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PubMed2004

Minimal intervention dentistry (MI) for South Africa.

The intention of Minimal Intervention Dentistry (MI) is to preserve as much tooth tissue as possible and to offer less threatening care to fearful patients. MI is based on a new understanding of cariology and the therapeutic value of biomimetic materials such as Glass lonomers. It has particular relevance to a new caries classification and employs treatment concepts such as selective caries removal and sealed restorations. The MI approach is appropriate for the vast majority of the population in South Africa; it addresses the public fear of dental treatment. It can be an effective private practice builder and is thus applicable to private and public services.

Biomimetic Materials↗

PubMedDate not supplied

Kinetic radical-scavenging activity of melatonin.

Carbon-centred free radicals can be involved in damage to biological systems under hypoxiclanoxic conditions as well as in ischaemia/reperfusion injury. The antioxidant activities of melatonin against carbon-centred radicals are poorly understood. The aim of this study was to investigate the antioxidant properties of melatonin against carbon-centred radicals in a biomimetic model system consisting of growing methyl methacrylate (MMA) radicals (poly-MMA radicals, PMMA*). The kinetics of the polymerization of MMA initiated by thermal decomposition of 2,2'-azobis(isobutyronitrile) (AIBN; R* radical) or benzoyl peroxide (BPO; PhCOO* radical) in the presence of melatonin were investigated by the induction period method under nearly anaerobic conditions. As melatonin concentrations increased, the length of the induction period (IP) increased, but for the BPO system the IP reached a plateau at a molar ratio of BPO to melatonin of 5:1, indicating that the oxidation of melatonin by PhCOO* was limited. At low concentrations of melatonin, the stoichiometric factor (n, the number of free radicals trapped by the antioxidant moiety) for melatonin was approximately 2, but as the melatonin concentration increased the n value decreased markedly to 0.1. These observations suggest that melatonin may possess catalytic activity contributing to radical avoidance. The initial rate of polymerization (Rp) in the BPO system was markedly suppressed by high concentrations of melatonin, suggesting a strong interaction between oxidative end-products formed from melatonin and PMMA*. Under conditions where n was about 2, the kinh values for melatonin in the BPO system and the AIBN system were 6.58 x 10(4) M(-1)S(-1) and 2.49 x 10(3) M(-1)s(-1), respectively. In the BPO system, the kinh of melatonin was of a similar magnitude to that of a-tocopherol, whereas in the AIBN system the kinh of melatonin was 100-fold greater than that of tocopherol. The present findings suggest that melatonin may be able to scavenge harmful carbon-centred radicals in vivo.

Antioxidants↗

PubMed2006

[Synthetic RGD-containing peptide K16GRGDSPC affected the adhesion, proliferation and osteogenic differentiation of rabbit bone marrow stromal cells on PLGA-[ASP-PEG] scaffold materials].

OBJECTIVE: To explore the effects of synthetic RGD-containing peptide K16GRGDSPC covalent bonding with PLGA-[ASP-PEG] scaffold materials on the adhesion, proliferation and osteogenic differentiation of rabbit bone marrow stromal cells (BMSCs). METHODS: The peptide was synthesized by solid-phase synthesis method and characterized by mass spectrometry and high pressure liquid chromatography. PLGA-[ASP-PEG] scaffold materials were modified with the peptide by cross linker Sulfo-LC-SPDP and detected by XPS. The BMSCs obtained from rabbit were cultured on PLGA-[ASP-PEG] modified with the peptide and those cultured on unmodified PLGA-[ASP-PEG] were also observed as control group. The adhesion and proliferation behaviors of the cells were analyzed by conventional precipitation method, micropipette aspiration technique, MTT assay and Coomassie Brilliant Blue dyes. The osteogenic differentiation of the cells was showed by the activity of alkaline phosphatase (ALP) assayed by ALP Assay Kit and the mRNA levels of ALP, osteocalcin (OCN), osteopontin (OPN) and collagen I assessed by real-time PCR (RT-PCR). Immunofluorescence stain was also used to detect the expression of core binding factor a1 (Cfba1) which was an osteogenic maker as well. RESULTS: The peptide was successfully manufactured and linked to the surface of the PLGA-[ASP-PEG] by the cross-linker. The abilities of adhesion and proliferation and the expressions of osteogenic makers of the cells were significantly higher than those of control group (P < 0.05). CONCLUSION: RGD-containing peptide K16GRGDSPC could promote the adhesion, proliferation and osteogenic differentiation of BMSCs on the biomimetic bone-matrix materials.

