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The response to wounds until healing requires the activity of many cell types coordinate in space and time, so that the types of cells in a wound and their localization may be of help to date lesions with respect to death, which would be useful in forensic pathology. Cells reacting to injury include dendritic cells; the early reaction of these cells to skin wounding has not yet been investigated in humans, which was the aim of this study. Samples of wounded and control skin were taken at autopsy and analyzed by affinity histochemistry. Both epidermal and dermal MHC-II+ cells increased transiently in number within the first hour after wounding, then decreased. In the epidermis the increase affected also CD1a+ cells, i.e. well differentiated Langherhans cells, which however increased less, earlier and for a shorter time period than MHC-II+ cells. Dermal MHC-II+ cells became part of a perivascular mononuclear cell infiltrate visible in the subpapillary dermis by 60 min after wounding, which contained also mast cells. The immediately perivascular MHC-II+ cells were DC-SIGN- and CD11c-, while MHC-II+, DC-SIGN+, CD11c+ dendritic cells were predominantly located at the periphery of infiltrates and some were near the epidermis. Mast cells underwent degranulation, besides increase in number, in the first hours after wounding. The results suggest that skin dendritic cells, including Langerhans cells, participate to the early response to wounding in concert with mast cells, and that subpapillary blood vessels are primary sites of cell infiltration during that response in humans. The results show that the ratio between CD1a positive and MHC-II positive cells in the epidermis, the degranulation index of mast cells and the relative volume of MHC-II positive cells in the dermis can be added to the tools useful to distinguish vital from post mortem lesions and, the first two of them, to estimate the interval between a lesion and death., (Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.)

Dendritic cells and their precursors express PPAR-gamma, whose stimulation has inhibitory effects on the maturation and function of dendritic cells in vivo. Dendritic cells can differentiate in vitro from CD133+ progenitors; the influence of PPAR-gamma stimulation on this process is unknown. We have addressed the effect of PPAR-gamma agonist rosiglitazone, at a concentration as used in clinics, on the differentiation of dendritic cells from human CD133+ progenitors. Cells were harvested from cord blood by density gradient and immunomagnetic separation, and cultured for 18 days with fetal calf serum, cytokines and 1 μmol/L rosiglitazone. Analyses included flow cytometry, electron microscopy and mixed lymphocyte reaction. As expected, control cells generated without rosiglitazone were dendritic, expressed MHC-II, CD80, CD83 and CD86 and stimulated mixed reaction potently. A minority of cells expressed the Langerhans cell marker CD207/langerin, but none contained Birbeck granules. With rosiglitazone much fewer cells were generated; they were all dendritic, expressed differentiation and maturation-related antigens in higher percentage and were better stimulators of lymphocytes than those generated without the drug. The vast majority of cells expressed CD207/langerin and many contained Birbeck granules, i.e. were full-fledged Langerhans cells. We conclude that stimulation of PPAR-gamma, while negatively affecting the number of generated cells, promotes the maturation of human cord blood CD133 positive precursors into efficient, immunostimulating dendritic cells with a Langerhans cell phenotype.

Langerhans cells are a dendritic cell type characteristic of the epidermis. Since mast cells secrete molecules potentially influent on dendritic cell differentiation, we have addressed the degree of proximity between these two cell types in biopsies of skin diseases characterized by massive influx of dendritic cell precursors. By fluorescence microscopy, avidin labeled mast cells were found in contact with CD1a+ dendritic cells. By electron microscopy, contacts between mast cells and blunty dendritic cells were found in areas corresponding to those where CD1a+ cells were localized by immunohistochemistry. We propose that mast cells can induce the differentiation of precursors into Langerhans cells through both the release of short range acting soluble factors and contact-mediating plasma membrane molecules.

Langerhans cells (LC) play a critical role in the induction of contact hypersensitivity (CH), and ultraviolet B radiation (UVR) impairs CH induction in UVB-susceptible (UVB-S) mice via a TNF-alpha-dependent mechanism. A possible explanation of this effect is that UVR impairs CH in UVB-S mice by immobilizing LC transiently in the epidermis and upper dermis, thereby preventing their timely migration to draining lymph nodes. To test this hypothesis we examined in vitro and in vivo the effects of in vivo UVR on migration of Ia(+) cells from skin of UVB-S and UVB-resistant (UVB-R) mice. Dorsal surfaces of ears of mice were irradiated with 400 J/cm(2) UVB and either explanted in vitro or transplanted orthotopically to the thoracic wall of syngeneic recipients. After 24, 48, and 72 h the epidermis was recovered from these explants/grafts and the number of Ia(+) cells determined by immunohistochemistry. Culture medium obtained after explants were removed was also evaluated for content of Ia(+) cells. The density of Ia(+)-bearing cells in the epidermis of cultured untreated skin explants and of grafted skin fell progressively for both UVB-S and UVB-R skin during the observation period. The rate of decline in Ia(+) cells density during this interval was greatly impaired if the skin was exposed to UVR prior to excision; this effect was seen equally in UVB-S and UVB-R skin. Recovery of Ia(+) cells in the medium after removal of cultured untreated skin explants was maximum after 24 h and comparable in UVB-S and UVB-R skin. However, the number of Ia(+) cells recovered in the medium from UVB-exposed skin was significantly reduced only if the skin donor was UVB-S. We conclude that the ability of UVR to impede LC migration from epidermis is significantly greater for UVB-S mice, accounting in part for the failure of these mice after UVR to acquire CH.

