Biomedical subjects

J Pallon

Publications and source records attributed to J Pallon.

13 recordsLinked to original sources

PubMed1999

Identification and characterization of a Streptococcus pyogenes ABC transporter with multiple specificity for metal cations.

Metal ions are crucial trace elements for bacteria infecting the human host. The LraI (lipoprotein receptor-associated antigen I) transporter in Streptococcus spp. belongs to the superfamily of ABC transporters. The transporter consists of a lipoprotein, an ATP-binding protein and a hydrophobic integral membrane protein. Here, we describe a new member of the LraI family in the important human pathogen Streptococcus pyogenes. The system was identified in silico by analysis of the S. pyogenes Genome Sequencing Project. The S. pyogenes operon exhibits an atypical organization compared with equivalents in other Streptococcus spp. The presence and atypical organization of the operon was verified in a number of S. pyogenes strains of different serotypes. Transcriptional analysis of the LraI operon demonstrates a polycistronic transcription attenuated by a stable stem-loop structure, which allows the lipoprotein to be expressed in larger quantities than the other two components. The localization of the native lipoprotein at the bacterial surface was shown by proteolytic digestion of S. pyogenes bacteria and NH2-terminal sequencing of a released lipoprotein fragment. Recombinant lipoprotein was expressed as a GST fusion protein, and studies of molecular interactions with metal radioisotopes demonstrated that the protein has affinity for Zn(II), Fe(III) and Cu(II). Zn(II) and Cu(II) were found to compete for the same binding site, whereas Fe(III) uses a second site. Also, proton-induced X-ray analysis of lipoprotein samples identified iron, copper and zinc. Finally, a mutant strain lacking a functional mtsABC operon was generated and showed reduced uptake of 55Fe and 65Zn compared with the wild-type strain. The operon encoding this novel ABC transporter with multiple specificity for metal cations is designated mtsABC, for metal transporter of Streptococcus.

ATP-Binding Cassette Transporters↗

PubMed1999

Elemental analysis mirrors epidermal differentiation.

Using a scanning nuclear microprobe, the distribution of elements and trace elements of skin cross sections of normal skin, non-lesional psoriatic skin and in dry atopic skin have been mapped. In non-lesional psoriatic skin and in dry atopic skin the epidermal Ca-gradient is higher than that of normal skin. In addition, abnormally high Fe and Zn levels were recorded in the stratum granulosum and corneum regions in the pathological skin. It is suggested that these findings correlate to an increased cell turnover in the basal cell layer of the psoriatic and atopic skins. The ratio of Ca/Zn in stratum corneum of paralesional psoriatic skin is approximately 8:1 compared to 12: 1 in normal skin and 15: 1 in atopic skin. This suggests that the differentiation process in paralesional psoriatic skin may actually be an example of disturbed programmed cell death.

Adult↗

PubMed1998

Normalization of epidermal calcium distribution profile in reconstructed human epidermis is related to improvement of terminal differentiation and stratum corneum barrier formation.

Calcium plays an important role in the regulation of cellular differentiation and desquamation of epidermal keratinocytes. In this study, we examined the calcium distribution in reconstructed epidermis in an attempt to understand the physiology of keratinocyte differentiation and desquamation in vitro. Ion capture cytochemistry (the potassium oxalate-pyroantimonate method) was employed to localize ionic calcium in reconstructed epidermis generated under three different culture conditions (in serum-containing medium, serum-free medium, and serum-free medium supplemented with retinoic acid), allowing a comparison of the physiology of incompletely and well-differentiated keratinocytes. The reconstructed epidermis generated in serum-containing medium showed features of incomplete differentiation, and compared with the native skin, a high calcium content within incompletely differentiated cells in the stratum corneum. Use of serum-free medium containing vitamin and lipid supplements led to a marked improvement of the stratum corneum ultrastructure and penetration pathway across the stratum corneum, indicating improved barrier formation of the reconstructed epidermis. In parallel, the calcium distribution pattern was normalized showing the highest levels of calcium in the stratum granulosum and low levels in the inner stratum corneum. Addition of retinoic acid to the serum-free medium resulted in an altered keratinocyte differentiation and re-appearance of large quantities of calcium precipitates in the stratum corneum. Proton probe X-ray microanalysis was applied to investigate the calcium distribution quantitatively in native and reconstructed epidermis generated in serum-free medium, and verified the calcium distribution demonstrated by the precipitation technique. Regardless of the presence or absence of calcium in the stratum corneum, all examined culture systems exhibited insufficient desquamation, which correlates with the finding that stratum corneum chymotryptic enzyme was present predominantly as an inactive precursor. This study demonstrates that improvement of the stratum corneum barrier properties in vitro is concurrent with the normalization of the epidermal calcium gradient, whereas deregulation of terminal differentiation correlates with an accumulation of calcium ions within incompletely differentiated corneocytes.

Adult↗

PubMed1997

Aspects on the physiology of human skin: studies using particle probe analysis.

