Application of enamel matrix derivative in conjunction with non-surgical therapy for treatment of moderate to severe periodontitis: A 12-month, randomized prospective, multicenter study.

BACKGROUND: Treatment of periodontitis aims to halt progressive bone and attachment loss and regenerate periodontal structures. In this study, the effect of using an enamel matrix derivative (EMD) as an adjunct to non-surgical periodontal therapy (test) versus non-surgical therapy alone (control) was evaluated. METHODS: A prospective, split-mouth, multicenter study evaluated scaling and root planing (SRP) with and without EMD in 51 patients presenting with moderate to severe periodontitis (PPD = 5 to 8 mm) in at least 2 pockets per contralateral quadrants within the same arch. The primary outcome variable was change in clinical attachment level (CAL) after 12 months. Secondary variables included probing pocket depth (PPD), bleeding on probing (BoP), gingival margin level, dentin hypersensitivity, and percent of pockets converted to sites no longer requiring surgical treatment. RESULTS: CAL changed significantly (P  < 0.001) from baseline to 12 months for both treatment modalities (test = -2.2 ± 1.5 mm versus control = -2.1 ± 1.3 mm) and similarly for PPD; the difference between groups was not significant. A significant difference, favoring test conditions, was observed in percentage of both healthy PPDs (pockets < 5 mm) and converted pockets (sites no longer requiring surgical treatment); 79.8% of test versus 65.9% of control sites. BoP decreased significantly more (P < 0.05) in test sites (BoP at 17.8% test versus 23.1% control). CONCLUSIONS: Both test and control treatments resulted in significant improvements in CAL and PPD. The adjunct use of EMD with SRP resulted in significantly greater improvements in overall periodontal health with less frequent BoP and a higher number of healthy PPDs.

Retrospective cohort study of 4,591 dental implants: Analysis of risk indicators for bone loss and prevalence of peri-implant mucositis and peri-implantitis.

BACKGROUND: Due to the risk of peri-implantitis, following dental implant placement, this study aimed to evaluate risk indicators associated with marginal bone loss from a retrospective open cohort study of 4,591 dental implants, placed in private practice, with 5- to 10-year follow-up. Furthermore, the prevalence of mucositis and peri-implantitis among the study cohort was evaluated, comparing strict versus relaxed criteria for bleeding on probing. METHODS: Periapical radiographs were used to evaluate changes in crestal bone level. Peri-implant soft tissue was evaluated using an ordinal mucosal index in comparison with the conventional binary threshold for bleeding (i.e., present or not). Periodontal probing depth was not evaluated. Linear mixed models were used to evaluate bone level over time, and other risk indicators, at the patient and implant level. RESULTS: Risk indicators found to have a significant impact on bone level included: autoimmune disease, heavy smoking, bisphosphonate therapy, implant location, diameter and design, and the presence of a bone defect at site of implantation. The prevalence of mucositis at the implant level was 38.6% versus 14.2% at 6 to 7 years, when using strict versus relaxed criteria, respectively. The prevalence of peri-implantitis after 6 to 7 years was 4.7% and 3.6% when using strict versus relaxed criteria, respectively. CONCLUSIONS: The results of this study identify several risk factors associated with bone loss. Furthermore, the prevalence of mucositis and peri-implantitis was shown to be lower at both the implant and the patient when using strict versus relaxed criteria based on the assessment of oral health surrounding dental implants.

Enamel matrix derivative: a review of cellular effects in vitro and a model of molecular arrangement and functioning.

