Synthesis and characterization of peptide grafted porous polymer microstructures

A scanning laser system has been used to generate three-dimensional trimethylolpropane trimethacrylate (TRIM) cross-linked poly(2-hydroxylethyl methacrylate) polymer microstructures through azo-bis(isobutyro)nitrile (AIBN) photopolymerization using a 20 x 0.5 NA microscope objective and 365 nm laser excitation. Macropores are observed to form without the use of porogens in regions of highest light flux. This is attributed to phase separation, which results from differences in monomer reactivity and miscibility. The microstructures were aminated and then protected with the photolabile protective group 6-nitroveratryloxycarbonyl (NVOC). This made it possible to selectively modify the microstructures with the same scanning laser system that was used to fabricate them, resulting in peptide grafted three-dimensional porous microstructures. On the basis of the absorbance of the dibenzofulvene-piperidine, these structures have an amine site density of approximately 0.1 nmol/feature. MALDI-TOF MS was used to characterize peptide photografted microstructures. N-Tris(2,4,6-trimethoxyphenyl)phosphonium (TMPP) labeling of the peptides greatly enhanced detection and allowed post-source decay sequencing of the peptides from the microstructures. The techniques described could be used to generate three-dimensional peptide grafted porous scaffolds for tissue engineering applications.     

A scalar analysis of landscape connectivity

Landscape connectivity is critical to the maintenance of spatially-structured populations and consists of both a structural component, which describes the shape, size and location of landscape features; and a biological component, which consists of both the response of individuals to landscape features, and the patterns of gene flow that result from those individual responses. Traditional studies of landscape connectivity have attempted to discern individual behavioral responses to landscape features, but this methodology is intractable for many species. This paper is an attempt to relate the components of landscape connectivity through the explicit treatment of their spatial and temporal scales. Traditional measures of structural and biological components of connectivity are reviewed and more recently developed methods for the analysis of scale for each are introduced. I then present a framework for the comparison of scalar phenomena based on Watt's unit pattern, describe the potential outcomes of the comparison and discuss the implications of each. Several testable hypotheses emerge from the analysis that may serve as a useful framework for the investigation of landscape connectivity in the future .     

A stimulus-dependent connectivity analysis of neuronal networks

We present an analysis of interactions among neurons in stimulus-driven networks that is designed to control for effects from unmeasured neurons. This work builds on previous connectivity analyses that assumed connectivity strength to be constant with respect to the stimulus. Since unmeasured neuron activity can modulate with the stimulus, the effective strength of common input connections from such hidden neurons can also modulate with the stimulus. By explicitly accounting for the resulting stimulus-dependence of effective interactions among measured neurons, we are able to remove ambiguity in the classification of causal interactions that resulted from classification errors in the previous analyses. In this way, we can more reliably distinguish causal connections among measured neurons from common input connections that arise from hidden network nodes. The approach is derived in a general mathematical framework that can be applied to other types of networks. We illustrate the effects of stimulus-dependent connectivity estimates with simulations of neurons responding to a visual stimulus. This research was supported by the National Science Foundation grants DMS-0415409 and DMS-0748417.     

Spectral analysis of nucleotide sequences

The data of Fourier-analysis of nucleotide sequences are discussed. The existence of reflexes corresponding to regular position of nucleotides (mainly T and G) with 3-base period is the most striking feature of both phage and viral nucleic acid sequences spectra. The amplitude and phase of the similar reflexes in the dinucleotide spectra obtained by digital computing of Fourier-transform, give specific information on amino acid composition, codon bias, amino acid relations. The width of frequency band characterizes a tendency to nucleotide clustering or to separate existence. The blurring of reflexes shows the disturbance of far order in the regular nucleotide "lattice". The two-dimensional spectral analysis supports the existence of far correlation in nucleotide positions.     

Spectral analysis of breathing pattern in man

The periodic oscillations of breathing pattern parameters were studied in 34 healthy subjects. In a three minutes' resting spirometric recording we determined the duration of inspiration, expiration and tidal volume in successive breaths and computed autocorrelation functions and power spectral density. Ten of the subjects were re-examined 2 years later. Pulmonary functions were examined in all of them. Rhythmic changes lasting several respiratory cycles were found in the breathing pattern. Rhythmic changes in the duration of inspiration, expiration and tidal volume differed from one another in the same individual. The spectrograms of the individual breathing pattern parameters in the same individual changed during a 2-year period. Despite individual differences, the power spectral density correlations in the same subject and between different subjects two years later, and the mean curves for power spectral densities, show that in all the subjects the power fell at values of 0 to 0.05 Hz and was then maintained at a roughly constant level. Differences in the spectrograms of the various parameters in the same subject can hardly be attributed to a feedback between peripheral receptors and respiratory centres. Rhythmic changes are probably of central origin.     