The cellular part of the skin, the epidermis, is a very thin structure, approximately 120 microns thick, a fact which has hindered the exploration of the physiology of the epidermis in normal and pathological conditions. An additional complication is the fact that the epidermis contains layers of cells at different stages of differentiation. Therefore, conventional physiological capillary probes cannot, with any satisfactory precision, be located within a specified cell of a specified layer of the skin in vivo. Hence, alternative ways for the exploration of skin physiology have been sought for. In the past, analysis of the elemental content of skin was done was done as bulk measurements, and surprisingly wide ranges of elemental content were recorded. The width of these ranges was most certainly due to the sampling methods used rather than the sensitivity of the chosen method of analysis. Also, these older measurements did not discriminate between the different strata, and therefore the information provided little if any substance for a functional analysis of processes involved in normal and pathological differentiation of the epidermis. Particle probes, however, have been able to overcome such methodological problems. Over a period of 15 years we have studied normal human skin, normal-looking, paralesional skin from psoriatics, and skin from persons suffering from atopic dermatitis using PIXE analysis. In recent years, trace elements have been shown to work as secondary messengers or regulatory substances. As an example, calcium (Ca2+) has proven to be a very important signalling substance in a great variety of cellular systems. Studies with the transmission electron microscope (TEM) as well as histochemical methods have allowed an understanding of the role of Ca2+ in the differentiation process of the epidermis. Ca2+ has also been shown to play an important role in apoptosis (programmed cell death), which is currently a hot subject for the obvious reason that the final differentiation step between the stratum granulosum level and the stratum corneum represents a particular aspect of programmed cell death. The importance of the balance between calcium and zinc in apoptosis has been clearly demonstrated in a number of cellular systems, but we have still to clarify the validity of topical treatment with Zn ointments in different skin conditions. Substantial iron (Fe) losses via psoriatic lesions were demonstrated more than two decades ago, and these data were given new meaning when we found that a more discrete loss occurs in clinically normal-looking psoriatic skin. Obviously, such findings stress the importance of understanding the relation between the elemental content and normal and abnormal physiology. The ultimate goal of particle probe studies is to provide an understanding of the formation of a mature stratum corneum with a functional barrier reflected in physiological/biochemical mechanisms behind the properties of changed skin in patients afflicted with skin disorders of genetic or constitutional origin. This paper aims to give an overview of the state of the art in skin physiology made possible through the use of particle probes.

Animals↗

PubMed1996

Nuclear microprobe applied to the study of coronary artery walls--a distinct look at atherogenesis.

The Nuclear Microprobe technique was employed to investigate the elemental concentration alterations due to atherosclerosis, of minor and trace elements at the different cellular layers and structures of freeze-dried cryosections of human coronary arteries. Nuclear Microprobe analyses enable to determine 7 elements, i.e., P, S, Cl, K, Ca, Fe and Zn in the artery walls. Early modifications of the vessels, due to the atherosclerosis progression, that cannot be detected with specific staining or conventional histological methods could be detected. These modifications comprise the disruption of elastic laminae and are apparently related to abnormal Fe and Zn depositions in the intimal region and surroundings of the elastic laminae. Later on, the calcification of these regions occur, contributing to an irreversible damage of the elastic laminae and leading to the atheroma growing and maturation. Moreover, the Fe and Zn content alterations and distribution in the vessel wall can characterize stages of the disease.

Arteriosclerosis↗

PubMed1996

Pixe analysis of pathological skin with special reference to psoriasis and atopic dry skin.

The nuclear microprobe is used for element analysis of human skin cross sections, providing new insight into the physiology of normal and pathological conditions. Special interest is focused on trace elements as they work as secondary messengers or regulatory substances. The distribution of ions in normal tissues serves as reference for pathological changes. Calcium (Ca2+) is assumed to take an important role in the differentiation process of the epidermis. This paper presents new data on elemental and trace elemental distributions in skin. Samples are prepared from skin biopsies obtained from patients with skin disorder and from individuals with no records of skin disorder serving as controls. Using the nuclear microprobe, both elemental maps and quantitative depth profiles are obtained. Previous findings of abnormal Fe distribution in psoriatic skin are confirmed, and new observations of altered Zn and Ca profiles in atopic skin are reported. The relation to possible physiological/biochemical mechanisms and apoptosis ("programmed cell death") is discussed. The study is a part in a larger survey aiming at an understanding of the formation of a mature stratum corneum with a functional barrier, and its changed properties in cases of skin disorder.

Apoptosis↗

PubMed1995

Distribution of calcium and sulphur in the blue-light-exposed rat retina.

