Conference Proceeding: Abstracts of 39th Annual Meeting of SCUR (Society for Cutaneous Ultrastructure Research) May 23-25, 2012, Lyon, France multiple Authors & MUSS WH (original) (raw)
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British Journal of Dermatology, 1994
Confocai scanning laser microscopy (CSLM). when used in conjunction with computerized image processing systems, provides a powerfu! tool for morphological and quantitative analyses of biological tissues. In this study, normal human epidermal sheets were stained by an indirect immunofluorescence method using anti-CDla monoclonal antibody. Positively stained epidermal Langerhans cells (LCs) were visualized using the Bio-Rad MRC-600 Confocal Imaging System. Images obtained from the confocal microscope were volumetrically rendered and quantitatively analysed using ANALYZE® (Version 40} running on a Sun SPARC 2 Workstation. Normal epidermal LCs were shown to be large disc-like structures with five to nine long dendritic processes per cell, orientated with their flat surfaces parallel to the skin surface. LCs form a monolayer network of cells distributed evenly throughout the suprabasa! layers of the epidermis, with no direct physical contact between dendritic processes. Mean LC density was estimated to be 582 per mm" (95% confidence intervals. CI ^ 23 3-940). and mean cell volume was 612//m^ (95% CI ^ 257-1020). LCs In sun-exposed sites were significantly lower in mean cell density, but larger in mean cell volume, than in covered sites. Mean surface area projected by LCs was estimated to be 26-8% (95% CI-18 9-34 2), and this value did not show significant regional or individual variation. Our data support the notion that epidermal LCs are organized in such a way as to maximize their surface area for efficient trapping of antigens, and a reduction in LC density per unit area in sun-exposed sites is compensated for by an increase in the mean cell volume.
12-09-21 Abstracts 39thSCUR Lyon2012final exd15741
Those of us working on the morphology of the permeability barrier tend to view any focal disruptions in the extracellular lamellar lipids of SC as a sign of barrier defect. While it holds true for human skin dysfunctions, and animal models for diseases, extending this view into all other species could lead to erroneous conclusions about optimal barrier functions that allow their successful environmental adaptations. The fallacy of such anthropomorphic view comes to light while evaluating adaptations of the skin barrier to various environments (ranging from terrestrial, aerial, aquatic and Fossorial) and specific types of stresses in both sub-mammalian species (Snakes, Aves) as well as Mammals from specialized habitats (Amphibious, aquatic, subterranean). My talk is focused on the diverse modifications to the basic theme of lamellar body secretion and processing of these probarrier lipids, and how this is exploited for successfully adaptating to specific environmental challenges. In some cases, facultative adaptations to short-term needs are also dependant on such modifications. The take-home message from these observations is that we should be asking 'when the barrier is good enough or optimal', rather than looking for the most water-tight permeability barrier.
Experimental Dermatology, 2007
Knowledge about the structural elements of skin and its appendices is an essential prerequisite for understanding their complex functions and interactions. The hence necessary morphological description across several orders of scale not only requires the investigation at the light microscopic level but also ultrastructural investigation, ideally on the identical sample. For a correlative and multimodal observation one unique preparation protocol is mandatory. As a compromise between sample sizes of >500 lm in diameter on the one hand and optimal preservation of antigenicity and morphology on the other, we developed a new preparation protocol that allows (i) 3D reconstruction of the resin-embedded sample by confocal light microscopy prior to (ii) direct immunolocalization of target proteins within selected sample planes by light and fluorescence microscopy or transmission electron microscopy. Alternatively, (iii) serial cryosections of the frozen sample can be taken for characterizing the sample in toto. With this unique approach we were able to fully demonstrate the structural complexity of axillary skin samples, increasing the structural resolution from 3D reconstruction of the whole gland up to ultrastructural investigations at the subcellular level. We could demonstrate that axillary sweat glands are not separately distributed, as has been assumed to date; instead, they seem to be intricately twisted into one another. This promotes the concept of a complex axillary sweat gland organ instead of single sweat gland entities.