Alkaline Phosphatase↗

PubMed1998

Assembly of phosphofructokinase-1 from Saccharomyces cerevisiae in extracts of single-deletion mutants.

Phosphofructokinase-1 from Saccharomyces cerevisiae is an octameric enzyme comprising two non-identical subunits, alpha and beta, which are encoded by the unlinked genes PFK1 and PFK2. In this paper, assembly and reactivation of the enzyme have been studied in cell-free extracts of single-deletion mutants. In contrast to the previously described lack of phosphofructokinase-1 activity in cell-free extracts of these mutants, we could measure a temporary enzyme activity immediately after lysis of protoplasts. This result supports the assumption that each of the subunits forms an enzyme structure which is active in vivo but not stable after cell disruption. Upon mixing of separately prepared cell-free extracts of both deletion mutants very low activity could be measured. About 40% of the wild-type activity was regained when both mutants were mixed prior to disruption. The reactivation rate could be slightly increased by addition of ATP and fructose 6-phosphate and was found to be a function of the growth state, particularly of the beta-subunit-carrying cells. The individual subunits did not interact with Cibacron Blue F3G-A, a biomimetic ligand of phosphofructokinase-1. After reassembly of both subunits in vitro a strong affinity of the reconstituted phosphofructokinase-1 to the dye-ligand was observed. The inability of the subunits to reconstitute under certain conditions seems to result from alterations of the intracellular environment following disruption. These changes give rise to induce an unproductive side reaction like self-aggregation of the subunits. Because reconstitution of phosphofructokinase-1 from S. cerevisiae behaves in a similar way to that of hemoglobin and luciferase, we would speculate a general mechanism for assembly of oligomeric proteins in vivo.

Blotting, Western↗

PubMed1998

Protein-like molecular architecture: biomaterial applications for inducing cellular receptor binding and signal transduction.

The development of biomaterials with desirable biocompatibility has presented a difficult challenge for tissue engineering researchers. First and foremost, materials themselves tend to be hydrophobic and/or thrombogenic in nature, and face compatibility problems upon implantation. To mediate this problem, researchers have attempted to graft protein fragments onto biomaterial surfaces to promote endothelial cell attachment and minimize thrombosis. We envisioned a novel approach, based on the capability of biomolecules to self-assemble into well-defined and intricate structures, for creating biomimetic biomaterials that promote cell adhesion and proliferation. One of the most intriguing self-assembly processes is the folding of peptide chains into native protein structures. We have developed a method for building protein-like structural motifs that incorporate sequences of biological interest. A lipophilic moiety is attached onto a N alpha-amino group of peptide chain, resulting in a "peptide-amphiphile." The alignment of amphiphilic compounds at the lipid-solvent interface is used to facilitate peptide alignment and structure initiation and propagation, while the lipophilic region absorbs to hydrophobic surfaces. Peptide-amphiphiles containing potentially triple-helical or alpha-helical structural motifs have been synthesized. The resultant head group structures have been characterized by CD spectroscopy and found to be thermally stable over physiological temperature ranges. Triple-helical peptide-amphiphiles have been applied to studies of surface modification and cell receptor binding. Cell adhesion and spreading was promoted by triple-helical peptide-amphiphiles. Cellular interaction with the type IV collagen sequence alpha 1(IV) 1263-1277 increased signal transduction, with both the time and level of induction dependent upon triple-helical conformation. Collectively, these results suggest that peptide-amphiphiles may be used to form stable molecular structure on biomaterial surfaces that promote cellular activities and improve biocompatibility.