The immunosuppressant drug cyclosporin A (CsA) is known to cause reduction in number, DNA synthesis and function of Langerhans cells (LC). Since also the differentiation of LC is known to be hampered in conditions of acquired immunodeficiency not due to drugs, we investigated whether this occurs with CsA. Rats were injected subcutaneously with CsA (5, 10 and 50 mgxkg(-1) x d(-1)) for three weeks; the skin was analyzed by Ia immunohistochemistry and by electron microscopy. Epidermal immunolabeled cells were 15+/-3.5 (mean +/- SEM) per 100 basal keratinocytes in untreated controls and 8.75+/-1.3, 4.75+/-1.0 and 1.7+/-1.2 upon increasing doses of CsA (p<0.01). By electron microscopy, monocytoid cells with deep invaginations of the plasma membrane and roundish LC poor in Birbeck granules appeared in the epidermis upon treatment. The results suggest that CsA inhibits the differentiation of LC precursors in the epidermis and that this can in part explain the selective increase in the risk of skin viral disease and cancer in chronically treated patients.

Acute, low-dose ultraviolet B (UVB) radiation impairs contact hypersensitivity induction in mice by a mechanism due at least in part to Langerhans cells alterations. To better define the effects of UVB on Langerhans cells, we have compared the action of this agent on the skin of intact mice and in skin explants incubated in vitro up to 24 h. Using immunofluorescence, we detected a reduction in the length of the dendrites of Langerhans cells and a significant reduction in the number of Ia-positive Langerhans cells per unit area within 2 h of UVB; these changes reversed within 24 h in vivo, but not in vitro. By electron microscopy, the number of dendritic cells per 100 basal keratinocytes increased in vivo, but decreased in vitro by 2 h after UVB, a discordance that was significant. On the contrary, the number of dendrite profiles per dendritic cell body decreased significantly 2 h after UVB, both in vivo and in vitro. Many epidermal dendritic cells, 2 h after UVB in vivo, were deficient in cytoplasmic organelles, whereas the few cells that remained after UVB in vitro retained their Birbeck granules, and displayed many, dilated cytoplasmic vesicles. We interpret these data to mean that low doses of UVB radiation destroy the functional and morphologic integrity of epidermal Langerhans cells, and that these cells are rapidly replaced by precursor cells that mature in situ into normal-appearing Langerhans cells.

Acute low-dose ultraviolet B radiation (UVR) impairs contact hypersensitivity (CH) induction in genetically defined strains of mice by a mechanism triggered by cis-urocanic acid (UCA) and dependent upon tumor necrosis factor-alpha (TNF-alpha). UVR, TNF-alpha, and cis-UCA cause similar morphologic changes among Langerhans cells, which spawns the speculation that UVR impairs CH induction in part by altering the Langerhans cell cytoskeleton. To examine this speculation, we studied the expression of vimentin in Langerhans cells after treatment with UVR, TNF-alpha, and cis-UCA. All treatments caused a reduction in expression of vimentin within the cytoplasm of Langerhans cells. Because partial loss of detectable vimentin may correlate with cytoskeletal disruption, we evaluated the effects of vinblastine, an agent that disrupts the cytoskeleton by disassembling microtubules, on Langerhans cell density and morphology. Epicutaneous treatment with vinblastine caused a reduction in Langerhans cell density, a loss of dendrites, and a reduction in vimentin expression. When dinitrofluorobenzene was painted on vinblastine-treated skin of BALB/c or C3H/HeN mice, only feeble CH was induced. Consequently, we propose that UVR prevents CH induction in susceptible mice by disrupting the cytoskeleton of Langerhans cells, thereby preventing them from carrying out their crucial role as antigen-presenting cells.

Recent studies show that Langerhans cell density increases in vital lesions and ligature marks when compared with post-mortal wounds. The enzyme, iNOS, has been established as a marker for estimating time of agony and serves in the regulation of dendritic cell behaviour. It is the aim of this paper, therefore, to evaluate the expression of this enzyme by Langerhans cells and the possible consequences that may be related to the production of nitric oxide by these cells in other types of lesions, including hanging furrows. The results show a greater expression of iNOS by Langerhans cells at the level of the hanging furrow when compared with other examined groups. Apart from an increase in the expression of iNOS, a large fraction of the mast cell population in the class II MHC molecules was observed in the hanging furrow. This corroborates that interactions between mast cells and dendritic cells are critical for the differentiation of these latter cellular types. Hence, the results suggest that iNOS plays a crucial role in the forensic practice of establishing the time interval in defining the vitality in hanging marks. [ABSTRACT FROM AUTHOR]

Hanging deaths are common and characterized by their brief survival time. Macroscopic and histological exams are frequently unable to distinguish vital lesions from post-mortem lesions. This pilot study investigates the dendritic and mast cells in hanging marks to establish vitality lesions. Skin specimens were taken from vital wounds, ligature marks, and post-mortem lesions. Cryosections were stained for haematoxylin eosin, avidin, CD1a and MHC class II+ antigens and examined under light or fluorescence microscopies. Using digitized photomicrographs the images were then analysed. Differences were found in the Langerhans cells and epidermal MHC density, the vital lesions and ligature marks and the other specimens. The results may prove useful in forensic practice when neither macro nor microscopic objective evidence of vitality is available. The results could very possibly support the hypothesis that the amount of time it takes to die from hanging is less than currently believed (< 5 min). [ABSTRACT FROM AUTHOR]