Spectral and spectral-temporal heart rate analysis in newborns

Spectral (in 3-min segments) and spectral-temporal (in 36-s windows with a 9-s step) analyses were carried out on the basis of cardiointerval recordings of newborns (full-term and preterm) during 24 h. It was found that each of the 3-min intervals of the 24-h cardiointerval recordings in the newborns had a unique spectrum; the spectra could be combined into several types. During 24 h, a spectrum with a marked LF component prevailed in the newborns. A spectrum with a peak in the HF region recorded in the newborns in the range from 0.4 to ～1 Hz was an indicator of the rhythmic pattern of the variation of cardiointervals related to respiratory waves. Owing to the diversity of patterns of the spectral behavior of cardiointervals in a newborn during 24 h, no single short segment of the cardiointerval recording could serve as an unambiguous characteristic of the newborn’s physiological state. The actual heart rate variability pattern could be seen by means of the spectraltemporal analysis, visualizing the dynamics of conversion of one spectral type to another, i.e., the change of the dominant system of regulation.     

Surface porous fibre-reinforced composite bulk bone substitute

The aim of this study was to describe and evaluate the significance of a porous surface with bioactive glass granules (S53P4) covering an artificial bulk material based on polymethylmetacrylate (PMMA) and fibre-reinforced composite (FRC) technology. Effort was focused particularly on characters of the porous surface and biomechanical properties of the material in vitro, and test in vivo the implant in reconstruction in an experimental long bone segment defect model. The defect, 10 mm in length, created in the shaft of rabbit tibia, was reconstructed by the implant and fixed by intramedullary K-wires. The implant was incorporated within 4 weeks by new bone growth from the host bone covering particularly its posterior surface and cortex/implant junctions with bridging trabecular bone. Later, at 8 weeks, new bone was found also at the cortex/implant interface and in the medullary canal of the implant. Histometric measurements revealed direct bone/implant surface contact in 34% at the interface. Bioactive glass granules in the porous surface evoked the most direct contact with bone. The implants manufactured from PMMA only served as a control group, and showed significantly lower osteoconductive properties. Biomechanical measurements in vitro of fibre-reinforced PMMA specimens revealed values for bending strength and the flexural modulus to match them to human bone. This artificial bulk bone material based on PMMA/FRC technology seems to have proposing properties to be used as a bone substitute on load-bearing conditions. This revised version was published online in July 2006 with corrections to the Cover Date.     

Detection and estimation of neural connectivity based on crosscorrelation analysis

Crosscorrelation analysis of simultaneously recorded activity of pairs of neurons is a common tool to infer functional neural connectivity. The adequacy of crosscorrelation procedures to detect and estimate neural connectivity has been investigated by means of computer simulations of small networks composed of fairly realistic modelneurons. If the mean interval of neural firings is much larger than the duration of postsynaptic potentials, which will be the case in many central brain areas excitatory connections are easier to detect than inhibitory ones. On the other hand, inhibitory connections are revealed better if the mean firing interval is much smaller than post-synaptic potential duration. In general the effects of external stimuli and the effects of neural connectivity do not add linearly. Furthermore, neurons may exhibit a certain degree of timelock to the stimulus. For these reasons the commonly applied shift predictor procedure to separate stimulus and neural effects appears to be of limited value. In case of parallel direct and indirect neural pathways between two neurons crosscorrelation analysis does not estimate the direct connection but instead an effective connectivity, which reflects the combined influences of the parallel pathways.Abbreviations ACH autocoincidence histogram - CCH crosscoincidence histogram - CPDF crossproduct density function - DCH difference crosscoincidence histogram - NACH nonsimultaneous autocoincidence histogram - NCCH nonsimultaneous crosscoincidence histogram - SCCH scaled crosscoincidence histogram - SDCH scaled difference crosscoincidence histogram     

Coherency and connectivity in oscillating neural networks: linear partialization analysis

 This paper studies the relation between the functional synaptic connections between two artificial neural networks and the correlation of their spiking activities. The model neurons had realistic non-oscillatory dynamic properties and the networks showed oscillatory behavior as a result of their internal synaptic connectivity. We found that both excitation and inhibition cause phase locking of the oscillating activities. When the two networks excite each other the oscillations synchronize with zero phase lag, whereas mutual inhibition between the networks resulted in an anti-phase (half period phase difference) synchronization. Correlations between the activities of the two networks can also be caused by correlated external inputs driving the systems (common input). Our analysis shows that when the networks exhibit oscillatory behavior and the rate of the common input is smaller than a characteristic network oscillator frequency, the cross-correlation functions between the activities of two systems still carry information about the mutual synaptic connectivity. This information can be retrieved with linear partialization, removing the influence of the common input. We further explored the network responses to periodic external input. We found that when the input is of a frequency smaller than a certain threshold, the network responds with bursts at the same frequency as the input. Above the threshold, the network responds with a fraction of the input frequency. This frequency threshold, characterizing the oscillatory properties of the network, is also found to determine the limit to which linear partialization works. Received: 20 October 1995 / Accepted in revised form: 20 May 1996     

