Biomedical subjects

James C Weaver

Publications and source records attributed to James C Weaver.

30 records · Page 2Linked to original sources

PubMed2004

High-resolution AFM imaging of intact and fractured trabecular bone.

Nanoscale structural analyses of biomineralized materials can frequently help elucidate important structure-function relationships in these complex organic-inorganic composites. Atomic force microscope (AFM) imaging of the exterior surface of trabecular bone reveals a densely woven structure of collagen fibrils, banded with a 67-nm periodicity, and densely packed mineral plates. The mineral plates on the collagen fibrils overlap and exhibit a large range of plate diameters from 30 to 200 nm. On the collagen fibrils, small nodular features, spaced 20-30 nm, run perpendicular to the fibrils. In some cases, these nodules are also seen on filaments extending between collagen fibrils. We hypothesize that these protrusions are noncollagenous proteins such as proteoglycans and may have collapsed into compact structures when the sample was dried. AFM images of pristine fractured surfaces reveal a dense array of mineral plates. In a few isolated locations, short sections of bare collagen fibrils are visible. In other regions, the existence of the underlying collagen fibrils can be inferred from the linear patterns of the mineral plates. Fractured samples, rinsed to remove mineral plates, reveal separated collagen fibrils on the fractured surfaces. These fibrils are often covered with protrusions similar to those observed on the exterior surfaces but are less organized. In addition, as on the exterior surfaces, there are sometimes small filaments extending between neighboring collagen fibrils. These studies provide important insights into the nanostructured architecture of this complex biocomposite.

Animals↗

PubMed2004

Influence of the degradation of the organic matrix on the microscopic fracture behavior of trabecular bone.

In recent years, the important role of the organic matrix for the mechanical properties of bone has become increasingly apparent. It is therefore of great interest to understand the interactions between the organic and inorganic constituents of bone and learn the mechanisms by which the organic matrix contributes to the remarkable properties of this complex biomaterial. In this paper, we present a multifaceted view of the changes of bone's properties due to heat-induced degradation of the organic matrix. We compare the microscopic fracture behavior (scanning electron microscopy; SEM), the topography of the surfaces (atomic force microscopy; AFM), the condition of bone constituents [X-ray diffraction (XRD), thermogravimetric analysis (TGA), and gel electrophoresis], and the macromechanical properties of healthy bovine trabecular bone with trabecular bone that has a heat-degraded organic matrix. We show that heat treatment changes the microfracture behavior of trabecular bone. The primary failure mode of untreated trabecular bone is fibril-guided delamination, with mineralized collagen filaments bridging the gap of the microcrack. In contrast, bone that has been baked at 200 degrees C fractures nondirectionally like a brittle material, with no fibers spanning the microcracks. Finally, bone that has been boiled for 2 h in PBS solution fractures by delamination with many small filaments spanning the microcracks, so that the edges of the microcracks become difficult to distinguish. Of the methods we used, baking most effectively weakens the mechanical strength of bone, creating the most brittle material. Boiled bone is stronger than baked bone, but weaker than untreated bone. Boiled bone is more elastic than untreated bone, which is in turn more elastic than baked bone. These studies clearly emphasize the importance of the organic matrix in affecting the fracture mechanics of bone.

Animals↗

PubMed2004

The tube cement of Phragmatopoma californica: a solid foam.

Phragmatopoma californica is a marine polychaete that builds protective tubes by joining bits of shell and sand grains with a secreted proteinaceous cement. The cement forms a solid foam (closed cells) via covalent crosslinking, as revealed by electron and laser scanning confocal microscopy. The cement contains extractable calcium and magnesium, and non-extractable phosphorus. Amino acid analysis demonstrated that the phosphorus is in the form of phosphoserine and that >90% of serine in the cement (i.e. 28 mol% of residues) is phosphorylated. In addition to previously identified basic proteins, the cement contains a highly acidic polyphosphoserine protein as a major component. We propose a model for the structure and bonding mechanism of the cement that has the following major features: (1) within the secretory pathway of cement gland cells, the electrostatic association of the oppositely charged proteins and divalent cations (Ca2+ and Mg2+) condense the cement proteins into dehydrated secretory granules; (2) the condensation of the cement leads to the separation of the solution into two aqueous phases (complex coacervation) that creates the closed cell foam structure of the cement; (3) rehydration of the condensed cement granules after deposition onto tube particles contributes to the displacement of water from the mineral substrate to facilitate underwater adhesion; and (4) after secretion, covalent cross-linking through oxidative coupling of DOPA gradually solidifies the continuous phase of the cement to set the porous structure.

