Quantitative and Computer Assisted Electron Microscopic and Microprobe Studies in Dermatology

Electron microscopes are not yet routine instruments in modern dermatological pathology even though they have provided detailed data about pathological changes in the skin for more than three decades. At present, dermatopathology is still dominated by light microscopy and especially so since the introduction of immunological techniques such as the use of monoclonal antibodies. These tools applied at electron microscopic resolution, however, may provide the ultimate identification of cells and subcellular components. In addition, electron microscopes have no peers in areas of quantitative investigation at subcellular levels, e.g. morphometry. The electron microprobe provides a unique tool in elemental analysis and may be used for the analysis of conventonally prepared specimens when foreign matter, not soluble in water, is deposited in the tissue. On the other hand, with water soluble substances the technique is most effective when freeze sections are utilized. This paper gives a selected review of the present day status of quantitative skin research as analysed with electron microscopy and the related technique of electron microprobe analysis

Proton Induced X-Ray Emission Analysis of Biological Specimens - Past and Future

Proton induced X-ray emission (PIXE) analysis is a comparatively new member of the family of spectrographic methods. In the last decade PIXE techniques have been applied to biological problems with great success. This review gives a condensed presentation of recent developments in biological (medical, zoological, and botanical) applications of PIXE analysis with special focus on factors which commonly influence the results, such as calibration, contamination, and preparation. The great advantage of PIXE analysis in studying physiologically important trace elements such as Zn, Mg, Fe, and Cu is underlined. Elemental mapping not only allows quantitative elemental analysis, but can also demonstrate the important differences in the morphological distributions of elements by comparing normal and pathological tissue

Hereditary Hair Changes Revealed by Analysis of Single Hair Fibres by Scanning Electron Microscopy

In many disorders with a genetic background the sparsity of scalp hairs may deter the clinician from trying to extract information from single hair fibres. Presenting a number of diverse conditions, we propose to show that simple measures can be taken in the doctor\u27s office which makes single fibre analysis a useful tool for assessment of factors involved in genetic disorders including the integument and its appendages. The paper is focussed on the utilization of the scanning electron microscope with the goal of demonstrating that pertinent information can be gained where information from transmission electron microscopy and other techniques are not immediately available

Disturbance of resting behaviour of broilers under different environmental conditions

Resting behaviours make up a large part of the daily time budget of broilers. However, in intensive broiler production systems disruptions of resting behaviours occur, where resting individuals get disturbed by active individuals. Such interruptions of resting behaviour may negatively affect the welfare of the birds but also disturb important functions of sleep related for instance to restoration and growth. This study investigated if a lower stocking density or the provision of separate resting places in the form of elevated platforms would result in less disruption of resting patterns of fast-growing broilers. Three different treatments were used, one with an elevated platform, one with lowered stocking density and a control setting. A randomized block design was used, consisting of six blocks, giving six to eight replicates of each treatment. Depending on treatment, 422-497 broilers (Ross 308) were kept per pen. Data on duration of resting bouts, occurrence of disturbances and position in pen during resting were collected on days 20 and 34 of age. An overall effect of treatment x position was found on duration of resting bouts both during day (P < 0.01) and night (P < 0.001). When resting on platforms the duration of resting bouts during daytime was longer compared to when birds were resting in open areas in the control groups (P = 0.04). During night the duration of resting bouts on platforms was longer compared to duration of resting bouts at all other locations in all treatments. In addition, resting position also had an overall effect on proportion of disturbances during day (P = 0.0018) and night (P = 0.0225). Resting on platforms reduced the number of physical disturbances of resting chickens compared to open areas in the control group both during day (P adj < 0.001) and night (P adj = 0.01). Generally, the level of disturbances was high in all treatments, suggesting that birds experience disrupted rest. As rest and sleep are vital needs, it is a welfare concern that chickens negatively affect other individuals' resting behaviour. A separate resting place appears to reduce disturbances to some extent and thus potentially increases the welfare of broiler chickens. However, provision of platforms is not enough to prevent frequent disturbances of resting and it is concluded that additional changes in housing conditions of broiler chickens are needed to improve their rest and sleep

Assessment of Qualitative and Quantitative Data from Pathological Hairs – A Critical Evaluation of Scanning Electron Microscope and Proton Induced X-Ray Emission Analyses

