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Biomedical subjects

P Nau

Publications and source records attributed to P Nau.

8 recordsLinked to original sources

Formation of the epidermal calcium gradient coincides with key milestones of barrier ontogenesis in the rodent.

The epidermal permeability barrier forms late in gestation, coincident with decreased lipid synthesis, increased lipid processing, and development of a mature, multi-layered stratum corneum. Prior studies have shown that changes in the epidermal Ca++ gradient in vivo regulate lamellar body secretion and lipid synthesis, and modulations in extracellular Ca++ in vitro also regulate keratinocyte differentiation. We asked here whether a Ca++ gradient forms in fetal epidermis in utero, and whether its emergence correlates with key developmental milestones of barrier formation and stratum corneum development. Using either ion precipitation or proton induced X-ray emission analysis of fetal mouse and rat skin, we showed that a Ca++ gradient is not present at gestational days 16-18, prior to barrier formation, and that a gradient forms coincident with the emergence of barrier competence (day 19, mouse; day 20, rat) prior to birth. These results are consistent with a role for Ca++ in the regulation of key metabolic events leading to barrier formation. Whether the calcium gradient is formed actively or passively remains to be determined.

Animals↗

Microwave incubation improves lipolytic enzyme preservation for ultrastructural cytochemistry.

Standard methods for the ultrastructural detection of lipase and sphingomyelinase activities in the skin result in considerable loss of structural preservation, often interfering with accurate delineation of enzyme localization in association with specific organelles. Moreover, poor preservation occurs, even after extensive aldehyde prefixation, owing to the prolonged incubation times needed to detect residual enzyme activity, which often require non-physiological conditions. A modified incubation protocol is described here, which uses microwave irradiation in conjunction with drastically shortened incubation times, resulting in both superior ultrastructural preservation and excellent localization in mammalian epidermis. This method should be useful generally not only for the study of lipase localization in skin, but also in conjunction with the cytochemical detection of a variety of enzymes in various types of tissue.

Animals↗

Congenital self-healing reticulohistiocytosis--a benign Langerhans cell disease.

An-11-day old girl was seen with brownish nodular lesions scattered over the body with emphasis on the face and scalp. Several lesions had started to involute. Tissue was studied by histopathology, immunohistopathology, routine electron microscopy, and immuno-electron microscopy using cryosubstitution and embedding in K4M. Immunohistopathology revealed that the cells of the dermal infiltrate were Langerhans cells. They expressed Leu 6 and HLA-DR. On routine electron microscopy no Birbeck granules were found in the dermal cells. Birbeck granules in epidermal Langerhans cells were deformed and often situated next to laminated dense bodies. The latter expressed Leu 6 and lysozyme on immuno-electron microscopy. It was concluded that congenital self-healing reticulohistiocytosis is a benign Langerhans cell disease in which Birbeck granules are transformed to laminated dense bodies and possibly degraded by lysosomal enzymes.

Female↗

Techniques in immuno-electron microscopy. I. Cryosubstitution.

Normal skin was cryoprotected by submerging it in a mixture of 30% dimethylformamide (DMF) in PBS or RPMI. Subsequently it was frozen in liquid propane gas. Cryosubstitution was carried out at -90 degrees C by using methanol to which uranyl acetate or osmium tetroxide were added. The tissue was embedded in either Lowicryl K4M at -40 degrees C or in Epon at +60 degrees C. The tissue was evaluated by its overall preservation of ultrastructural details and by its labeling intensity after incubation with either anti-desmoglein or anti-type VII collagen monoclonal antibodies. The mixture of DMF and PBS caused an electron-dense precipitate within the cell. The overall morphology was better in Epon-embedded material than in K4M-embedded material. However, the labeling was best in K4M material. Regardless of whether the tissue was embedded in Epon or K4M, the addition of osmium tetroxide markedly reduced the degree of labeling.

Freeze Substitution↗

Ultrastructural localization of lectin binding sites in human melanocytes.

The ultrastructural localization of carbohydrate residues in human melanocytes of normal epidermis and of one compound naevus was studied. The following lectins were used in a post-embedding technique: 1. peanut agglutinin (PNA), which reacts specifically with N-acetylgalactosamine; 2. Concanavalia ensiformis (Con A) indicating alpha-D-glucose and alpha-D-mannose binding sites; 3. Ulex europaeus agglutinin (UEA I) specific for alpha-L-fucose; 4. Wheat germ agglutinin (WGA), reacting specifically with N-acetyl-glucosamine and neuraminic acid (sialic acid); and 5. Limax flavus agglutinin (LFA), also specific for sialic acid (Neu5Ac-alpha-2,3-Gal and Neu5Ac-alpha-2,6-Gal). When incubated with WGA, Con A and LFA strong labelling was seen within the cytoplasm and in the plasma membrane of melanocytes, whereas incubations with PNA and UEA I revealed an occasional gold particle only. The determination of the distribution of carbohydrate residues in normal melanocytes is a prerequisite for future studies of abnormal melanocytes.

Arachis↗

Human peripheral blood accessory cell: isolation by hypotonic density gradient, functional, and phenotypical characterization.

In order to purify the human peripheral blood-derived accessory cell that cooperate with T lymphocytes in the process of mitogenic stimulation, we developed a new density gradient separation. This was based on the principle of hypotonic swelling of the cells to obtain a differential change of the buoyant densities of cells. By this method, we have obtained a highly accessory cell-depleted lymphocyte fraction whose proliferative response to sodium periodate stimulation was almost aborted. Another fraction containing high accessory cell activity was further divided into Fc-receptor-positive and -negative cells. The latter revealed the highest accessory activity for T lymphocyte periodate stimulation. The cells were characterized according to a number of markers and appeared to resemble lymphoid dendritic cells. Compared with the monocyte/macrophage fraction, they showed veils and dendritiform elongations and expressed reduced values of monocyte/macrophage specific markers. Compared with the high accessory activity of these cells, monocytes/macrophages expressed a low accessory activity.

Antigen-Presenting Cells↗

An analysis of public posting in reducing speeding behavior on an urban highway.

The effects of public posting on speeding behavior on an urban highway were assessed using a reversal design. During baseline the percentage of drivers speeding was measured with a concealed radar unit. During the daily posting condition a highway sign was installed which provided feedback on the percentage of drivers not speeding yesterday and the best record to date. This sign was then covered and reintroduced. Results indicated that the sign was effective in reducing speeding behavior. Furthermore, the effects were most pronounced in reducing the speeds of the faster drivers. Next, daily and weekly postings were compared with the sign alone without numerical feedback: results revealed that the weekly posting condition was as effective as the daily posting condition, but that the sign had no influence when numbers were not posted. Finally, the weekly posting procedure remained effective during a 6-month follow-up condition.

Journal Article↗