo. Moreover, the utilization of autologous dermal fibroblasts implanted on a biomaterial represents a promising approach for possible future clinical applications aimed at inducing new bone formation.

Quality of thermoplasticized and single point root fillings assessed by micro-computed tomography.

AIM To evaluate ex vivo the quality of root fillings completed by two thermoplasticized gutta-percha techniques (Thermafil and System B) and a cold gutta-percha technique (single point) by μCT analysis. METHODOLOGY A total of 30 freshly extracted human single-rooted permanent teeth were selected. Root canals were prepared with ProTaper Universal instruments and then randomly divided into three groups (n = 10) depending on the filling technique. In group 1, canals were filled with a single-point technique; group 2 was filled with Thermafil; in group 3 System B was used. In group 1 and group 3, the root filling was performed using ProTaper Universal gutta-percha points, in group 2 Thermafil obturators were used; AH-Plus sealer was used in all groups. Assessment of the root filling was carried out by μCT, using a desktop X-ray micro focus CT scanner. Percentage of root canal filling materials and voids was calculated for each specimen. Data were statistically analysed using Kruskal-Wallis test (P < 0.05). RESULTS Mean percentages of filling materials were 98.379 ± 1.204 in the single-point group, 99.023 ± 1.457 in Thermafil group, and 98.167 ± 3.432 in System B group. No statistically significant difference was found amongst the groups. CONCLUSION All techniques produced comparable results in terms of percentage of filling and void distribution.

Gas-in-Liquid Foam Templating as a Method for the Production of Highly Porous Scaffolds

In the present work, a novel synthetic methodology for the preparation of scaffold of biopolymeric nature is described. In particular, a porous gelatin scaffold was prepared by foam templating. The gas phase, nitrogen, was generated by means of the reaction between sulfamic acid and sodium nitrite in situ a concentrated solution of gelatin and in the presence of a suitable polymeric surfactant in association with sodium dodecyl sulfate. The foam was prepared at a temperature of 45 degrees C and then let gel at 5 degrees C. After purification, the physical gel was auto-cross-linked with EDC and freeze-dried. The scaffold synthesized with this technique presents a morphology characterized by voids of spherical symmetry highly interconnected by a plurality of interconnects, and, as a consequence, is particularly suited for cell culturing. In more quantitative terms, voids and interconnects are characterized by an average diameter of 230 and 90 microm, respectively. Preliminary tests of cell culturing demonstrated the suitability of such a scaffold for tissue engineering applications.

Synthetic bone substitute engineered with amniotic epithelial cells enhances bone regeneration after maxillary sinus augmentation.

Background Evidence has been provided that a cell-based therapy combined with the use of bioactive materials may significantly improve bone regeneration prior to dental implant, although the identification of an ideal source of progenitor/stem cells remains to be determined. Aim In the present research, the bone regenerative property of an emerging source of progenitor cells, the amniotic epithelial cells (AEC), loaded on a calcium-phosphate synthetic bone substitute, made by direct rapid prototyping (rPT) technique, was evaluated in an animal study. Material And Methods Two blocks of synthetic bone substitute (∼0.14 cm3), alone or engineered with 1×106 ovine AEC (oAEC), were grafted bilaterally into maxillary sinuses of six adult sheep, an animal model chosen for its high translational value in dentistry. The sheep were then randomly divided into two groups and sacrificed at 45 and 90 days post implantation (p.i.). Tissue regeneration was evaluated in the sinus explants by micro-computer tomography (micro-CT), morphological, morphometric and biochemical analyses. Results And Conclusions The obtained data suggest that scaffold integration and bone deposition are positively influenced by allotransplantated oAEC. Sinus explants derived from sheep grafted with oAEC engineered scaffolds displayed a reduced fibrotic reaction, a limited inflammatory response and an accelerated process of angiogenesis. In addition, the presence of oAEC significantly stimulated osteogenesis either by enhancing bone deposition or making more extent the foci of bone nucleation. Besides the modulatory role played by oAEC in the crucial events successfully guiding tissue regeneration (angiogenesis, vascular endothelial growth factor expression and inflammation), data provided herein show that oAEC were also able to directly participate in the process of bone deposition, as suggested by the presence of oAEC entrapped within the newly deposited osteoid matrix and by their ability to switch-on the expression of a specific bone-related protein (osteocalcin, OCN) when transplanted into host tissues.

Morphological comparison of PVA scaffolds obtained by gas foaming and microfluidic foaming techniques.

In this article, we have exploited a microfluidic foaming technique for the generation of highly monodisperse gas-in-liquid bubbles as a templating system for scaffolds characterized by an ordered and homogeneous porous texture. An aqueous poly(vinyl alcohol) (PVA) solution (containing a surfactant) and a gas (argon) are injected simultaneously at constant flow rates in a flow-focusing device (FFD), in which the gas thread breaks up to form monodisperse bubbles. Immediately after its formation, the foam is collected and frozen in liquid nitrogen, freeze-dried, and cross-linked with glutaraldehyde. In order to highlight the superior morphological quality of the obtained porous material, a comparison between this scaffold and another one, also constituted of PVA but obtained with a traditional gas foaming technique, was carried out. Such a comparison has been conducted by analyzing electron microscopy and X-ray microtomographic images of the two samples. It turned out that the microfluidic produced scaffold was characterized by much more uniform porous texture than the gas-foaming one as witnessed by narrower pore size, interconnection, and wall thickness distributions. On the other side, scarce pore interconnectivity, relatively low pore volume, and limited production rate represent, by now, the principal disadvantages of microfluidic foaming as scaffold fabrication method, emphasizing the kind of improvement that this technique needs to undergo.