Amino Acid Sequence↗

PubMed1998

Examination of protein adsorption in fluidized bed and packed bed columns at different temperatures using frontal chromatographic methods.

The influences of various experimental parameters on the dynamic adsorption capacity (DAC) and the dynamic adsorption rate (DAR) of a biomimetic affinity silica-based adsorbent in fluidized and packed bed columns operated under plug flow conditions and at different temperatures have been investigated with different inlet concentrations of hen egg white lysozyme (HEWL) and human serum albumin (HSA). The DACs as well as the DARs of both the fluidized and packed beds were examined at 10% saturation (i.e., at the QB value) and the experimental data compared with the corresponding data obtained from batch equilibrium adsorption procedures. Parameters examined included the fluid superficial velocity and protein concentration and their effect on the binding capacity and column efficiency. Consistent with various results reported from this and other laboratories on the behavior of biospecific affinity adsorbents derived from porous silica and zirconia particles, adsorbents prepared from Fractosil 1000 were found to exhibit appropriate rheological characteristics in fluidized bed systems under the experimental conditions. Moreover, changes in temperature resulted in a more significant effect on the breakthrough profiles of HSA compared to HEWL with the immobilized Cibacron Blue F3G-A with Fractosil 1000 adsorbent. This result suggests that temperature effects can possibly be employed profitably in some processes as part of a strategy to enhance column performance with fluidized bed systems for selective recovery of target proteins. At relatively low superficial velocities of the feed, the DARs with HEWL and HSA were similar for both the fluidized and packed bed column systems, whereas, at high superficial velocities, the DARs for these proteins were larger with the packed bed columns.

Adsorption↗

PubMed1998

Osteoclastic resorption of bone-like apatite formed on a plastic disk as an in vitro assay system.

We have investigated the applicability of a simple and inexpensive osteoclastic assay system using bone-like apatite-coated polyethyleneterephthalate (PET) disks. A 1 microm thick apatite layer, uniform and homogeneous bone-mineral-like with no organic components, was made on PET disks using a biomimetic process. As substrates for an osteoclastic assay, these coated disks were compared with dentine as well as with bone-like or heat-treated apatite of various thicknesses on apatite- and wollastonite-containing glass ceramic (A-W GC) disks. The unfractionated bone cells, including osteoclasts, of a neonatal rabbit were seeded onto these substrates. By scanning electron microscopic examination, the resorption lacunae of the thick bone-like apatite clearly showed track-like shapes at various depths, similar to those of dentine although the border between the A-W GC and the apatite was unclear. In contrast, those of heat-treated apatite showed small and shallow shapes with irregular margins, quite different from those of dentine. By reducing the thickness of bone-like apatite to 1 microm as well as using PET as its substrate, the margins of the resorption lacunae became quite clear, and with the use of phase-contrast microscopy during culture, osteoclasts and resorption pits could be precisely observed. The resorbed area, easily measured with the aid of bright-field microscopy and an image analyzer, was found to have increased in a time-dependent manner and at the end of 4 days of culture was not statistically different from that of dentine.

Animals↗

PubMed1999

Composition and structure of the apatite formed on PET substrates in SBF modified with various ionic activity products.

