An alternate method for extracting DNA from environmentally challenged teeth for improved DNA analysis.

A grinding-free method to extract DNA from teeth via a direct minimal-invasive retrograde approach to the pulp cavity and dentine was compared to a standard grinding/pulverisation method. This alternate method uses endodontic dental files to access the root canals and pulp cavity for tissue and dentine harvest via the apical end of the roots and avoids mechanical damage to the crown and root morphology. In contrast, other methods require pulverisation of the whole root or tooth, transection or destruction of the occlusal surface to gain access to the DNA in the root canals and pulp chamber. This study compared two methods for preparing dentine powder from the roots of environmentally challenged teeth for forensic DNA analysis. We found that although the filing method was more laborious, and produced less dentine powder, the amount of amplifiable DNA per milligram of powder was substantially higher with the filing method compared to grinding the entire root. In addition, the number of short tandem repeat (STR) alleles detected and the peak height ratios of the STR profiles were notably higher. Although several other methods of extracting DNA-rich tissue from the pulp chamber of teeth have previously been reported, the method presented in this study is minimally invasive, thereby allowing the preservation of tooth and crown morphology.

Investigating the use of smartphones for learning purposes by Australian dental students.

BACKGROUND: Mobile Internet devices and smartphones have at present a significant potential as learning tools and the development of educational interventions based on smartphones have attracted increasing attention. OBJECTIVE: The objective of this study was to obtain a deeper insight in the nature of students' use of smartphones, as well as their attitudes towards educational use of mobile devices in order to design successful teaching interventions. METHOD: A questionnaire was designed, aiming to investigate the actual daily habitual use, as well as the attitudes of dental students towards smartphones for their university education purposes. The survey was used to collect data from 232 dental students. RESULTS: Of the 232 respondents, 204 (87.9%) owned a smartphone, and 191 (82.3%) had access to third generation (3G) mobile carriers. The most popular devices were the iPhone and Android. Most of the respondents had intermediate smartphone skills and used smartphones for a number of learning activities. Only 75/232 (32.3%) had specific educational applications installed, while 148/232 (63.7%) used smartphones to access to social media and found it valuable for their education (P<.05). Students accessing social media with their smartphones also showed significantly more advanced skills with smartphones than those who did not (P<.05). There was no significant association between age group, gender, origin, and smartphone skills. There was positive correlation between smartphone skills and students' attitudes toward improving access to learning material (r=.43, P<.05), helping to learn more independently (r=.44, P<.05), and use of smartphones by teaching staff (r=.45, P<.05). CONCLUSION: The results in this study suggest that students use smartphones and social media for their education even though this technology has not been formally included in the curriculum. This might present an opportunity for educators to design educational methods, activities, and material that are suitable for smartphones and allow students to use this technology, thereby accommodating students' current diverse learning approaches.

A clinically-feasible protocol for using human platelet lysate and mesenchymal stem cells in regenerative therapies.

The transplantation of human stem cells seeded on biomaterials holds promise for many clinical applications in cranio-maxillo-facial tissue engineering and regenerative medicine. However, stem cell propagation necessary to produce sufficient cell numbers currently utilizes fetal calf serum (FCS) as a growth supplement which may subsequently transmit animal pathogens. Human platelet lysate (HPL) could potentially be utilized to produce clinical-grade stem cell-loaded biomaterials as an appropriate FCS substitute that is in line with clinically-applicable practice. The goal of this study was to investigate whether HPL can be successfully used to propagate human mesenchymal stem cells (HMSCs) seeded on clinically-approved collagen materials under clinically-applicable conditions using FCS as a control. HMSCs were isolated from bone marrow and cultured in the presence of 10% FCS or 10% HPL. Characterization of HMSCs was performed by flow cytometry and through osteogenic and adipogenic differentiation assays. Proliferative capacity of HMSCs on both matrices was investigated by mitochondrial dehydrogenase assays (WST) and tissue coverage scanning electron microscopy (SEM). The isolated HMSC differentiated into osteogenic and adipogenic cells authenticating the multipotentiality of the HMSCs. WST tests and the SEM images demonstrated that HPL was generally superior to FCS in promoting growth of seeded HMSCs. For all other tests HPL supported HMSCs at least equal to FCS. In conclusion, HPL is an effective growth factor to allow expansion of clinical-grade HMSCs on clinically-approved biomaterials for maxillofacial and oral implantology applications.

Nanospiderwebs: artificial 3D extracellular matrix from nanofibers by novel clinical grade electrospinning for stem cell delivery.

Novel clinical grade electrospinning methods could provide three-dimensional (3D) nanostructured biomaterials comprising of synthetic or natural biopolymer nanofibers. Such advanced materials could potentially mimic the natural extracellular matrix (ECM) accurately and may provide superior niche-like spaces on the subcellular scale for optimal stem-cell attachment and individual cell homing in regenerative therapies. The goal of this study was to design several novel "nanofibrous extracellular matrices" (NF-ECMs) with a natural mesh-like 3D architecture through a unique needle-free multi-jet electrospinning method in highly controlled manner to comply with good manufacturing practices (GMP) for the production of advanced healthcare materials for regenerative medicine, and to test cellular behavior of human mesenchymal stem cells (HMSCs) on these. Biopolymers manufactured as 3D NF-ECM meshes under clinical grade GMP-like conditions show higher intrinsic cytobiocompatibility with superior cell integration and proliferation if compared to their 2D counterparts or a clinically-approved collagen membrane.