Network analysis of intrinsic functional brain connectivity in Alzheimer's disease

Functional brain networks detected in task-free (“resting-state”) functional magnetic resonance imaging (fMRI) have a small-world architecture that reflects a robust functional organization of the brain. Here, we examined whether this functional organization is disrupted in Alzheimer's disease (AD). Task-free fMRI data from 21 AD subjects and 18 age-matched controls were obtained. Wavelet analysis was applied to the fMRI data to compute frequency-dependent correlation matrices. Correlation matrices were thresholded to create 90-node undirected-graphs of functional brain networks. Small-world metrics (characteristic path length and clustering coefficient) were computed using graph analytical methods. In the low frequency interval 0.01 to 0.05 Hz, functional brain networks in controls showed small-world organization of brain activity, characterized by a high clustering coefficient and a low characteristic path length. In contrast, functional brain networks in AD showed loss of small-world properties, characterized by a significantly lower clustering coefficient (p<0.01), indicative of disrupted local connectivity. Clustering coefficients for the left and right hippocampus were significantly lower (p<0.01) in the AD group compared to the control group. Furthermore, the clustering coefficient distinguished AD participants from the controls with a sensitivity of 72% and specificity of 78%. Our study provides new evidence that there is disrupted organization of functional brain networks in AD. Small-world metrics can characterize the functional organization of the brain in AD, and our findings further suggest that these network measures may be useful as an imaging-based biomarker to distinguish AD from healthy aging.     

Spectral K-edge subtraction imaging of experimental non-radioactive barium uptake in bone

PurposeTo evaluate the feasibility of using non-radioactive barium as a bone tracer for detection with synchrotron spectral K-edge subtraction (SKES) technique.MethodsMale rats of 1-month old (i.e., developing skeleton) and 8-month old (i.e., skeletally mature) were orally dosed with low dose of barium chloride (33 mg/kg/day Ba²⁺) for 4 weeks. The fore and hind limbs were dissected for imaging in projection and computed tomography modes at 100 μm and 52 μm pixel sizes. The SKES method utilizes a single bent Laue monochromator to prepare a 550 eV energy spectrum to encompass the K-edge of barium (37.441 keV), for collecting both ‘above’ and ‘below’ the K-edge data sets in a single scan.ResultsThe SKES has a very good focal size, thus limits the ‘crossover’ and motion artifacts. In juvenile rats, barium was mostly incorporated in the areas of high bone turnover such as at the growth plate and the trabecular surfaces, but also in the cortical bone as the animals were growing at the time of tracer administration. However, the adults incorporated approximately half the concentration and mainly in the areas where bone remodeling was predominant and occasionally in the periosteal and endosteal layers of the diaphyseal cortical bone.ConclusionsThe presented methodology is simple to implement and provides both structural and functional information, after labeling with barium, on bone micro-architecture and thus has great potential for in vivo imaging of pre-clinical animal models of musculoskeletal diseases to better understand their mechanisms and to evaluate the efficacy of pharmaceuticals.     

Spectral karyotyping analysis of human and mouse chromosomes

Classical banding methods provide basic information about the identities and structures of chromosomes on the basis of their unique banding patterns. Spectral karyotyping (SKY), and the related multiplex fluorescence in situ hybridization (M-FISH), are chromosome-specific multicolor FISH techniques that augment cytogenetic evaluations of malignant disease by providing additional information and improved characterization of aberrant chromosomes that contain DNA sequences not identifiable using conventional banding methods. SKY is based on cohybridization of combinatorially labeled chromosome-painting probes with unique fluorochrome signatures onto human or mouse metaphase chromosome preparations. Image acquisition and analysis use a specialized imaging system, combining Sagnac interferometer and CCD camera images to reconstruct spectral information at each pixel. Here we present a protocol for SKY analysis using commercially available SkyPaint probes, including procedures for metaphase chromosome preparation, slide pretreatment and probe hybridization and detection. SKY analysis requires approximately 6 d.     

