Methods of studying the planar distribution of objects in histological sections of brain tissue

This article reviews the statistical methods that have been used to study the planar distribution, and especially clustering, of objects in histological sections of brain tissue. The objective of these studies is usually quantitative description, comparison between patients or correlation between histological features. Objects of interest such as neurones, glial cells, blood vessels or pathological features such as protein deposits appear as sectional profiles in a two-dimensional section. These objects may not be randomly distributed within the section but exhibit a spatial pattern, a departure from randomness either towards regularity or clustering. The methods described include simple tests of whether the planar distribution of a histological feature departs significantly from randomness using randomized points, lines or sample fields and more complex methods that employ grids or transects of contiguous fields, and which can detect the intensity of aggregation and the sizes, distribution and spacing of clusters. The usefulness of these methods in understanding the pathogenesis of neurodegenerative diseases such as Alzheimer's disease and Creutzfeldt-Jakob disease is discussed. © 2006 The Royal Microscopical Society.

Quantification of the pathological changes in the temporal lobe of patients with a novel neurofilamentopathy: neurofilament inclusion disease (NID)

Objective: To quantify the densities of neurofilament inclusions (NI), swollen achromatic neurons, surviving neurons and glial cells in a novel neurofilamentopathy neurofilament inclusion disease (NID). Material: Sectionsof temporal lobe from 4 cases of NID stained with an antibody raised to neurofilament proteins. Method: Densities of the pathological changes were estimated in the various gyri of the temporal lobe, hippocampus and dentate gyrus. Results: Densities of the NI and swollen achromatic neurons (SN) were greater in the cerebral cortical gyri than in the hippocampus and dentate gyrus. Lesion density was relatively constant between gyri and between the CA sectors of the hippocampus. In cortical gyri, the density of the NI, SN and glial cell nuclei was greater in laminae II/III than laminae V/VI. Densities of the NI were negatively correlated with the surviving neurons and positively correlated with the glial cell nuclei. The density of the SN was positively correlated with that of the surviving neurons. Conclusion: The pathology of NID morphologically resembles that of Pick's disease (PD) and corticobasal degeneration (CBD), but there are distinct differences between NID and these disorders supporting the hypothesis that NID is a novel and unique type of neurodegenerative disease.

Clustering of neuronal inclusions in "dementia with neurofilament inclusions"

Dementia with neurofilament inclusions (DNI) is a new disorder characterized clinically by early-onset dementia and histologically by the presence of intraneural inclusions immunopositive for neurofilament antigens but lacking tau and α-synuclein reactivity. We studied the clustering patterns of the neurofilament inclusions (NI) in regions of the temporal lobe in three cases of DNI to determine whether they have the same spatial patterns as inclusions in the tauopathies and α-synucleinopathies. The NI exhibited a clustered distribution (mean size of clusters 400 μm, range 50-800 μm, SD 687.8) in 24/28 of the areas studied. In 22 of these areas, the clusters exhibited a regular distribution along the tissue parallel to the pia mater or alveus. In 3 cortical areas, there was evidence of a more complex pattern in which the NI clusters were aggregated into larger superclusters. In 6 cortical areas, the size of the clusters approximated to those of the cells of origin of the cortico-cortical pathways but in the remaining areas cluster size was smaller than 400 μm. Despite the unique molecular profile of the NI, their spatial patterns are similar to those shown by filamentous neuronal inclusions in the tauopathies and α-synucleinopathies.

An analysis of returned medicines in primary care

Objective: The number of pharmaceutical items issued on prescription is continually rising and contributing to spiralling healthcare costs. Although there is some data highlighting the quantity, in terms of weight of medicines returned specifically to community pharmacies, little is known about the specific details of such returns or other destinations for wasted medications. This pilot study has been designed to investigate the types and amounts of medicines returned to both general practices (GPs) and associated local community pharmacies determining the reasons why these medicines have been returned. Method: The study was conducted in eight community pharmacies and five GP surgeries within East Birmingham over a 4-week period. Main outcome Measure: Reason for return and details of returned medication. Results: A total of 114 returns were made during the study: 24 (21.1) to GP surgeries and 90 (78.9) to community pharmacies. The total returns comprised 340 items, of which 42 (12.4) were returned to GPs and 298 (87.6) to pharmacies, with the mean number of items per return being 1.8 and 3.3, respectively. Half of the returns in the study were attributed to the doctor changing or stopping the medicine; 23.7 of returns were recorded as excess supplies or clearout often associated with patients' death and 3.5 of returns were related to adverse drug reactions. Cardiovascular drugs were most commonly returned, amounting to 28.5 of the total drugs returned during the study. Conclusions: The results from this pilot study indicate that unused medicines impose a significant financial burden on the National Health Service as well as a social burden on the United Kingdom population. Further studies are examining the precise nature of returned medicines and possible solutions to these issues. © Springer 2005.

