EDENTULISM PROBLEM: USE OF STEM CELLS FOR BONE REGENERATION AND BONE INFLAMMATION

In this study we have started from a real odontoiatric clinic problem, which affects a high percentage of patients: edentulism. Implant therapy, started by Prof. Branemark in the seventies thanks to the discovery of osseointegration, has developed in the last 15 years. This therapy represents a valid solution for edentulous patients, although it cannot always be used. Severe osseous resorption or anatomical limitations hinder the use of implant and force the employ of biomaterials and/or autologous bone, which, according to the latest scientific research, represents the gold standard. However, intraoral bone graft is not always well tolerated by patients and, in cases of severe atrophy, the quantity of bone that can be taken from the oral cavity is not enough for complete regeneration. Although implant therapy represents the best solution for edentulous patients, in the latest years a biological complication has become more and more frequent: peri-implantitis. It represents a serious problem since it begins with an inflammation of peri-implant hard tissues that clinically leads to a loss of peri-implant alveolar bone and, eventually, to the loss of the implant as a whole. According to the most recent literature, this serious problem affects 10% of the patients and 4% of the implant sites after 10 years of follow-up. To make this pathology even more serious is the little knowledge that clinicians and researchers have about early diagnosis, etiopathogenesis and therapy. The purpose of this study is to address both edentulism and consequent osseous regeneration, as well as the problem of peri-implantitis from a biological point of view and with the help of tissue engineering. In particular, we wanted to test the ability of stem cells taken from adult tissue to favor - in shorter time - osseous regeneration and implant osseointegration both in vitro and on small and large animals. Furthermore, we have tested the anti-inflammatory ability of stem cells in bone tissue. Finally, with regard to the problem of peri-implantitis, we have searched for predictive genetic factors in order to possibly identify patients at risk. Preliminary results of studies performed both in vitro and on animal enable us to state that: -- It is possible to isolate stem cells from different adult tissues (adipose tissue, dental pulp) and test their genetic stability. -- For the cells isolated from dental pulp, we could identify an important connection between patient’s age and ability to differentiate and proliferate. -- Stem cells combined with different scaffolds are able to foster osseous regeneration faster. -- It has been recognized an actual anti-inflammatory ability of stem cells in bone tissue. -- Preliminary results on the use of CGH as genetic predictive technique for peri-implantitis are encouraging since they foreground a correlation between the genetic alteration of some chromosomal tracts and clinical onset of the disease

Porcine bone scaffolds adsorb growth factors secreted by MSCs and improve bone tissue repair

An ideal tissue-engineered bone graft should have both excellent pro-osteogenesis and pro-angiogenesis properties to rapidly realize the bone regeneration in vivo . To meet this goal, in this work a porcine bone scaffold was successfully used as a Trojan horse to store growth factors produced by mesenchymal stem cells (MSCs). This new scaffold showed a time-dependent release of bioactive growth factors, such as vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF), in vitro . The biological effect of the growth factors-adsorbed scaffold on the in vitro commitment of MSCs into osteogenic and endothelial cell phenotypes has been evaluated. In addition, we have investigated the activity of growth factor-impregnated granules in the repair of critical-size defects in rat calvaria by means of histological, immunohistochemical, and molecular biology analyses. Based on the results of our work bone tissue formation and markers for bone and vascularization were significantly increased by the growth factor-enriched bone granules after implantation. This suggests that the controlled release of active growth factors from porcine bone granules can enhance and promote bone regeneratio

Plasma rich in growth factors in human extraction sockets: a radiographic and histomorphometric study on early bone deposition

Objectives To determine whether and to what extent the additional application of plasma rich in growth factors ( PRGF ) to an extraction socket may influence the early bone deposition, as assessed by micro‐computed tomography (micro‐ CT ) scan as well as histomorphometric markers. Material and methods Twenty‐eight patients (age range: 34–74 years) contributing 36 extraction sockets were included in the study. Sockets were either treated with PRGF ( PRGF group; 18 sites in 11 patients) or left to spontaneous healing (control group; 18 sites in 17 patients). Radiographic and histomorphometric analysis was performed on bone cores trephined from each healing socket after 4–6 (T1) or 7–10 (T2) weeks of healing. Results Patients treated with PRGF application showed (i) similar bone volume and tissue mineral content, (ii) a trend, although not statistically significant, toward a greater number of CD 68+ cells (at T 1 and T 2) and v VW + cells (at T1), and (iii) a similar OCN staining score throughout the study, when compared with control group. Conclusions Plasma rich in growth factors‐treated group did not show any enhancement in early (4 and 8 weeks) bone deposition compared with control group.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/100296/1/clr12033.pd