Comparison of in vitro biocompatibility of NanoBone(®) and BioOss(®) for human osteoblasts.

INTRODUCTION: Scaffolds for bone tissue engineering seeded with the patient's own cells might be used as a preferable method to repair bone defects in the future. With the emerging new technologies of nanostructure design, new synthetic biomaterials are appearing on the market. Such scaffolds must be tested in vitro for their biocompatibility before clinical application. However, the choice between a natural or a synthetic biomaterial might be challenging for the doctor and the patient. In this study, we compared the biocompatibility of a synthetic bone substitute, NanoBone(®) , to the widely used natural bovine bone replacement material BioOss(®) . MATERIAL AND METHODS: The in vitro behaviour of human osteoblasts on both materials was investigated. Cell performance was determined using scanning electron microscopy (SEM), cell vitality staining and four biocompatibility tests (LDH, MTT, WST, BrdU). RESULTS: We found that both materials showed low cytotoxicity and good biocompatibility. The MTT proliferation test was superior for Nanobone(®) . DISCUSSION: Both scaffolds caused only little damage to human osteoblasts and justify their clinical application. However, NanoBone(®) was able to support and promote proliferation of human osteoblasts slightly better than BioOss(®) in our chosen test set-up. The results may guide doctors and patients when being challenged with the choice between a natural or a synthetic biomaterial. Further experiments are necessary to determine the comparison of biocompatibility in vivo.

Proliferation assessment of primary human mesenchymal stem cells on collagen membranes for guided bone regeneration.

PURPOSE: Human mesenchymal stem cells (hMSCs) hold the potential for bone regeneration because of their self-renewing and multipotent character. The goal of this study was to evaluate the influence of collagen membranes on the proliferation of hMSCs derived from bone marrow. A special focus was set on short-term eluates derived from collagen membranes, as volatile toxic materials washed out from these membranes may influence cell behavior during the short time course of oral surgery. MATERIALS AND METHODS: The proliferation of hMSCs seeded directly on a collagen membrane (BioGide) was evaluated quantitatively using the cell proliferation reagent WST-1 (4-3-[4-iodophenyl]-2-[4-nitrophenyl]-2H-[5-tetrazolio]-1, 3--benzol-disulfonate) and qualitatively by scanning electron microscopy. Two standard biocompatibility tests, namely the lactate dehydrogenase and MTT (3-[4, 5-dimethyl-2-thiazolyl]-2, 5-diphenyl-2H-tetrazoliumbromide) tests, were performed using hMSCs cultivated in eluates from membranes incubated for 10 minutes, 1 hour, or 24 hours in serum-free cell culture medium. The data were analyzed statistically. RESULTS: Scanning electron microscopy showed large numbers of hMSCs with well-spread morphology on the collagen membranes after 7 days of culture. The WST test revealed significantly better proliferation of hMSCs on collagen membranes after 4 days of culture compared to cells cultured on a cover glass. Cytotoxicity levels were low, peaking in short-term eluates and decreasing with longer incubation times. CONCLUSION: Porcine collagen membranes showed good biocompatibility in vitro for hMSCs. If maximum cell proliferation rates are required, a prewash of membranes prior to application may be useful.

Ceramic scaffolds produced by computer-assisted 3D printing and sintering: characterization and biocompatibility investigations.

Hydroxyapatite (HAP) and tricalcium phosphate (TCP) are two very common ceramic materials for bone replacement. However, in general HAP and TCP scaffolds are not tailored to the exact dimensions of the defect site and are mainly used as granules or beads. Some scaffolds are available as ordinary blocks, but cannot be customized for individual perfect fit. Using computer-assisted 3D printing, an emerging rapid prototyping technique, individual three-dimensional ceramic scaffolds can be built up from TCP or HAP powder layer by layer with subsequent sintering. These scaffolds have precise dimensions and highly defined and regular internal characteristics such as pore size. External shape and internal characteristics such as pore size can be fabricated using Computer Assisted Design (CAD) based on individual patient data. Thus, these scaffolds could be designed as perfect fit replacements to reconstruct the patient's skeleton. Before their use as bone replacement materials in vivo, in vitro testing of these scaffolds is necessary. In this study, the behavior of human osteoblasts on HAP and TCP scaffolds was investigated. The commonly used bone replacement material BioOss(R) served as control. Biocompatibility was assessed by scanning electron microscopy (SEM), fluorescence microscopy after staining for cell vitality with fluorescin diacetate (FDA) and propidium iodide (PI) and the MTT, LDH, and WST biocompatibility tests. Both versions were colonised by human osteoblasts, however more cells were seen on HAP scaffolds than TCP scaffolds. Cell vitality staining and MTT, LDH, and WST tests showed superior biocompatibility of HAP scaffolds to BioOss, while BioOss was more compatible than TCP. Further experiments are necessary to determine biocompatibility in vivo. Future modifications of 3D printed scaffolds offer advantageous features for Tissue Engineering. The integration of channels could allow for vascular and nerve ingrowth into the scaffold. Also the complex shapes of convex and concave articulating joint surfaces maybe realized with these rapid prototyping techniques.