I investigated the effect of inhibiting NO production in the psoriatic plaque. A 70% inhibition of NO was attained but overall there was no clinical benefit in 17 patients whereas with topical steroids benefit was very apparent with only 12 subjects. Immunocytochemical examination of these patients revealed a 50% reduction in endothelial cells, following L-NMMA treatment, supporting an important effect of NO in angiogenesis. ICAM-1 was also suppressed and there was an increase in CD8 positive cells and Langerhans' cells following L-NMMA. Surprisingly there was no inhibition of keratinocyte proliferation with L-NMMA. It is possible that some of the immunosuppressive effects of NO were inhibited, thus promoting some aspects of psoriasis at the same time as inhibiting others. Finally the direct effects of NO on normal skin were examined using acidified nitrite as a topically applied NO donor. In keeping with the observations in psoriasis there was a reduction in Langerhans' cells which lost dendricity and migrated from the epidermis. This suggests that NO may have some role in activating Langerhans's cells and thereby in antigen presentation. NO increased the adhesion molecules ICAM-1 and VCAM-1, attracting increased numbers of macrophages, T cells and neutrophils but had no effect on mast cell numbers. There was a tendency for higher doses of NO to produce lower levels of cellular infiltration, suggesting an immunosuppressive effect of higher doses. Nitrosotyrosine staining was not marked, suggesting only minor formation of peroxynitrite and this led to recoverable cytotoxicity manifesting as cloudy swelling of keratinocytes. There was some increase in wild type p53 in basal keratinocytes. Associated with this was an increase in apoptosis which was modest but manifested in the epidermis and dermis and was greater with the duration of exposure to NO. There was a high background level of cell proliferation and these cells were no more numerous in the NO treated skin than in the normal skin. However, there were significantly more proliferating cells in skin exposed to the higher dose of NO than those exposed to low dose suggesting that NO may promote keratinocyte proliferation at certain concentrations.

Occupational skin diseases is one of the most commonest occupational disease groups accounting for about a fifth of all occupational diseases in the UK. Current guidance from the HSE for skin health surveillance is the skin inspection by a responsible person in the workplace. The use of teledermatology can be attractive to reliably conduct skin surveillance in the workplace, and a tool box to take reproducible standardised photographs from the hands of workers in the workplace was developed. Aim of this thesis was to assess diagnostic accuracy and validity of this toolkit with visual inspection as criterion standard for the presence of minor or major hand dermatitis and by scoring the hands and photographs with the validated Hand Eczema Severity Index. Workers from four different occupations were recruited over a 7 month period and 332 workers were assessed on a repeatedly basis producing 1212 assessments. Skin hydration and transepidermal water loss was measured and a symptoms questionnaire was completed by every participant. A high prevalence of skin problems was found in our study: 70% of the participants presented at least once over the study period with minor or major skin symptoms. A high intrarater reliability compared to the visual assessment was shown for the teledermatologic assessment with an agreement of 88%, kappa of 0.79, and a positive likelihood ratio of 7.4 and negative likelihood ratio of 0.07; about 5% of participants with normal skin were over-diagnosed compared to the visual inspection. The interrater reliability was low. The biophysical parameter did not distinguish between normal and affected skin. The tool kit has shown to produce reliable and standardised high quality photographs, the assessment of the photographs showed a very good intrarater agreement to the criterion standard. The toolkit would allow regular skin surveillance with minimal interruption in the workplace and with reliable results from the assessment.