Micro-computerized tomographic analysis of radicular and canal morphology of premolars with long oval canals

OBJECTIVE The aims of this in vitro study were to measure root and canal diameters, root and canal diameter ratios, mean taper of the root canal and of each root canal section, and radicular wall thickness at different levels in premolars with long oval root canals. STUDY DESIGN Thirty human premolars, with single long oval canals were selected for this study. The specimens were analyzed with micro-computerized tomography. The cross-sections corresponding to 1, 2, 3, 4, 5, 7, 9, 11, and 13 mm from the radiologic apex were analyzed to measure the mesiodistal (MD) and buccolingual (BL) diameters of the canals and the thickness of the root and the walls. The BL/MD ratios of the canal (DeltaC) and the root (DeltaR) diameters were calculated, as was as the mean taper in both a BL and an MD dimension. Multiple logistic regression analyses were performed with a level of significance of P = .05. RESULTS At all levels of analysis, the BL diameter was greater than the MD diameter for both the canal and the root. Generally, DeltaC and DeltaR increased coronally. Buccal and lingual wall thicknesses were greater than mesial and distal at all levels. Canal diameters at 1 mm from the apex were >0.30 mm in the shorter-oval diameter and >0.40 mm in the longer-oval diameter. In all root segments the BL taper was greater than the MD taper. CONCLUSIONS An oval canal anatomy was frequently present even in the most apical sections of the root canals. A high correlation was established between the shape of the root canal and the corresponding root.

Present and future in the use of micro-CT scanner 3D analysis for the study of dental and root canal morphology

The goal of the present article is to illustrate and analyze the applications and the potential of microcomputed tomography (micro-CT) in the analysis of tooth anatomy and root canal morphology. The authors performed a micro-CT analysis of the following different teeth: maxillary first molars with a second canal in the mesiobuccal (MB) root, mandibular first molars with complex anatomy in the mesial root, premolars with single and double roots and with complicated apical anatomy. The hardware device used in this study was a desktop X-ray microfocus CT scanner (SkyScan 1072, SkyScan bvba, Aartselaar, Belgium). A specific software ResolveRT Amira (Visage Imaging) was used for the 3D analysis and imaging. The authors obtained three-dimensional images from 15 teeth. It was possible to precisely visualize and analyze external and internal anatomy of teeth, showing the finest details. Among the 5 upper molars analyzed, in three cases, the MB canals joined into one canal, while in the other two molars the two mesial canals were separate. Among the lower molars two of the five samples exhibited a single canal in the mesial root, which had a broad, flat appearance in a mesiodistal dimension. In the five premolar teeth, the canals were independent; however, the apical delta and ramifications of the root canals were quite complex. Micro-CT offers a simple and reproducible technique for 3D noninvasive assessment of the anatomy of root canal systems.

Evaluation of guided bone regeneration in rabbit femur using collagen membranes.

&NA; The aim of the study was to evaluate the mechanical performance and the structure of neoformed bone around hydroxyapatite‐coated titanium fixtures according to guided bone regeneration techniques. Ten hydroxyapatitecoated titanium fixtures were inserted in the femurs of five rabbits, in which a cortical defect was created and after the insertion of the fixture, covered with a resorbable membrane obtained from bovine Achilles tendon collagen Type I (A implant). In the same femur, a second fixture was inserted in similar cavities without application of the membrane (B implant). After 60 days, the animals were sacrificed, and block sections of the femoral bone con taining the implants were embedded in polymethylmetacrylate and subjected to tensile shear‐stress at break testing. After the detachment of the implants from the bone, their surfaces were examined with a scanning electron microscope. Tensile shear‐stress values for A and B implant specimens were comparable to some extent, but the former had a lower performance. In this regard, scanning electron microscope observations showed that the neoformed cortical bone present cervically around implant A was much thicker than around implant B. (Implant Dent 2000;9:219‐225)

Role of X-ray microtomography in tissue engineering

The structure and architecture of scaffolds are crucial factors in scaffolds-based tissue engineering since they affect the functionality of the tissue engineering construct and the eventual application in health care. Therefore, effective scaffold assessment techniques are required right at the initial stages of research and development so as to select or design scaffolds with suitable properties. Furthermore, since the biological performances of a scaffold is evaluated with respect to its capacity of favouring cell adhesion, proliferation as well as production of extracellular matrix, it is important to have an analytical technique able to monitor the various stages of cell culture both in vitro and especially in vivo. Finally, the development of a vascular network inside the cell scaffold construct is a fundamental requisite for achieving a full integration of the developing tissue with the host tissue. Also in this respect it is mandatory to assess the propensity of the scaffold to be permeated by blood vessels. In the review, it will be shown how X-ray microtomography (micro-CT) can give fundamental information regarding all the three aspects outlined above.