Amino Acids↗

PubMed2003

Molecular biology of demosponge axial filaments and their roles in biosilicification.

For hundreds of years, the skeletal elements of marine and freshwater sponges have intrigued investigators with a diverse array of remarkably complex morphologies. Early studies of demosponge monaxonal megascleres revealed the presence of a central organic axial filament running their entire length. Until recently, however, the precise function of these axial filaments was largely unknown. The spicules from the temperate Eastern Pacific demosponge, Tethya aurantia, comprise approximately 75% of the dry weight of this species, facilitating the large-scale isolation and purification of the biosilica-associated proteins. Silicateins, the most abundant proteins comprising the axial filaments of these spicules, prove to be members of a well-known superfamily of proteolytic and hydrolytic enzymes and can be easily collected after silica demineralization with hydrofluoric acid. Consistent with these findings, the intact filaments are more than simple, passive templates; in vitro, they actively catalyze and spatially direct the hydrolysis and polycondensation of silicon alkoxides to yield silica at neutral pH and low temperature. Catalytic activity also is exhibited by the monomeric subunits obtained by disaggregation of the protein filaments and those produced from recombinant DNA templates cloned in bacteria. These proteins also can be used to direct the polymerization of organosilicon polymers (silicones) from the corresponding organically functionalized silicon alkoxides. Based on these observations, the silicateins are currently being used as models for the design of biomimetic agents with unique catalytic and structure-directing properties. The presence of axial filaments in a diversity of spicule types and the evolutionary implications of these findings are also discussed.

Amino Acid Sequence↗

PubMed2003

An approach to electrical modeling of single and multiple cells.

Previous theoretical approaches to understanding effects of electric fields on cells have used partial differential equations such as Laplace's equation and cell models with simple shapes. Here we describe a transport lattice method illustrated by a didactic multicellular system model with irregular shapes. Each elementary membrane region includes local models for passive membrane resistance and capacitance, nonlinear active sources of the resting potential, and a hysteretic model of electroporation. Field amplification through current or voltage concentration changes with frequency, exhibiting significant spatial heterogeneity until the microwave range is reached, where cellular structure becomes almost "electrically invisible." In the time domain, membrane electroporation exhibits significant heterogeneity but occurs mostly at invaginations and cell layers with tight junctions. Such results involve emergent behavior and emphasize the importance of using multicellular models for understanding tissue-level electric field effects in higher organisms.

Cells↗

PubMed2003

Characterization of two molluscan crystal-modulating biomineralization proteins and identification of putative mineral binding domains.

Ethylenediamine-tetraacetic acid extracted water-soluble matrix proteins in molluscan shells secreted from the mantle epithelia are believed to control crystal nucleation, morphology, orientation, and phase of the deposited mineral. Previously, atomic force microscopy demonstrated that abalone nacre proteins bind to growing step edges and to specific crystallographic faces of calcite, suggesting that inhibition of calcite growth may be one of the molecular processes required for growth of the less thermodynamically stable aragonite phase. Previous experiments were done with protein mixtures. To elucidate the role of single proteins, we have characterized two proteins isolated from the aragonitic component of nacre of the red abalone, Haliotis rufescens. These proteins, purified by hydrophobic interaction chromatography, are designated AP7 and AP24 (aragonitic protein of molecular weight 7 kDa and 24 kDa, respectively). Degenerate oligonucleotide primers corresponding to N-terminal and internal peptide sequences were used to amplify cDNA clones by a polymerase chain reaction from a mantle cDNA library; the deduced primary amino acid sequences are presented. Preliminary crystal growth experiments demonstrate that protein fractions enriched in AP7 and AP24 produced CaCO(3) crystals with morphology distinct from crystals grown in the presence of the total mixture of soluble aragonite-specific proteins. Peptides corresponding to the first 30 residues of the N-terminal sequences of both AP7 and AP24 were generated. The synthetic peptides frustrate the progression of step edges of a growing calcite surface, indicating that sequence features within the N-termini of AP7 and AP24 include domains that interact with CaCO(3). CD analyses demonstrate that the N-terminal peptide sequences do not possess significant percentages of alpha-helix or beta-strand secondary structure in solution. Instead, in both the presence and absence of Ca(II), the peptides retain unfolded conformations that may facilitate protein-mineral interaction.