Influence of check and field size on the visual evoked magnetic response to a pattern shift stimulus

A decrease in the check size of a pattern shift stimulus increases the latency and amplitude of the visual evoked potential (VEP) P100. In addition, for a given check size, decreasing the size of the stimulus field increases the latency and amplitude of the P100. These results imply that the central regions of the retina make a significant contribution to the generation of the electrical P100. However, the corresponding magnetic P100m may have a different origin. We have studied the effects of check and field size on the P100m in five normal subjects using a DC-Squid, second-order gradiometer. Magnetic responses were recorded at the positive maximum of the P100m over the occipital scalp to six check sizes (10-100') presented in a large (13 degrees 34') and small (5 degrees 14') field and to a large check (100') presented in seven field sizes (1 degree 45' - 15 degrees 10'). No responses were recorded to any check size with a small field. Decreasing the check size presented in a large field increased latency of the P100m by approx. 30 ms while the amplitude of the response decreased with the largest reduction occurring between 70' and 12' checks. Using a large check, latency increased and amplitude decreased as the field size was reduced. The latency changes in response to check and field size were similar to those described for the VEP although the magnitudes of the magnetic changes were greater. Unlike the VEP, amplitude responses were maximal when large checks were presented in a large stimulus field. This suggests that regions outside the central retina make a more significant contribution to the visual evoked magnetic response than they do to the VEP, and that the P100m may be useful clinically in the study of diseases that affect the more peripheral regions of the retina.

Analysis of spatial patterns in histological sections of brain tissue using a method based on regression

A method of determining the spatial pattern of any histological feature in sections of brain tissue which can be measured quantitatively is described and compared with a previously described method. A measurement of a histological feature such as density, area, amount or load is obtained for a series of contiguous sample fields. The regression coefficient (β) is calculated from the measurements taken in pairs, first in pairs of adjacent samples and then in pairs of samples taken at increasing degrees of separation between them, i.e. separated by 2, 3, 4,..., n units. A plot of β versus the degree of separation between the pairs of sample fields reveals whether the histological feature is distributed randomly, uniformly or in clusters. If the feature is clustered, the analysis determines whether the clusters are randomly or regularly distributed, the mean size of the clusters and the spacing of the clusters. The method is simple to apply and interpret and is illustrated using simulated data and studies of the spatial patterns of blood vessels in the cerebral cortex of normal brain, the degree of vacuolation of the cortex in patients with Creutzfeldt-Jacob disease (CJD) and the characteristic lesions present in Alzheimer's disease (AD). Copyright (C) 2000 Elsevier Science B.V.

Analysis of spatial patterns in histological sections of brain tissue

A method is described which enables the spatial pattern of discrete objects in histological sections of brain tissue to be determined. The method can be applied to cell bodies, sections of blood vessels or the characteristic lesions which develop in the brain of patients with neurodegenerative disorders. The density of the histological feature under study is measured in a series of contiguous sample fields arranged in a grid or transect. Data from adjacent sample fields are added together to provide density data for larger field sizes. A plot of the variance/mean ratio (V/M) of the data versus field size reveals whether the objects are distributed randomly, uniformly or in clusters. If the objects are clustered, the analysis determines whether the clusters are randomly or regularly distributed and the mean size of the clusters. In addition, if two different histological features are clustered, the analysis can determine whether their clusters are in phase, out of phase or unrelated to each other. To illustrate the method, the spatial patterns of senile plaques and neurofibrillary tangles were studied in histological sections of brain tissue from patients with Alzheimer's disease.