Biocompatibility and antibacterial properties of zirconium nitride coating on titanium abutments: An in vitro study

Improving soft tissue attachment and reducing bacterial colonization on titanium abutments are key factors for the long-term maintenance of healthy soft and hard peri-implant tissues. This in vitro study was conducted to compare the biocompatibility and antibacterial activity of four different surfaces: uncoated Ti6Al4V, anodized, and coated with titanium nitride or zirconium nitride. Surface topography was investigated with a high-resolution system for measuring surface finishes. Human gingival fibroblast (HGF) adhesion and proliferation were examined using MTT assay, Scanning Electron Microscopy (SEM) imaging, immunofluorescence analysis and real-time PCR for selected target genes. The hemolysis and AMES tests were performed to assess the chemical compounds' blood compatibility and mutagenic potential, respectively. Antibacterial activity was tested against five bacterial strains isolated from the oral cavity (Streptococcus salivarius, S. sanguinis, S. mutans, S. sobrinus, S. oralis), and the percentage of dead bacteria was calculated. Roughness measurements confirmed a substantial similarity between the surfaces and their compatibility with clinical applications. MTT assay, SEM analysis and immunofluorescence staining showed adhesion and proliferation of HGFs cultured on all the examined surfaces. PCR confirmed that HGFs produced extracellular matrix components efficiently on all the surfaces. No hemolytic activity was detected, and the AMES test confirmed the surfaces' clinical safety. For all tested bacterial strains, biofilms grown on the zirconium nitride surface showed a higher percentage of dead bacteria than on the other disks. The titanium nitride surface inactivated bacterial biofilms, too, but to a lesser extent

Platform switching on wide-diameter external hex implants a finite element analysis

Objectives: The objective of this work was to use finite element analysis to compare the effect of forces coming to bear on abutments 4.1 or 5.0 mm in diameter connected to a 5.0 mm implant (i.e. with or without platform swit - ching). Study design: A 3D CAD model of a 5 x 11.5 mm external hex implant was developed, complete with a connection screw and either of two abutments, one 4.1 and the other 5 mm in diameter, to assess the influence of two loading conditions, i.e. 200 N loaded either axially or off center on the top of the abutment. Results and conclusions: In the symmetrically loaded models, greater stresses were transmitted to the bone in the area below the neck of the implant in the case of the wider-diameter abutment. When the narrower abutment was considered, the stress lines remained confined to the metal and were transferred to the bone in a more distal position. When the stresses in the bone where compared under non-symmetrical loading of the larger- and smaller-diameter abutments, the stresses reached lower values in the latter case. These findings indicate that platform switching (i.e. coupling a 4.1 mm abutment with a 5 mm implant) achieves a better, more even distribution of the peri-implant stresses deriving from simulated occlusal loads on the bone margins

The biological properties of OGI surfaces positively act on osteogenic and angiogenic commitment of mesenchymal stem cells

Osteogenesis process displays a fundamental role during dental implant osteointegration. In the present work, we studied the influence of Osteon Growth Induction (OGI) surface properties on the angiogenic and osteogenic behaviors of Mesenchymal Stem cells (MSC). MSC derived from dental pulp and HUVEC (Human Umbilical Vein Endothelial Cells) were grown in on OGI titanium surfaces, and cell proliferation and DNA synthesis were evaluated by MTT [3-(4,5-dimethylthiazol-2yl)-2,5-diphenyltetrazolium bromide] test and DNA quantification. Gene expression has been performed in order to evaluate the presence of mRNA related to endothelial and osteogenesis markers. Moreover, morphological and biochemical analyses of osteogenesis commitments has been performed. On OGI surfaces, MSC and HUVEC are able to proliferate. Gene expression profiler confirms that MSC on OGI surfaces are able to express endothelial and osteogenic markers, and that these expression are higher compared the expression on control surfaces. In conclusion On OGI surfaces proliferation, expression and morphological analyses of angiogenesis-associated markers in MSC are promoted. This process induces an increasing on their osteogenesis commitmen