Amino Acid Sequence↗

PubMed2003

Nanostructural features of demosponge biosilica.

Recent interest in the optical and mechanical properties of silica structures made by living sponges, and the possibility of harnessing these mechanisms for the synthesis of advanced materials and devices, motivate our investigation of the nanoscale structure of these remarkable biomaterials. Scanning electron and atomic force microscopic (SEM and AFM) analyses of the annular substructure of demosponge biosilica spicules reveals that the deposited material is nanoparticulate, with a mean particle diameter of 74+/-13 nm. The nanoparticles are deposited in alternating layers with characteristic etchant reactivities. Further analyses of longitudinally fractured spicules indicate that each deposited layer is approximately monoparticulate in thickness and exhibits extensive long range ordering, revealing an unanticipated level of nanoscale structural complexity. NMR data obtained from differentially heated spicule samples suggest that the etch sensitivity exhibited by these annular domains may be related to variation in the degree of silica condensation, rather than variability in the inclusion of organics. In addition, AFM phase imaging in conjunction with results obtained from HF and alkaline etching experiments suggest that at various stages in spicule biosynthesis, regions of unusually low silica condensation are deposited, indicating a possible interruption in normal spicule formation. While this discovery of nanoparticulate silica aggregation in demosponge skeletal elements is likely to reflect the intrinsic kinetic tendency of silica to form such particles during polycondensation, the heirarchical organization of these nanoparticles is biologically unique.

Animals↗

PubMed2003

Childhood leukemia: electric and magnetic fields as possible risk factors.

Numerous epidemiologic studies have reported associations between measures of power-line electric or magnetic fields (EMFs) and childhood leukemia. The basis for such associations remains unexplained. In children, acute lymphoblastic leukemia represents approximately three-quarters of all U.S. leukemia types. Some risk factors for childhood leukemia have been established, and others are suspected. Pathogenesis, as investigated in animal models, is consistent with the multistep model of acute leukemia development. Studies of carcinogenicity in animals, however, are overwhelmingly negative and do not support the hypothesis that EMF exposure is a significant risk factor for hematopoietic neoplasia. We may fail to observe effects from EMFs because, from a mechanistic perspective, the effects of EMFs on biology are very weak. Cells and organs function despite many sources of chemical "noise" (e.g., stochastic, temperature, concentration, mechanical, and electrical noise), which exceed the induced EMF "signal" by a large factor. However, the inability to detect EMF effects in bioassay systems may be caused by the choice made for "EMF exposure." "Contact currents" or "contact voltages" have been proposed as a novel exposure metric, because their magnitude is related to measured power-line magnetic fields. A contact current occurs when a person touches two conductive surfaces at different voltages. Modeled analyses support contact currents as a plausible metric because of correlations with residential magnetic fields and opportunity for exposure. The possible role of contact currents as an explanatory variable in the reported associations between EMFs and childhood leukemia will need to be clarified by further measurements, biophysical analyses, bioassay studies, and epidemiology.

Animals↗

PubMed2003

Structure-function studies of the lustrin A polyelectrolyte domains, RKSY and D4.

The lustrin superfamily represents a unique group of biomineralization proteins localized between layered aragonite mineral plates (i.e., nacre layers) in mollusk shell. These proteins not only exhibit elastomeric behavior within the mineralized matrix, but also adhesion to the aragonite-containing composite layer. One member of the lustrin superfamily, Lustrin A, has been sequenced; the protein is organized into defined, modular sequence domains that are hypothesized to perform separate functions (i.e., force unfolding, mineral adhesion, intermolecular binding) within the Lustrin A protein. Using nuclear magnetic resonance (NMR) and in vitro mineralization assays, we investigated structure-function relationships for two Lustrin A putative mineral binding domains, the 30 AA Arg, Lys, Tyr, Ser-rich (RKSY) and the 24 AA Asp-rich (D4) sequence regions domain of the Lustrin A protein. The results indicate that both sequences adopt open, unfolded structures that represent either extended or random coil states. Using geologic calcite overgrowth assays and scanning electron microscopic analyses, we observe that the RKSY polypeptide does not significantly perturb calcium carbonate growth. However, the D4 domain does influence crystal growth in a concentration-dependent manner. Collectively, our data indicate that D4, and not the RKSY domain, exhibits structure-function activity consistent with a mineral binding region.