An apatite layer was formed on polyethyleneterephthalate (PET) substrates by the following biomimetic process. PET substrates were placed on granular particles of a CaO-SiO2-based glass in simulated body fluid (SBF) with ion concentrations nearly equal to those of human blood plasma to form apatite nuclei on their surfaces (first treatment). They then were soaked in modified SBFs, the ion concentrations of which were changed to give a variation in ionic activity product of apatite (IP), in order to make the apatite nuclei grow (second treatment). The Ca/P atomic ratio and the lattice constant c of the formed apatite decreased from 1.54 to 1.40 and from 6.880 to 6.838 A, respectively, with increasing ion concentrations from 0.75 to 2.00 times those of SBF, that is, with increasing IP from 10(-96.6) to 10(-91.9). This was attributed to an increase in the concentration of HPO4(2-) ion substituting for the PO4(3-) ion sites, which gave an increase in the Ca2+ in the apatite. Even the apatite formed in 1.00 SBF showed a Ca/P ratio of 1.51 and lattice constants a of 9.432 A and c of 6.870 A. The Ca/P ratio and lattice constant c were smaller and the lattice constant a was larger than those of the bone apatite; its Ca/P ratio and its lattice constants a and c, were 1.65, 9.419 A, and 6.88 A, respectively. This was attributed to the lower content (2.64 wt%) of the CO3(2-) ion substituting for the PO4(3-) ion sites of the apatite compared to that of the bone apatite (5.80 wt%). The lower content of the CO3(2-) ion in the apatite might be caused by the lower concentration of HCO3- ion in 1.00 SBF compared to that in human blood plasma.

Apatites↗

PubMed2000

Growth of continuous bonelike mineral within porous poly(lactide-co-glycolide) scaffolds in vitro.

Strategies to engineer bone have focused on the use of natural or synthetic degradable materials as scaffolds for cell transplantation or as substrates to guide bone regeneration. The basic requirements of the scaffold material are biocompatibility, degradability, mechanical integrity, and osteoconductivity. A major design problem is satisfying each of these requirements with a single scaffold material. This study addresses this problem by describing an approach to combine the biocompatibility and degradability of a polymer scaffold with the osteoconductivity and mechanical reinforcement of a bonelike mineral film. We report the nucleation and growth of a continuous carbonated apatite mineral on the interior pore surfaces of a porous, degradable polymer scaffold via a one step, room temperature incubation process. A 3-dimensional, porous scaffold of the copolymer 85:15 poly(lactide-co-glycolide) was fabricated by a solvent casting, particulate leaching process. Fourier transform IR spectroscopy and scanning electron microscopy (SEM) analysis after different incubation times in a simulated body fluid (SBF) demonstrate the growth of a continuous bonelike apatite layer within the pores of the polymer scaffold. Quantification of phosphate on the scaffold displays the growth and development of the mineral film over time with an incorporation of 0.43 mg of phosphate (equivalent to 0.76 mg of hydroxyapatite) per scaffold after 14 days in SBF. The compressive moduli of polymer scaffolds increased fivefold with formation of a mineral film after a 16-day incubation time as compared to control scaffolds. In summary, this biomimetic treatment provides a simple, one step, room temperature method for surface functionalization and subsequent mineral nucleation and growth on biodegradable polymer scaffolds for tissue engineering.

Analysis of Variance↗

PubMedDate not supplied

Affinity chromatography of yeast alcohol dehydrogenase using immobilized monochlorotriazine colourless compounds.

Four new symmetrical colourless biomimetics bearing two monochlorotriazine rings have been rationally designed and synthesized. These immobilized colourless ligands and one analogue of Cibacron Blue F3GA on Sepharose CL-4B were used to purify alcohol dehydrogenase from baker's yeast extract. Twenty-two-fold purification with 78% enzyme recovery was achieved in a single step with specific elution of NAD+ (5 mM) from the colourless absorbent comprised of two ortho-aminobenzene sulphonates as terminal rings, which is comparable to the result obtained from the analogue of Cibacron Blue F3GA. Differential spectra between ligand-enzyme complexes and free ligands were studied.

Alcohol Dehydrogenase↗

PubMedDate not supplied

The ion channel switch biosensor.

A biosensor technology is described which provides a direct measurement for functional molecular interactions, at the surface of a tethered bilayer membrane, through the electrical transduction of chemically modified ion-channels. High sensitivity of analyte detection is achieved due to the large flux of ions transmitted through the ion channel. The biomimetic sensor surface allows the molecular recognition to be measured in complex biological matrices (such as blood and sera) without compromising sensitivity. We have used the sensor for activity and concentration measurements for a range of analytes, which include bacteria, DNA, proteins and drugs. We have a quantitative model for the biosensor performance which is described by three-dimensional molecular interactions with the membrane surface and two-dimensional molecular interactions within the tethered bilayer.