Intraepidermal cell surface fine structure: Preservation and examination at high resolution
The Anatomical Record, 1979
Morphological and functional studies on cell surfaces have been limited largely to cultured cells because of injury wrought to cells of solid tissues by commonly employed mechanical, enzymatic, or chelator dispersal methods. By using the staphylococcal epidermolytic toxin we avoided this problem; the toxin cleaves the intercellular spaces of human and mouse squamous epithelia without ultrastructural evidence of cytotoxicity. We studied the cell surface topography of neonatal mouse epidermis obtained two hours after injection of highly purified epidermolytic toxin. Immediately after sacrifice intraepithelial surfaces were exposed while the animals were immersed in fixative. Specimens were either freeze-fractured or embedded for transmission electron microscopy, or were critical-point-dried prior to platinumkarbon replication for transmission and scanning electron microscopy. Replicas could be prepared for transmission electron microscopy only if they were first stabilized with parloidion and then cleaned with both bleach and 40% chromate. By using these four complementary morphological methods (freezefracture, scanning electron microscopy, transmission electron microscopy of surface replicas, and standard thin sections), we could positively identify external membrane structures. The convoluted surface was studded by tenuous microvilli, scattered 15-20 nm particles, and hemispherical desmosomal mounds. Desmosomal plaques displayed randomly arrayed 15-20 nm globular particles comparable in distribution and density to particles observed in freeze-fractured desmosomes, and suggesting that desmosomal integral membrane particles span the external leaflet of the plasma membrane.
Real-Time Visualization of Macromolecule Uptake by Epidermal Langerhans Cells in Living Animals
Journal of Investigative Dermatology, 2012
As a skin-resident member of the dendritic cell (DC) family, Langerhans cells (LCs) are generally regarded to function as professional antigen presenting cells. Here we report a simple method to visualize the endocytotic activity of LCs in living animals. BALB/c mice received subcutaneous injection of FITC-conjugated dextran (DX) probes into the ear skin and were then examined under confocal microscopy. Large numbers of FITC + epidermal cells became detectable 12-24 h after injection as background fluorescence signals began to disappear. Most (>90%) of the FITC + epidermal cells expressed Langerin, and >95% of Langerin + epidermal cells exhibited significant FITC signals. To assess intracellular localization, Alexa Fluor 546-conjugated DX probes were locally injected into IAβ-EGFP knock-in mice and Langerin-EGFP-DTR mice-three dimensional rotation images showed close association of most of the internalized DX probes with MHC class II molecules, but not with Langerin molecules. These observations support the current view that LCs constantly sample surrounding materials, including harmful and innocuous antigens, at the environmental interface. Our data also validate the potential utility of the newly developed imaging approach to monitor LC function in wild-type animals. Recent advances in mouse genetic engineering and intravital confocal imaging have enabled direct visualization of LC behaviors in living animals.
Multimodal label-free imaging of living dermal equivalents including dermal papilla cells
Stem cell research & therapy, 2018
Despite the significant progress in the development of skin equivalents (SEs), the problem of noninvasively assessing the quality of the cell components and the collagen structure of living SEs both before and after transplantation remains. Undoubted preference is given to in vivo methods of noninvasive, label-free monitoring of the state of the SEs. Optical bioimaging methods, such as cross-polarization optical coherence tomography (CP OCT), multiphoton tomography (MPT), and fluorescence lifetime imaging microscopy (FLIM), present particular advantages for the visualization of such SEs. In this study, we simultaneously applied several visualization techniques for skin model examination. We investigated the structure and quality of dermal equivalents containing dermal papilla (DP) cells and dermal fibroblasts (FBs) using CP OCT, MPT, and FLIM. Both the energy metabolism of the cell components and the structuring of the collagen fibrils were addressed. Based on the data from the fluo...