Amino Acid Sequence↗

PubMed2002

Understanding conditions for which biological effects of nonionizing electromagnetic fields can be expected.

Scientific interest in the interaction of nonionizing electromagnetic fields with biological systems is longstanding, but often still controversial. Theories, models and computer simulations have usually emphasized physical interactions with subsystems (e.g. cell membranes) of a biological system. By extending this first necessary physical step to a second step of explicitly and quantitatively considering chemical changes, increased understanding appears possible. In the case of "strong fields", the role of field-altered chemistry is important to electrochemotherapy [Biochem. Pharmacol. 42, Suppl. (1991) 567] and creation of transdermal microconduits [Bioelectrochem. Bioenerg. 49 (1999) 11; J. Controlled Release 61 (1999) 185; J. Invest. Dermatol. 116 (2001) 40] For "weak fields" (a topic with much more controversy) consideration of chemical change shows that organized multicellular systems can be understood to respond to extremely small electric [Chaos 8 (1998) 576] or magnetic fields [Nature 405 (2000) 707]. In contrast, isolated individual cells interacting via voltage-gated channels [Proc. Natl. Acad. Sci. 92 (1995) 3740; Biophys. J. 75 (1998) 2251; Bioelectromagnetics 20 (1999) 102], or processes without "temperature compensation" [Biophys. J. 76 (1999) 3026], appear implausible. Satisfactory understanding is likely only if experimental and theoretical work is reconciled, which should therefore be emphasized. The interaction of electromagnetic fields with biological systems is of interest because of fundamental scientific curiosity, potential medical benefits and possible human health hazards.

Animals↗

PubMed2002

Theoretical analysis of localized heating in human skin subjected to high voltage pulses.

Electroporation, the increase in the permeability of bilayer lipid membranes by the application of high voltage pulses, has the potential to serve as a mechanism for transdermal drug delivery. However, the associated current flow through the skin will increase the skin temperature and might affect nearby epidermal cells, lipid structure or even transported therapeutic molecules. Here, thermal conduction and thermal convection models are used to provide upper and lower bounds on the local temperature rise, as well as the thermal damage, during electroporation from exponential voltage pulses (70 V maximum) with a 1 ms and a 10 ms pulse time constant. The peak temperature rise predicted by the conduction model ranges from 19 degrees C for a 1 ms time constant pulse to 70 degrees C for the 10 ms time constant pulse. The convection (mass transport) model predicts a 18 degrees C peak rise for 1 ms time constant pulses and a 51 degrees C peak rise for a 10 ms time constant pulse. The convection model compares more favorably with previous experimental studies and companion observations of the local temperature rise during electroporation. Therefore, it is expected that skin electroporation can be employed at a level which is able to transport molecules transdermally without causing significant thermal damage to the tissue.

Electrophysiology↗

PubMed2002

Kinetics of the temperature rise within human stratum corneum during electroporation and pulsed high-voltage iontophoresis.

Electroporation is believed to be a nonthermal phenomenon at the membrane level. However, the effects of associated processes, such as Joule heating, should be considered. Because electroporation of skin, specifically the stratum corneum (SC), occurs at highly localized sites, the heating is expected to conform locally to the sites of electroporation. Significant localized heating was found to be strongly dependent on the voltage and duration of the high-voltage pulses. Specifically, a localized temperature rise was predicted theoretically and confirmed by experiments, with only a small rise (about 17 degrees C) for short, large pulses (1 ms, 100 V across the SC), but was increased (about 54 degrees C) for long, large pulses (300 ms, 60 V across the SC). The latter case appears to result in irreversible structural changes like vesicularization of the lipid lattice. These results support the hypothesis that electroporation occurs within the SC and that additional processes, such as localized heating, may be important.

Electrochemistry↗