Biosensing Techniques↗

PubMed2000

Introduction of heteroatom-based substituents into 1,4-dihydropyridines by means of a halogen-mediated, oxidative protocol: diamination, sulfonylation, sulfinylation, bis-sulfanylation, and halo-phosphonylation processes

The natural tendency of 1,4-dihydropyridines to undergo "biomimetic" oxidation to afford pyridinium salts can be switched off and, through the use of reagents that interact electrophilically with the enamine moiety present in the heterocyclic system, it is possible to promote alternative oxidations. In this way, efficient regio- and stereocontrolled synthetic methods have been developed that lead to diversely substituted di- and tetrahydropyridines. These include iodoazidation, diamination, bis-sulfonamidation, sulfonylation, sulfinylation, thiocyanation, sulfanylation, bis-sulfanylation, and halo-phosphonylation processes.

Journal Article↗

PubMed1999

New Routes to Hydrogen Peroxide: Alternatives for Established Processes?

The reduction of dioxygen to hydrogen peroxide occurs in the presence of copper and palladium catalysts. The first mentioned biomimetic process uses primary alcohols as the reductant, while in the second, in which the key feature is the use of chelating nitrogen ligands, CO fulfills this function (see scheme). Whether these two new methods will become commercial scale and compete with the established anthraquinone method for production remains unanswered.

Journal Article↗

PubMed2001

Mechanical properties of the dentinoenamel junction: AFM studies of nanohardness, elastic modulus, and fracture.

The dentinoenamel junction (DEJ) is a complex and poorly defined structure that unites the brittle overlying enamel with the dentin that forms the bulk of the tooth. In addition, this structure appears to confer excellent toughness and crack deflecting properties to the tooth, and has drawn considerable interest as a biomimetic model of a structure uniting dissimilar materials. This work sought to characterize the nanomechanical properties in the region of the DEJ using modified AFM based nanoindentation to determine nanohardness and elastic modulus. Lines of indentations traversing the DEJ were made at 1-2 microm intervals from the dentin to enamel along three directions on polished sagittal sections from three third molars. Nanohardness and elastic modulus rose steadily across the DEJ from bulk dentin to enamel. DEJ width was estimated by local polynomial regression fits for each sample and location of the mechanical property curves for the data gradient from enamel to dentin, and gave a mean value of 11.8 microm, which did not vary significantly with intratooth location or among teeth. Nanoindentation was also used to initiate cracks in the DEJ region. In agreement with prior work, it was difficult to initiate cracks that traversed the DEJ, or to produce cracks in the dentin. The fracture toughness values for enamel of 0.6-0.9 MPa . m(1/2) were in good agreement with recent microindentation fracture results. Our results suggest that the DEJ displays a gradient in structure and that nanoindenation methods show promise for further understanding its structure and function.

Algorithms↗

PubMed2001

Fibronectin modulates macrophage adhesion and FBGC formation: the role of RGD, PHSRN, and PRRARV domains.