Analysis of single hair fibres in genetic disorders is a desirable complement to the clinical diagnosis. The scanning electron microscope (SEM) allows detailed study of the surface morphology of hair fibres which may explain some mechanical characteristics of the pathological hair. Quantitative elemental data may indicate biochemical or metabolic abnormalities. In this preliminary study we assess the feasibility of combining SEM and proton induced X-ray emission (PIXE) analysis on single hair fibres from 12 cases of genetic disease influencing the integument status. We conclude that SEM is a valuable tool in the analysis of hair pathology. The macro-PIXE technique involves some methodological and technical problems which in many cases are likely to be solved by using a proton microbeam. However, this means that routine methods have to be abandoned and careful selection of the material for analysis is an imperative necessity

Elemental Content of Anagen Hairs in a Normal Caucasian Population Studies with Proton Induced X-Ray Emission (PIXE)

The elemental content of anagen hair fibers in a Caucasian population of healthy females and males in the age range 10-69 years was performed to constitute a baseline for further studies of pathological conditions. Proton induced X-ray emission (PIXE) analyses were performed on single hair fibers in triplicates from 103 individuals in order to determine sulfur, zinc, calcium, and chlorine content. The hair fibers were all anagen hairs collected from a site little influenced by genetic and hormonal influences 1.5 cm above the right ear of the probands. An area 5-8 mm from the follicle bottom was chosen for minimize effect the of analysis in all cases hair-do contamination. The average sulfur content was 0.049 g/g and the average zinc content 170 μg/g. These results were not significantly influenced by chloroform/ethanol rinsing before analysis. The calcium and chlorine contents were 330 μg/g and 0.0033 g/g respectively. The latter data are expected to be more seriously influenced by external factors (e.g., contamination) than sulfur and zinc. No correlation between elemental concentration and sex was found for sulfur and zinc in the present material. PIXE analysis of single hair fibers yields valuable information on the elemental composition of hair fibers and can be rapidly and efficiently performed after simple mounting procedures

Resting behavior of broilers reared with or without artificial brooders

Rest and sleep are important for the welfare of mammals and birds. A large part of the daily time budget of broiler chickens is taken up by resting behavior and the quality of resting is important. However, in intensive broiler production systems, disruptions of resting behaviors are common. These disruptions of resting behavior could be negative for the health and growth of the birds. This study investigated if artificial brooders that provide a delimited and darker resting place, away from active birds, reduce disruptions of resting behavior compared to a control situation without artificial brooders. Six pens of each treatment were used in the same building, keeping 60 chickens (Ross 308) per pen. The artificial brooders were removed at 21 days of age. Data on disturbances and duration of resting bouts and activity between resting bouts were collected on 20 and 34 days of age. Also, as an indicator of the quality of rest, the animals' cognitive performance was evaluated in a spatial learning test that was performed at 11 days of age. The results showed that birds housed in pens with access to brooders have longer resting bouts (260.7 +/- 5.2 vs. 132.8 +/- 5.3s, p < 0.001) and are less likely to be disturbed during resting by other individuals (0.15 vs. 0.48, p < 0.001). The effect of the artificial brooders on both the duration of resting bouts and the proportion of disturbances remained after the removal of the brooders at 21 days of age. The duration of activity between resting bouts was shorter if the resting bout was ended by a disturbance (9.98 +/- 1.0 vs. 61.0 +/- 2.4s, p < 0.001). Birds reared with brooders were more likely to solve the spatial learning task (0.5 vs. 0.27, p < 0.01), but those succeeding were not faster at solving it. Broilers may be exposed to disrupted rest due to the lack of a dedicated resting place separated from areas with high activity. Using artificial brooders reduces disturbances but does not eliminate them. Therefore, additional changes to the housing conditions or management will be needed to prevent disturbances