BACKGROUND: Blue-light exposure inhibits cytochrome oxidase and may therefore inhibit retinal metabolism. The reduced metabolism decreases the extrusion of calcium from the photoreceptor cell. Overload of calcium is proposed as one of the factors that lead to photoreceptor degeneration after light exposure. The light-induced photoreceptor degeneration can be ameliorated by calcium overload blocker. In the present study the calcium concentration was measured in the inner and outer segment layer of the rat retina. METHODS: Six eyes were exposed to blue (404 nm) light at a retinal dose of 380 kJ/m2. Five eyes served as the control group. The calcium and sulphur distributions were measured with a nuclear microprobe in the freeze-dried rat retina. The proton beam size was 12 x 12 microns and the energy of the protons was 2.55 MeV. The calcium concentration was calculated using sulphur as a reference. RESULTS: The level of calcium per milligram sulphur was 21 micrograms (range 17-23 micrograms) in the inner segment of the control retina. It increased to 62 micrograms/mg sulphur (range 57-67 micrograms) and 61 micrograms/mg sulphur (range 58-66 micrograms) 1 h and 12 h after blue-light exposure, respectively. CONCLUSION: The findings of the present study support the idea that accumulation of calcium in the inner segment layer is one of the factors that cause photoreceptor degeneration.

Animals↗

PubMed1995

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.

Apoptosis↗

PubMed1994

New applications of the nuclear microprobe for biological samples.

The continuing development of the Nuclear Microprobe (NMP) has opened up new fields of applications in biology and medicine. Quantitative multielemental analysis in small sections of samples can be performed routinely. The use of techniques such as scanning transmission ion microscopy makes imaging as well as mass normalization possible at submicron resolution. Recent medical applications include studies on thin cryosections prepared from autopsies and biopsies, as well as single cells grown directly on the backing foil used in the NMP analyses. The purpose of the single cell analysis is often pharmacological, e.g., testing of new drugs, their uptake and distribution. New applications, for instance, in food chemistry, ecology and evolutionary genetics, are also taking advantage of the high analytical sensitivity of the NMP in combination with its imaging capability.

Alzheimer Disease↗

PubMed1993

Quantitative elemental mapping of biomedical specimens using the nuclear microprobe.

Quantitative and structural elemental information is available from the nuclear microprobe through a time consuming fit of the (X-ray) spectrum at each point. An alternative technique is proposed which greatly shortens the analysis procedure and allows an increased number of samples to be processed. The method discussed here is to create elemental maps which, when they are divided by the charge/pixel and multiplied by a scaling factor, will form quantitative maps. The scaling factors are obtained from a calibration procedure comparing a large number of fitted X-ray spectra with the corresponding contents of selected energy windows. The technique also allows the reduction of artefacts due to spectral overlap, assuming that a simple background model can be used.

Animals↗

PubMed1991

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.

Animals↗

PubMed1990

Accumulation of calcium in substantia nigra lesions induced by status epilepticus. A microprobe analysis.

Status epilepticus of sufficient duration (greater than 30 min) causes a unique lesion of substantia nigra pars reticulata (SNPR), and of globus pallidus (GP). This lesion, which encompasses a pan-necrotic destruction of neurons and glial cells seems to develop during ongoing seizures. We decided to investigate if the lesion is accompanied by net calcium accumulation. Seizures of 20 and 60 min duration were induced by the administration of flurothyl, and the tissue was frozen in situ either at the end of the seizure periods, or after recovery periods of 15 or 120 min. The total calcium and potassium contents of caudoputamen, neocortex, GP and SNPR were measured using particle induced X-ray emission (PIXE) in the microprobe mode. Seizures of 20 min duration did not cause net accumulation of calcium. When the duration of seizures was extended to 60 min the results varied depending on the location. In caudoputamen, which does not incur neuronal damage, no calcium accumulation was observed. In GP and SNPR, such a rise was unequivocally demonstrated, with calcium content increasing to about 150% of controls. The increase in calcium correlated to a decrease in potassium content. It is concluded that epileptic cell death occurs pari passu with accumulation of calcium although it cannot be stated that this accumulation is the cause of the cell death. It is speculated that seizures increase the permeability of the blood-brain barrier to calcium, and that enhanced blood to tissue transfer increases the calcium load of metabolically strained cells.

Animals↗

PubMed1989

High concentrations of chromium in lung tissue from lung cancer patients.

The pulmonary chromium content was determined by plasma atomic emission spectrometer (DCP-AES) from 53 lung cancer and 43 control patients, and compared with smoking habits, severity of emphysema and occupational history. The chromium content from the lung cancer patients was higher than that from the smoking (P less than 0.025) or nonsmoking control patients (6.4 +/- 4.3, 4.0 +/- 4.0, and 2.2 +/- 0.6 microgram/g dry weight, respectively). A positive correlation between the pulmonary chromium and smoking time (P less than 0.025) and the severity of emphysema (P less than 0.001) was found in the control but not in the cancer patients. The difference in the pulmonary chromium content was greatest between those lung cancer and control patients who were light smokers or had mild emphysema. This group of lung cancer patients included subjects with occupational exposure to chromium. The possibility of occupational cancer should be considered especially with light smokers. The grade of emphysema and metals such as chromium accumulating from tobacco could serve as objective indicators of smoking.

Aged↗