To probe the role of human plasma fibronectin in modulating human blood-derived macrophage adhesion and fusion to form multinucleated foreign-body giant cells (FBGC), a series of biomimetic oligopeptides based on the functional structure of fibronectin was designed and synthesized. Peptides incorporated the RGD and PHSRN integrin-binding sequences from FIII-10 and FIII-9 modules, respectively, and the PRRARV sequence from the C-terminal heparin-binding domain, either alone or in combination. Peptides were immobilized onto a polyethyleneglycol-based polymer substrate. The following conclusions were reached. Fibronectin modulated macrophage adhesion and the extent (i.e., size) of FBGC formation on control surfaces in the presence of serum proteins. Macrophages adhered to all substrates with relatively subtle differences between adhesion mediated by RGD, PHSRN, PRRARV, or combinations thereof. beta1-integrin subunit was essential in macrophage adhesion to peptide-grafted networks in a receptor-peptide specific manner, whereas beta3-integrin subunit was less important. Macrophage adhesion to PRRARV was mediated primarily by the direct interaction with integrins. RGD or PHSRN alone did not provide an adequate substrate for macrophage fusion to form FBGCs. However, the PHSRN synergistic site and the RGD site in a single oligopeptide provided a substrate for FBGC formation that was statistically comparable to that on the positive control material in the presence of serum proteins. This response was highly dependent upon the relative orientation between RGD and PHSRN. PRRARV did not support FBGC formation. These results demonstrate the importance of fibronectin and, specifically, the synergy between RGD and PHSRN domains, in supporting macrophage fusion to form FBGCs.

Amino Acid Sequence↗

PubMed2000

Multicomponent domino reactions for the synthesis of biologically active natural products and drugs.

A main issue in modern synthetic organic chemistry, which deals with the preparation of natural products, pharmaceuticals, diagnostics, agrochemicals, and other important materials, is the improvement of efficiency, the avoidance of toxic reagents, the reduction of waste, and the responsible treatment of our resources. One of the ways to fulfill these goals is the development and use of domino processes, which consist of several bond-forming reactions and which allow the highly efficient synthesis of complex molecules starting from simple substrates. Herein, the combination of several catalytic bond-forming transformations is clearly most appropriate. The synthesis of the enantiopure alkaloid (-)-hirsutine 22, which has a strong inhibitorial effect on influenza A viruses, was accomplished using a biomimetic domino Knoevenagel-hetero-Diels Alder-solvolysis-hydrogenation process. In a similar way the alkaloids (+)-dihydrocorynantheine 23 and (-)-dihydroantirhine 24 as well as heterosteroids 62, D-homosteroids 65 and 68, and azasteroids 25 are prepared. In addition, novel steroid alkaloids 26 are accessible by a combination of the formation of an iminium salt, a hydride shift, and an alkylation. The anti-leukemic pentacyclic (-)-cephalotaxine 27 is obtained by a combination of two Pd-catalyzed reactions.

Biological Products↗

PubMed2001

Bioorganic applications of semisynthetic DNA-protein conjugates.

Semisynthetic DNA-protein conjugates are versatile molecular tools useful, for instance, in the self-assembly of high-affinity reagents for immunological detection assays, the fabrication of highly functionalized laterally microstructured biochips, and the biomimetic "bottom-up" synthesis of nanostructured supramolecular devices. This concept paper summarizes the current state-of-the-art concerning the synthesis, characterization, and applications of such hybrid molecules, and also draws perspectives on future developments.

Binding Sites↗

PubMed2001

Iron porphyrins as models of cytochrome c oxidase.

A series of iron porphyrins has been synthesized as models of cytochrome c oxidase; their activity as 4e catalysts in the reduction of dioxygen has been studied at pH 7. These compounds have been obtained by grafting very different residues onto the same iron complex, namely tripodal tetraamines, pickets, and straps, in order to change the environment of the metal center. In the case of porphyrins bearing a tripodal cap, the secondary amines have been alkylated with different substituents so as to modify the electronic environment of the distal pocket. Surprisingly, when the iron porphyrin is functionalized with four identical acrylamido pickets, the resulting complex exhibits biomimetic activity in that it catalyzes oxygen reduction with almost no production of hydrogen peroxide. The crystal structure of the redox-inactive zinc(II) analogue is reported; this shows how the metal influences the spatial arrangement of the four pickets through axial coordination and hydrogen bonding. Even a bis-strapped iron porphyrin, for which no dimerization or self-aggregation can occur at the electrode surface, acts as a 4e catalyst for O2 reduction. It is thus demonstrated that at pH close to physiological values, the iron porphyrin is an intrinsically efficient catalyst for the reduction of oxygen to water.

Catalysis↗