Margaret Tzaphlidou

The collagen fibril—A model system for studying the staining and fixation of a protein

A collagen fibril is made up of long rod-like molecules regularly D-staggered with respect to one another. This means that (i) its axially projected fine structure, resolvable to approximately 2 nm in electron micrographs, repeats D-periodically (D = 67 nm), and (ii) the amino acid residues contributing to each element of the fine structure can be inferred from sequence data. Electron-optical data from a fibril D-period can can therefore be correlated directly with chemical data. Such correlations confirm the electrostatic nature of the staining reaction when a fibril is positively stained. After negative staining, the principal factor determining the small-scale distribution of stain is local exclusion by bulky amino acid side-chains. (Bulkiness is the average cross-sectional area, or plumpness, of a side-chain.) A small superimposed positive staining contribution can also be detected. Fixation of collagen by aldehydes and diimidoesters occurs via an initial reaction with lysyl (and hydroxylsyl) side-chains and alpha-amino groups, followed by secondary cross-linking reactions that differ from fixative to fixative. These secondary reactions determine the nature and abundance of the cross-links and the extent to which they influence subsequent staining behaviour.

Structural variations of collagen in normal and pathological tissues: role of electron microscopy.

The spectrum of ultrastructural appearances assumed by collagen in normal and pathological tissues is illustrated using techniques of thin section transmission electron microscopy and computer-assisted analysis. The normal fibrillar collagen types are described in order to provide a basis for comparing other normal and abnormal forms. In normal tissues, the anchoring fibril and basal lamina (basement membrane) represent tissue structures largely containing collagen but differing significantly in organisation from normal types I to III fibrillar collagen. In pathological tissue, deviations from normal fine structure are reflected in abnormal aggregates of collagen fibrils (amianthoid and skeinoid fibres) and abnormalities in fibril diameter and cross-sectional profile. Fibrous and segment long-spacing collagen represent two further organisational variants of collagen, the former found widely in pathological tissues, the latter very rarely. Much remains to be discovered about these abnormal collagen variants-their mode of formation, the cells that produce them, and their roles. They also present a challenge for the collagen biologist formulating hypotheses of collagen fibril assembly and molecular organisation.

Automated Epileptic Seizure Detection Methods: A Review Study

Epilepsy is a neurological disorder with prevalence of about 1-2% of the world’s population (Mormann, Andrzejak, Elger & Lehnertz, 2007). It is characterized by sudden recurrent and transient disturbances of perception or behaviour resulting from excessive synchronization of cortical neuronal networks; it is a neurological condition in which an individual experiences chronic abnormal bursts of electrical discharges in the brain. The hallmark of epilepsy is recurrent seizures termed epileptic seizures. Epileptic seizures are divided by their clinical manifestation into partial or focal, generalized, unilateral and unclassified seizures (James, 1997; Tzallas, Tsipouras & Fotiadis, 2007a, 2009). Focal epileptic seizures involve only part of cerebral hemisphere and produce symptoms in corresponding parts of the body or in some related mental functions. Generalized epileptic seizures involve the entire brain and produce bilateral motor symptoms usually with loss of consciousness. Both types of epileptic seizures can occur at all ages. Generalized epileptic seizures can be subdivided into absence (petit mal) and tonic-clonic (grand mal) seizures (James, 1997).

Bone Architecture: Collagen Structure and Calcium/Phosphorus Maps

Bone collagen structure in normal and pathological tissues is presented using techniques of thin section transmission electron microscopy and morphometry. In pathological tissue, deviations from normal fine structure are reflected in abnormal arrangements of collagen fibrils and abnormalities in fibril diameter. The relationships between these bone structural changes and the skeletal calcium/phosphorus ratio are discussed. Calcium/phosphorus ratio is measured by X-ray absorptiometry and computed microtomography.

Determination of calcium, phosphorus, and the calcium/phosphorus ratio in cortical bone from the human femoral neck by neutron activation analysis.

Concentrations of Ca and P as well as the Ca/P ratio were estimated in intact cortical bone samples from the femoral neck of healthy humans, 33 women and 45 men, aged from 15 to 55 yr using instrumental neutron activation analysis. Mean values (M +/- SD) for the investigated parameters (on dry weight basis) were: 23.0 +/- 3.9%, 10.7 +/- 2.4% and 2.17 +/- 0.31, respectively. No statistically significant differences of the above parameters were observed related either to age or sex. The mean values for Ca, P and Ca/P ratio were within a very wide range of published data and close to their median. The individual variation for the Ca/P ratio in cortical bone from the healthy human femoral neck was lower than those for Ca and P separately. This means that specificity of Ca/P ratio is better than those of Ca and P concentrations are and may be more reliable for diagnosis of bone disorders.