BACKGROUND: Enamel matrix derivative (EMD), the active component of Emdogain®, is a viable option in the treatment of periodontal disease owing to its ability to regenerate lost tissue. It is believed to mimic odontogenesis, though the details of its functioning remain the focus of current research. OBJECTIVE: The aim of this article is to review all relevant literature reporting on the composition/characterization of EMD as well as the effects of EMD, and its components amelogenin and ameloblastin, on the behavior of various cell types in vitro. In this way, insight into the underlying mechanism of regeneration will be garnered and utilized to propose a model for the molecular arrangement and functioning of EMD. METHODS: A review of in vitro studies of EMD, or components of EMD, was performed using key words "enamel matrix proteins" OR "EMD" OR "Emdogain" OR "amelogenin" OR "ameloblastin" OR "sheath proteins" AND "cells." Results of this analysis, together with current knowledge on the molecular composition of EMD and the structure and regulation of its components, are then used to present a model of EMD functioning. RESULTS: Characterization of the molecular composition of EMD confirmed that amelogenin proteins, including their enzymatically cleaved and alternatively spliced fragments, dominate the protein complex (>90%). A small presence of ameloblastin has also been reported. Analysis of the effects of EMD indicated that gene expression, protein production, proliferation, and differentiation of various cell types are affected and often enhanced by EMD, particularly for periodontal ligament and osteoblastic cell types. EMD also stimulated angiogenesis. In contrast, EMD had a cytostatic effect on epithelial cells. Full-length amelogenin elicited similar effects to EMD, though to a lesser extent. Both the leucine-rich amelogenin peptide and the ameloblastin peptides demonstrated osteogenic effects. A model for molecular structure and functioning of EMD involving nanosphere formation, aggregation, and dissolution is presented. CONCLUSIONS: EMD elicits a regenerative response in periodontal tissues that is only partly replicated by amelogenin or ameloblastin components. A synergistic effect among the various proteins and with the cells, as well as a temporal effect, may prove important aspects of the EMD response in vivo.

Lipid redistribution in phosphatidylserine-containing vesicles adsorbing on titania.

Lipid vesicles (liposomes) exhibit a wide range of behavior at inorganic (oxide) surfaces. A complete understanding of the vesicle-surface interactions, and of the ensuing transformations surface adsorbed liposomes undergo, has proven elusive. This is at least in part due to the large number of degrees of freedom of the system comprising vesicles with their molecular constituents, substrate surface, and electrolyte solution. The least investigated among these degrees of freedom are those intrinsic to the vesicles themselves, involving rearrangements of lipid molecules. In this study, the adsorption of two-component vesicles (phosphatidylcholine:phosphatidylserine) on titanium dioxide was investigated by dual polarization interferometry. Mixtures of these two lipids containing more than 20% of phosphatidylserine form supported bilayers on titania, with phosphatidylserine predominantly facing the surface of the oxide. The purpose of this investigation is to ascertain whether redistribution of phosphatidylserine occurs already in the adsorbing vesicles. Indeed, this was found to be the case. A possible mechanism of this process is discussed.

Micro-well arrays for 3D shape control and high resolution analysis of single cells.

In addition to rigidity, matrix composition, and cell shape, dimensionality is now considered an important property of the cell microenvironment which directs cell behavior. However, available tools for cell culture in two-dimensional (2D) versus three-dimensional (3D) environments are difficult to compare, and no tools exist which provide 3D shape control of single cells. We developed polydimethylsiloxane (PDMS) substrates for the culture of single cells in 3D arrays which are compatible with high-resolution microscopy. Cell adhesion was limited to within microwells by passivation of the flat upper surface through 'wet-printing' of a non-fouling polymer and backfilling of the wells with specific adhesive proteins or lipid bilayers. Endothelial cells constrained within microwells were viable, and intracellular features could be imaged with high resolution objectives. Finally, phalloidin staining of actin stress fibers showed that the cytoskeleton of cells in microwells was 3D and not limited to the cell-substrate interface. Thus, microwells can be used to produce microenvironments for large numbers of single cells with 3D shape control and can be added to a repertoire of tools which are ever more sought after for both fundamental biological studies as well as high throughput cell screening assays.

Nanopatterns with biological functions.