Dental Pulp Stem Cells and Tissue Engineering Strategies for Clinical Application on Odontoiatric Field

Recent advances in tissue engineering have drawn scientists to test the possibility of tooth engineering and regeneration. Tooth regeneration is normally referred to as the regeneration of the entire tooth or root that can be integrated into the jaw bone. This technology is still at its infancy and when it matures, it may be used to restore missing teeth and replace artificial dental implants When the tooth is damaged but still in a reparable condition, regeneration of parts of the tooth structure can prevent or delay the loss of the whole tooth. To engineer and regenerate a whole tooth, the cell source, tissue engineering strategies and specific scaffolds needed to be correct choose. Indeed, for example, to repair partly lost tooth tissues such as PDL, dentin, and pulp, one or two particular types of dental stem cells may be sufficient to fulfill the need. In light of such considerations, aim of the present chapter is to define the main strategies to isolate dental pulp stem cells, their characterisation and differentiation, tissue enngineering strategies and clinical applications for the creation of artificial tissue useful in odontoiatric field

Decellularization and Delipidation Protocols of Bovine Bone and Pericardium for Bone Grafting and Guided Bone Regeneration Procedures

The combination of bone grafting materials with guided bone regeneration (GBR) membranes seems to provide promising results to restore bone defects in dental clinical practice. In the first part of this work, a novel protocol for decellularization and delipidation of bovine bone, based on multiple steps of thermal shock, washes with detergent and dehydration with alcohol, is described. This protocol is more effective in removal of cellular materials, and shows superior biocompatibility compared to other three methods tested in this study. Furthermore, histological and morphological analyses confirm the maintenance of an intact bone extracellular matrix (ECM). In vitro and in vivo experiments evidence osteoinductive and osteoconductive properties of the produced scaffold, respectively. In the second part of this study, two methods of bovine pericardium decellularization are compared. The osmotic shock-based protocol gives better results in terms of removal of cell components, biocompatibility, maintenance of native ECM structure, and host tissue reaction, in respect to the freeze/thaw method. Overall, the results of this study demonstrate the characterization of a novel protocol for the decellularization of bovine bone to be used as bone graft, and the acquisition of a method to produce a pericardium membrane suitable for GBR applications

Genetical stability and osteogenic ability of mesenchimal stem cells on demineralized bone matrices

Journal of Osseointegration Volume 7, Issue 1, 1 March 2015, Pages 2-7 Open Access Genetical stability and osteogenic ability of mesenchimal stem cells on demineralized bone matrices (Article) Pozzuoli, A.a, Gardin, C.b, Aldegheri, R.a, Bressan, E.c, Isola, M.d, Calvo-Guirado, J.L.e, Biz, C.a, Arrigoni, P.a, Feroni, L.b, Zavan, B.b a Department of Surgical,Oncological and Gastroenterological Sciences, University of Padua, Padua, Italy b Department of Biomedical Sciences, University of Padua, Padua, Italy c Department of Neurosciences, University of Padua, Padua, Italy d Department of Animal Medicine, Production and Health (MAPS), Italy e Department of General Dentistry, Faculty of Medicine and Dentistry, University of Murcia, Murcia, Spain Hide additional affiliations View references (44) Abstract Aim: Tissue engineering is a rapidly expanding field with regard to the use of biomaterials and stem cells in the orthopedic surgery. Many experimental studies have been done to understand the best characteristics of cells, materials and laboratory methods for safe clinical applications. The aim of this study was to compare the ability of 2 different human demineralized bone matrices (DBMs), the one enriched and the other not enriched with hyaluronic acid, to stimulate in vitro the proliferation and the osteogenic differentiation of human adipose-derived stem cells (ADSCs) seeded onto an osteoconductive scaffold. Materials and Methods: ADSCs were isolated, by enzymatic digestion, from abdominal adipose tissue of 5 patients undergoing cosmetic lipoaspiration surgery. ADSCs were then seeded onto a 3D scaffold in the presence of the two different osteoinductive matrices of human demineralized bone and evaluated for proliferation and osteogenic differentiation. The safety of the methods was verified using array-Comparative Genomic Hybridization (array-CGH). Results: ADSCs were able to differentiate in osteogenic sense. Both DBMs showed the ability to induce osteogenic differentiation of the cells. Conclusion: array-CGH showed no changes at genome level, thus confirming the safety of materials and method