Analysis of population dispersion in histological sections

Stereology and other image analysis methods have enabled rapid and objective quantitative measurements to be made on histological sections. These mesurements may include total volumes, surfaces, lengths and numbers of cells and blood vessels or pathological lesions. Histological features, however, may not be randomly distributed across a section but exhibit 'dispersion', a departure from randomness either towards regularity or aggregation. Information of population dispersion may be valuable not only in understanding the two-or three-dimensional structure but also in elucidating the pathogenesis of lesions in pathological conditions. This article reviews some of the statistical methods available for studying dispersion. These range from simple tests of whether the distribution of a histological faeture departs significantly from random to more complex methods which can detect the intensity of aggregation and the sizes, distribution and spacing of the clusters.

Dispersion of classic beta-amyloid deposits around blood vessels in Alzheimer’s disease

The spatial pattern of the classic (‘cored’) type of beta-amyloid (Abeta) deposit was studied in the upper laminae of the superior temporal gyrus in 9 cases of sporadic Alzheimer’s disease (SAD). Abeta stained tissue was counterstained with collagen IV to study the relationships between the spatial distribution of the classic deposits and the blood vessel profiles. Both the classic deposits and blood vessel profiles were distributed in clusters. In all cases, there was a spatial correlation between the clusters of the classic deposits and the larger diameter (>10 micron) blood vessel profiles and especially the vertically penetrating arterioles. In only 1 case, was there a significant spatial correlation between the clusters of the classic deposits and the smaller diameter (<10 micron) capillaries. In 9/11 cases, the clusters of the classic deposits were significantly larger than those of the clusters of the larger blood vessels. In addition, the density of the classic deposits declined as a negative exponential function with distance from the vertically penetrating arterioles. These results suggest that the classic Abeta deposits cluster around the larger blood vessels in the frontal cortex and that diffusion of proteins from these blood vessels could be involved in the pathogenesis of the classic deposits in SAD.

Clustering patterns of neurofibrillary tangles in Alzheimer’s disease

Clustering of cellular neurofibrillary tangles (NFT) was studied in the cerebral cortex and hippocampus in cases of Alzheimer’s disease (AD) using a regression method. The objective of the study was to test the hypothesis that clustering of NFTs reflects the degeneration of the cortico-cortical pathways. In 25/38 (66%) of analyses of individual brain areas, a significant peak to trough and peak to peak distance was obtained suggesting that the clusters of NFTs were regularly distributed in bands parallel to the tissue boundary. In analyses of cortical tissues with regularly distributed clusters, peak to peak distance was between 1000 and 1600 microns in 13/24 (54%) of analyses, >1600 microns in 10/24 (42%) and <1000 microns in 1/24 (4%) of analyses. A regular distribution of NFT clusters was less evident in the CA sectors of the hippocampus than in the cortex. Hence, in a significant proportion of brain areas, the spacing of NFT clusters along the cerebral cortex was consistent with the predicted distribution of the cells of origin of specific cortico-cortical projections. However, in many brain regions, the sizes of the NFT clusters were larger than predicted which may be attributable to the spread of NFTs to adjacent groups of cells as the disease progresses.

Degeneration of cortical neurons associated with diffuse beta-amyloid (Abeta) deposits in Alzheimer’s disease

The frequency of morphological abnormalities in neuronal perikarya which were in contact with diffuse beta-amyloid (Abeta) deposits in patients with Alzheimer’s disease (AD) was compared with neurons located adjacent to the deposits. Morphological abnormalities were also studied in elderly, non-demented (ND) cases with and without diffuse Abeta deposits. In AD and ND cases with Abeta deposits, an increased proportion of neurons in contact with diffuse deposits exhibited at least one abnormality compared with neurons located adjacent to the deposits. Neurons in contact with diffuse deposits exhibited a greater frequency of abnormalities of shape, nuclei, nissl substance and had a higher frequency of cytoplasmic vacuoles compared with adjacent neurons. A greater frequency of abnormalities of shape, nissl substance and in the frequency of displaced nuclei were also observed in neurons adjacent to diffuse deposits in AD compared with ND cases. With the exception of absent nuclei, morphological abnormalities adjacent to diffuse deposits in ND cases were similar to those of ND cases without Abeta deposits. These results suggest that neuronal degeneration is associated with the earliest stages of Abeta deposit formation and is not specifically related to the formation of mature senile plaques.