In vivo measurement of radius calcium/phosphorus ratio by X-ray absorptiometry.

We describe a new method for assessing the skeletal Ca/P ratio in vivo using X-ray absorptiometry. By placing cerium (Ce) and samarium (Sm) filters in the X-ray beam from a commercial X-ray source (Norland), mean photon energies of 39 and 89 keV were obtained. The instrument was designed to take measurements of the forearm, at a site located at the distal 1/3 of the radius. The system was calibrated with three bone phantoms: Ca10(PO4)6(OH)2, Ca(HPO4)(H2O)2 and Ca(HPO4)2(H2O)). The precision for measuring the Ca/P ratio in the human radius was 2.3% CV for a skin dose to the forearm ranging from 0.3 to 0.4 mGy, depending on the width of the arm. The Ca/P ratio of the radius was significantly lower in patients with postmenopausal osteoporosis than in premenopausal controls.

Calcium and phosphorus concentrations and the calcium/phosphorus ratio in trabecular bone from the femoral neck of healthy humans as determined by neutron activation analysis.

The Ca and P concentrations as well as the Ca/P ratio were estimated in intact trabecular bone samples from the femoral neck of healthy humans, 34 women and 44 men, aged from 15 to 55 years, using instrumental neutron activation analysis. The mean values (M+/-SD) for the investigated parameters (on a dry-weight basis) were: 12.1+/-3.0%, 5.94+/-1.71%, 2.07+/-0.25 and 10.9+/-2.5%, 5.30+/-1.23%, 2.07+/-0.22 for females and males, respectively. A statistically significant (p</=0.05) decrease of Ca concentration with age was found only for males while the P concentration and the Ca/P ratio were not affected by age. No statistically significant sex-related differences were observed in any of the parameters. The mean values for Ca, P and the Ca/P ratio lay close to the median of the very wide range of published data. The individual variation for the Ca/P ratio in trabecular bone from the healthy human femoral neck was lower than those for Ca and P separately. This suggests that the specificity of the Ca/P ratio is better than that of Ca and P concentrations and may be more reliable for the diagnosis of bone disorders.

Calcium, phosphorus, calcium-phosphorus ratio in rib bone of healthy humans.

Calcium and phosphorus concentrations as well as the Ca/P ratio were estimated in intact rib bone samples from healthy humans, 37 women and 45 men, aged 15–55 yr. For Ca and P concentration measurements, instrumental neutron activation analysis was used. The mean values (mean±SD) for the investigated parameters were 19.3±4.5% of dry bone weight, 8.42±2.14% of dry bone weight, and a ratio of 2.33±0.34, respectively. Statistically significant differences for the above parameters were not observed to be related either to age or sex. The mean values for Ca, P, and the Ca/P ratio were within a very wide range of published data and close to their medians. The individual variation for the Ca/P ratio in rib bone from healthy humans was lower than those for Ca and P taken separately. An indication is that the specificity of the Ca/P ratio improves upon that for Ca and P concentrations and may be more reliable in the diagnosis of bone disorders.

Structural destabilization of chignolin under the influence of oscillating electric fields

The structural response of chignolin to 1 V/nm electric fields of different frequencies has been studied with molecular dynamics simulations and stochastic modeling. It was found that oscillating electric fields induce conformational changes to chignolin that are frequency dependent. For frequencies comparable with or smaller from the orientational self-diffusion rate, the peptide destabilizes after performing an oscillatory motion between the two possible directions of the electric field axis. For higher frequencies the field effects are averaged out and chignolin performs a Brownian rotation diffusion maintaining its native conformation. Stochastic modeling can describe chignolin’s oscillatory motion equally well with the molecular dynamics simulations. The time needed for these changes to take place has a stochastic nature depending, beyond frequency, on factors related with the hydrogen bonds’ stability and their geometrical arrangement in the structure.

The effects of inflammation-mediated osteoporosis (IMO) on the skeletal Ca/P ratio and on the structure of rabbit bone and skin collegen

The relationships between the skeletal Ca/P ratio (used as an index of bone quality) and structural changes in the bone and skin was studied in inflammatory mediated osteoporosis (IMO). The bone Ca/P ratio in IMO rabbits was significantly lower than in controls. Also, severe alterations were detected at the ultrastructural level in bone and skin collagen fibrils from IMO rabbits.