Spectral analysis and fingerprinting for biomedia characterisation

Classical culture media, as well as domestic and/or industrial wastewater treated by biological processes, have a complex composition. The on-line and/or in situ determination of some substances is possible, but expensive, as sample collection and pre-treatment are often necessary with strict rules of sterility. More global methods can be used to detect rapidly "accidents" such as the appearance of an undesirable by-product in a fermentation broth or of a toxic substance in wastewater. These methods combine a "hard" part, for sensing, and a "soft" part, for data treatment. Among potential "hard" candidates, spectroscopy can be the basis for non-invasive and non-destructive measuring systems. Some of them have been already tested in situ: ultra-violet-visible, infra-red (mid or near), fluorescence (mono-dimensional, two-dimensional or synchronous), dielectric, while others, more sophisticated, such as mass spectrometry, coupled or not to pyrolysis, nuclear magnetic resonance and Raman spectroscopy, have been proposed. All these methods provide spectra, i.e. large sets of data, from which meaningful information should be rapidly extracted, either for analysis or fingerprinting. The recourse to data-mining techniques (the "soft" part) such as principal components analysis, projection on latent structures or artificial neural networks, is a necessary step for that task. A review of techniques, mostly based on spectroscopy, with examples taken in the bioengineering field in general is proposed.     

Spectral analysis of cytochromes inAquaspirillum magnetotacticum

The respiratory chain ofAquaspirillum magnetotacticum strain MS-1 cells denitrifying microaerobically included a-, a₁-, b-, c-, cd₁-, and o-type hemes. More than 85% of the total cytochromes detected were of the c type. Virtually all of the a and b types were detected in cell membranes, whereas 70% of the c-type hemes were soluble. Large quantities of soluble c-type hemes were released with periplasm by freezing and thawing cells. Soluble c₅₅₁ occurred in two forms: as a single compound of apparent molecular weight of 17,000 daltons, which bound CO, and, together with d₁ heme, as a component of nitrite reductase. Both a₁-type hemes (which usually comprise part of the low aeration cytochrome oxidase) and o types (usually part of the high aeration oxidase) were simultaneously expressed in microaerobically grown denitrifying cells ofA. magnetotacticum; this indicated branching of the respiratory chain.     

Modeling of dynamic fracture and damage in two-dimensional trabecular bone microstructures using the cohesive finite element method

Trabecular bone fracture is closely related to the trabecular architecture, microdamage accumulation, and bone tissue properties. Micro-finite-element models have been used to investigate the elastic and yield properties of trabecular bone but have only seen limited application in modeling the microstructure dependent fracture of trabecular bone. In this research, dynamic fracture in two-dimensional (2D) micrographs of ovine (sheep) trabecular bone is modeled using the cohesive finite element method. For this purpose, the bone tissue is modeled as an orthotropic material with the cohesive parameters calculated from the experimental fracture properties of the human cortical bone. Crack propagation analyses are carried out in two different 2D orthogonal sections cut from a three-dimensional 8 mm diameter cylindrical trabecular bone sample. The two sections differ in microstructural features such as area fraction (ratio of the 2D space occupied by bone tissue to the total 2D space), mean trabecula thickness, and connectivity. Analyses focus on understanding the effect of the rate of loading as well as on how the rate variation interacts with the microstructural features to cause anisotropy in microdamage accumulation and in the fracture resistance. Results are analyzed in terms of the dependence of fracture energy dissipation on the microstructural features as well as in terms of the changes in damage and stresses associated with the bone architecture variation. Besides the obvious dependence of the fracture behavior on the rate of loading, it is found that the microstructure strongly influences the fracture properties. The orthogonal section with lesser area fraction, low connectivity, and higher mean trabecula thickness is more resistant to fracture than the section with high area fraction, high connectivity, and lower mean trabecula thickness. In addition, it is found that the trabecular architecture leads to inhomogeneous distribution of damage, irrespective of the symmetry in the applied loading with the fracture of the entire bone section rapidly progressing to bone fragmentation once the accumulated damage in any trabeculae reaches a critical limit.     

Cross-linking connectivity in bone collagen fibrils: the COOH-terminal locus of free aldehyde.

Quantitative analyses of the chemical state of the 16c residue of the alpha 1 chain of bone collagen were performed on samples from fetal (4-6-month embryo) and mature (2-3 year old) bovine animals. All of this residue could be accounted for in terms of three chemical states, in relative amounts which depended upon the age of the animal. Most of the residue was incorporated into either bifunctional or trifunctional cross-links. Some of it, however, was present as free aldehyde, and the content increased with maturation. This was established by isolating and characterizing the aldehyde-containing peptides generated by tryptic digestion of NaB3H4-reduced mature bone collagen. We have concluded that the connectivity of COOH-terminal cross-linking in bone collagen fibrils changes with maturation in the following way: at first, each 16c residue in each of the two alpha 1 chains of the collagen molecule is incorporated into a sheet-like pattern of intermolecular iminium cross-links, which stabilizes the young, nonmineralized fibril as a whole. In time, some of these labile cross-links maturate into pyridinoline while others dissociate back to their precursor form. The latter is likely due to changes in the molecular packing brought about by the mineralization of the collagen fibrils. The resultant reduction in cross-linking connectivity may provide a mechanism for enhancing certain mechanical characteristics of the skeleton of a mature animal.