Dispersal of soredia from individual soralia of the lichen Hypogymnia physodes (L.) Nyl. in a simple wind tunnel

Dispersal of soredia from individual soralia of the lichen Hypogymnia physodes (L.) Nyl. was studied using a simple wind tunnel constructed in the field. Individual lobes with terminal soralia were placed in the wind tunnel on the adhesive surface of dust particle collectors. Air currents produced by a fan were directed over the surface of the lobes. The majority of soredia were deposited within 5 cm of the source soralium but some soredia were dispersed to at least 80 cm at a wind speed of 6 m s-1. Variation in wind speed had no statistically significant effect on the total number of soredial clusters deposited averaged over soralia but the mean size of cluster and the distance dispersed were greater at higher wind speeds. The number of soredia deposited was dependent on the orientation of the soralium to the air currents. More soredia were deposited with the soralium facing the fan at a wind speed of 9 m s-1. Moisture in the form of a fine mist reduced substantially the number of soredia deposited at a wind speed of 6 m s-1 but had no effect on the mean number of soredia per cluster or on the mean distance dispersed. The data suggest: (1) that wind dispersal from an individual soralium is influenced by wind speed, the location of the soralium on the thallus and the level of moisture and (2) that air currents directed over the surfaces of thalli located on the upper branches of trees would effectively disperse soredia of H. physodes vertically and horizontally within a tree canopy. © 1994.

Effect of osmotic and matrix potential on Saprolegnia diclina and S. ferax

Growth of biomass and sporulation of pathogenic and non-pathogenic Saprolegnia species was markedly decreased at reduced water potentials. Oogonium and zoosporangium formation were more sensitive to reduced osmotic and matrix potentials than growth in biomass. Although little difference was observed between the effects of matrix and osmotic potentials, the Saprolegnia species investigated responded differently to those solutes utilized in control of osmotic potential. Biomas, oogonium and zoosporangium formation were greater in the presence of reduced osmotic potentials mediated by mannitol than equivalent potentials mediated by potassium chloride. Endogenous potassium levels varied little with reduced matrix or osmotic potentials. Conversly, mannitol content of colonies exposed to reduced osmotic potentials mediated by mannitol initailly increased while endogenous amino acid levels were observed to rise in response to moderately reduced water potentials. Sensitivity of Saprolegnia species to reduced potantials and effects on substrate colonization are discussed in the light of these observations.

DNA Mutation of the progranulin (GRN) gene in familial frontotemporal lobar degeneration (FTLD):a study of the pathology of nine cases

Frontotemporal lobar degeneration (FTLD) with transactive response (TAR) DNA-binding protein of 43kDa (TDP-43) proteinopathy (FTLD-TDP) is a neurodegenerative disease characterized by variable neocortical and allocortical atrophy principally affecting the frontal and temporal lobes. Histologically, there is neuronal loss, microvacuolation in the superficial cortical laminae, and a reactive astrocytosis. A variety of TDP-43 immunoreactive changes are present in FTLD-TDP including neuronal cytoplasmic inclusions (NCI), neuronal intranuclear inclusions (NII), dystrophic neurites (DN) and, oligodendroglial inclusions (GI). Many cases of familial FTLD-TDP are caused by DNA mutations of the progranulin (GRN) gene. Hence, the density, spatial patterns, and laminar distribution of the pathological changes were studied in nine cases of FLTD-TDP with GRN mutation. The densities of NCI and DN were greater in cases caused by GRN mutation compared with sporadic cases. In cortical regions, the commonest spatial pattern exhibited by the TDP-43 immunoreactive lesions was the presence of clusters of inclusions regularly distributed parallel to the pia mater. In approximately 50% of cortical gyri, the NCI exhibited a peak of density in the upper cortical laminae while the GI were commonly distributed across all laminae. The distribution of the NII and DN was variable, the most common pattern being a peak of NII density in the lower cortical laminae and DN in the upper cortical laminae. These results suggest in FTLD-TDP caused by GRN mutation: 1) there are greater densities of NCI and DN than in sporadic cases of the disease, 2) there is degeneration of the cortico-cortical and cortico-hippocampal pathways, and 3) cortical degeneration occurs across the cortical laminae, the various TDP-43 immunoreactive inclusions often being distributed in different cortical laminae.

Genetics of ocular disease Part 1: Basic concepts: cells, chromosomes and inheritance

This series of articles describes the basic elements of genetics necessary to understand the new advances and the impact these advances will have on the study and treatment of ocular disease. The first article describes the patterns of inheritance of human characteristics, how they are transmitted between the generations and the structure of chromosomes.