Both curiosity and a desire for efficiency have advanced our ability to manipulate materials with great precision on the micrometer and, more recently, on the nanometer scale. Certainly, the semiconductor and integrated circuit industry has put the pressure on scientist and engineers to develop better and faster nanofabrication techniques. Furthermore, our curiosity as to how life works, and how it can be improved from a medical perspective, stands to gain a great deal from advances in nanotechnology. Novel nanofabrication techniques are opening up the possibilities for mimicking the inherently nano-world of the cell, i.e., the nanotopographies of the extracellular matrix (ECM) and the nanochemistry presented on both the cell membrane and the ECM. In addition, biosensing applications that rely on fabrication of high-density, precision arrays, e.g., DNA or gene chips and protein arrays, will gain significantly in efficiency and, thus, in usefulness once it becomes possible to fabricate heterogeneous nanoarrays. Clearly, continued advances in nanotechnology are desired and required for advances in biotechnology. In this review, we describe the leading techniques for generating nanopatterns with biological function including parallel techniques such as extreme ultraviolet interference lithography (EUV-IL), soft-lithographic techniques (e.g., replica molding (RM) and microcontact printing (muCP)), nanoimprint lithography (NIL), nanosphere lithography (NSL) (e.g., colloid lithography or colloidal block-copolymer micelle lithography) and the nanostencil technique, in addition to direct-writing techniques including e-beam lithography (EBL), focused ion-beam lithography (FIBL) and dip-pen nanolithography (DPN). Details on how the patterns are generated, how biological function is imparted to the nanopatterns, and examples of how these surfaces can and are being used for biological applications will be presented. This review further illustrates the rapid pace by which advances are being made in the field of nanobiotechnology, owing to an increasing number of research endeavors, for an ever increasing number of applications.

Light-induced in situ patterning of DNA-tagged biomolecules and nanoparticles.

We present an in situ method for the selective manipulation of DNA-tagged nano-objects such as vesicles or gold colloids in aqueous solution, at neutral pH. The method makes use of the photosensitizer concept found in photodynamic therapy. Here, single-stranded DNA is immobilized onto a surface via the biotin/streptavidin linkage. If the streptavidin is fluorescently labeled, reactive species will be created during laser-induced photobleaching of the label. These reactive species can then completely or partly suppress the DNA hybridization and cause the removal of the streptavidin. The technique thereby enables a dynamic on-off control over surface density of immobilized DNA-tagged nano-objects. Furthermore, combining this in situ manipulation of DNA with prepatterning of single-stranded DNA in the micro and later in the nano range provides a means for the dynamic patterning required for applications in biosensing and nanotechnology.

Surface engineering approaches to micropattern surfaces for cell-based assays.

The ability to produce patterns of single or multiple cells through precise surface engineering of cell culture substrates has promoted the development of cellular bioassays that provide entirely new insights into the factors that control cell adhesion to material surfaces, cell proliferation, differentiation and molecular signaling pathways. The ability to control shape and spreading of attached cells and cell-cell contacts through the form and dimension of the cell-adhesive patches with high precision is important. Commitment of stem cells to different specific lineages depends strongly on cell shape, implying that controlled microenvironments through engineered surfaces may not only be a valuable approach towards fundamental cell-biological studies, but also of great importance for the design of cell culture substrates for tissue engineering. Furthermore, cell patterning is an important tool for organizing cells on transducers for cell-based sensing and cell-based drug discovery concepts. From a material engineering standpoint, patterning approaches have greatly profited by combining microfabrication technologies, such as photolithography, with biochemical functionalization to present to the cells biological cues in spatially controlled regions where the background is rendered non-adhesive ("non-fouling") by suitable chemical modification. The focus of this review is on the surface engineering aspects of biologically motivated micropatterning of two-dimensional (flat) surfaces with the aim to provide an introductory overview and critical assessment of the many techniques described in the literature. In particular, the importance of non-fouling surface chemistries, the combination of hard and soft lithography with molecular assembly techniques as well as a number of less well known, but useful patterning approaches, including direct cell writing, are discussed.

Creation of a functional heterogeneous vesicle array via DNA controlled surface sorting onto a spotted microarray.

Membrane protein microarrays are expected to play a key role in the future of drug screening and discovery. The authors present a method for the creation of functional heterogeneous vesicle arrays via DNA controlled surface sorting. Complexes of streptavidin and biotinylated DNA are spotted onto a biomolecule- and cell-resistant surface of biotinylated poly(L-lysine)-grafted-poly(ethylene glycol). Two kinds of vesicles functionalized with either the membrane-binding protein annexin A5 or loaded with bovine serum albumin, are tagged with DNA, mixed together, and guided to predefined spots on the surface. The authors show that the spotted complexes remain active and selective and that the background is resistant towards nonspecific adsorption of the vesicles and the proteins.