Human Skin Physiology Studied by Particle Probe Microanalysis

Particle probe methods (electron probe and proton probe X-ray microanalysis) have been applied to investigate the distribution of elements and water over the different layers of the epidermis. For major elements, electron probe X-ray microanalysis (XRMA) provides the advantage of superior spatial resolution, but for trace element analysis the more sensitive proton probe (particle induced X-ray emission, PIXE) analysis has to be used. On a dry weight basis, the concentration of S is rather constant across the epidermis, whereas the concentrations of P, K, Cl and Na show gradients with high levels in stratum germinativum (basale) and stratum spinosum but low levels in the stratum granulosum and stratum corneum. Essentially, Fe and Zn are confined to the basal region in normal skin. The concentration of Ca, however, increased steadily from the basal region to the stratum corneum. The probe technique allows quantitative analysis of stratum-specific changes in elemental content in a variety of pathological conditions, e.g., changes induced by nickel, detergents and other chemicals, or in psoriatic skin. Of particular interest are findings of increased Fe and Zn in non-involved psoriatic skin. Since the different layers of the skin have different elemental concentrations and react differently under pathological conditions, the probe techniques are far superior to bulk chemical analysis in elucidating physiological and pathological processes in the skin

Resting behavior of broilers reared with or without artificial brooders

Rest and sleep are important for the welfare of mammals and birds. A large part of the daily time budget of broiler chickens is taken up by resting behavior and the quality of resting is important. However, in intensive broiler production systems, disruptions of resting behaviors are common. These disruptions of resting behavior could be negative for the health and growth of the birds. This study investigated if artificial brooders that provide a delimited and darker resting place, away from active birds, reduce disruptions of resting behavior compared to a control situation without artificial brooders. Six pens of each treatment were used in the same building, keeping 60 chickens (Ross 308) per pen. The artificial brooders were removed at 21 days of age. Data on disturbances and duration of resting bouts and activity between resting bouts were collected on 20 and 34 days of age. Also, as an indicator of the quality of rest, the animals' cognitive performance was evaluated in a spatial learning test that was performed at 11 days of age. The results showed that birds housed in pens with access to brooders have longer resting bouts (260.7 ± 5.2 vs. 132.8 ± 5.3s, p &lt; 0.001) and are less likely to be disturbed during resting by other individuals (0.15 vs. 0.48, p &lt; 0.001). The effect of the artificial brooders on both the duration of resting bouts and the proportion of disturbances remained after the removal of the brooders at 21 days of age. The duration of activity between resting bouts was shorter if the resting bout was ended by a disturbance (9.98 ± 1.0 vs. 61.0 ± 2.4s, p &lt; 0.001). Birds reared with brooders were more likely to solve the spatial learning task (0.5 vs. 0.27, p &lt; 0.01), but those succeeding were not faster at solving it. Broilers may be exposed to disrupted rest due to the lack of a dedicated resting place separated from areas with high activity. Using artificial brooders reduces disturbances but does not eliminate them. Therefore, additional changes to the housing conditions or management will be needed to prevent disturbances

Antibodies to citrullinated proteins and differences in clinical progression of rheumatoid arthritis

Antibodies to citrullinated proteins (anti-cyclic-citrullinated peptide [anti-CCP] antibodies) are highly specific for rheumatoid arthritis (RA) and precede the onset of disease symptoms, indicating a pathogenetic role for these antibodies in RA. We recently showed that distinct genetic risk factors are associated with either anti-CCP-positive disease or anti-CCP-negative disease. These data are important as they indicate that distinct pathogenic mechanisms are underlying anti-CCP-positive disease or anti-CCP-negative disease. Likewise, these observations raise the question of whether anti-CCP-positive RA and anti-CCP-negative RA are clinically different disease entities. We therefore investigated whether RA patients with anti-CCP antibodies have a different clinical presentation and disease course compared with patients without these autoantibodies. In a cohort of 454 incident patients with RA, 228 patients were anti-CCP-positive and 226 patients were anti-CCP-negative. The early symptoms, tender and swollen joint count, and C-reactive protein level at inclusion, as well as the swollen joint count and radiological destruction during 4 years of follow-up, were compared for the two groups. There were no differences in morning stiffness, type, location and distribution of early symptoms, patients' rated disease activity and C-reactive protein at inclusion between RA patients with and without anti-CCP antibodies. The mean tender and swollen joint count for the different joints at inclusion was similar. At follow-up, patients with anti-CCP antibodies had more swollen joints and more severe radiological destruction. Nevertheless, the distribution of affected joints, for swelling, bone erosions and joint space narrowing, was similar. In conclusion, the phenotype of RA patients with or without anti-CCP antibodies is similar with respect to clinical presentation but differs with respect to disease course