Bovine Dermatitis digitalis (DD) ist eine weltweit verbreitete Infektionskrankheit bei Rindern, die primär die plantare Haut über dem Kronenrand nahe des Zwischenzehenspalts der Hinterklauen betrifft. Schmerzhafte ulzero-proliferative Läsionen mit akuten und chronischen Erscheinungsformen führen zu Verhaltensänderungen und Lahmheit der Tiere. DD hat damit einen erheblichen Einfluss auf deren Wohl und ihre Leistungen. Zahlreiche Untersuchungen zur Ätiologie der Krankheit ergaben, dass es sich um das Zusammenspiel verschiedener Ursachen handelt. Einer synergistischen multifaktoriellen Infektion mit starker Beteiligung von Bakterien der Gattung Treponema kommt dabei besondere Bedeutung zu. Aspekte wie Tierhaltung, Hygienestandards und genetische Prädispositionen wurden ebenfalls intensiv untersucht. Nichtsdestotrotz bleiben Infektionsherde, Transmissionsrouten und Pathomechanismen weitgehend unklar. Zum besseren Verständnis der Ereignisse, die zu DD-Läsionen führen, sollte im Zuge dieser Arbeit ein organotypisches Zellkulturmodell der bovinen Haut erstellt werden, welches in späteren Versuchen mit dem Krankheitserreger zum Einsatz kommen soll. Verlässliche und reproduzierbare Techniken zur Isolation und Kultur von bovinen primären Keratinozyten und Fibroblasten wurden etabliert; geeignete Zellkulturmedien für die Langzeitkultivierung und –aufbewahrung der Hautzellen wurden identifiziert. Zur Erstellung des Hautmodells wurden zwei verschiedene Ansätze miteinander verglichen. Der zweite Ansatz, bei dem Keratinozyten direkt auf ein dermales Äquivalent, d.h. ein Pad aus bovinem Kollagen I mit eingesäten post-mitotischen Fibroblasten, gesät wurden, brachte ein vielversprechendes Hautmodell hervor. Die inkorporierten post-mitotischen Fibroblasten wiesen eine charakteristische Zellmorphologie mit intakten Nuklei auf. Die terminale Differenzierung der Keratinozyten auf dem dermalen Äquivalent wurde mittels Immunfluoreszenzfärbungen mit Antikörpern gegen die Markerproteine Keratin 14 und Desmoglein 1 gezeigt. Die Ergebnisse erster Experimente mit Treponema spp. verdeutlichen, dass das Hautäquivalent ein geeignetes Modell zur Untersuchung der Pathogenese der DD darstellt. / Bovine digital dermatitis (DD) is a worldwide occurring, infectious disease in cattle primarily affecting the plantar skin above the coronary band near the interdigital cleft on hind feet. Painful ulceroproliferative lesions with acute and chronic appearances lead to behavioral changes and lameness. Hence, DD has a major impact on animal welfare and performance. Substantial efforts in investigating the etiology of the disease revealed a synergistic origin with evidence for a multibacterial infection and the strong involvement of bacteria from the genus Treponema. As the interaction between host, pathogen and environment is not negligible, surrounding circumstances such as housing, general hygiene and genetic predispositions have been investigated intensively. Nevertheless, infection reservoirs, transmission routes and pathomechanisms remain widely unclear. To better understand the cellular and molecular events during Treponema-infection of bovine skin, it was the specific aim of this study to establish an organotypic in vitro skin model, which could be challenged with the causative agent of the disease. A technique to reliably and reproducibly isolate primary keratinocytes and fibroblasts from the site of infection was established. Appropriate cell culture media for the long-term cultivation and storage of bovine skin cells were identified. Two different methods to develop the skin model were compared. The second strategy in which keratinocytes were directly seeded on top of a dermal equivalent, i.e. a bovine collagen type I pad with embedded post-mitotic fibroblasts, gave rise to a promising organotypic skin equivalent. The incorporated post-mitotic fibroblasts showed a characteristic cell morphology with intact nuclei. The terminal differentiation of the keratinocytes on top of the dermal equivalent was shown with anti-K14 and anti-Dsg1 immunofluorescence stainings. The results of initial Treponema-experiments proved that the skin equivalent is a suitable model to investigate the underlying mechanisms during Treponema-infection of bovine skin and hence, the pathogenesis of DD.