Epidermal keratinocytes studied by X-ray microanalysis, with special reference to contact dermatitis : In vivo and in vitro studies of functional changes in human cells

Sammanfattning: By determining the intracellular elemental content, information can be obtained on the physiological status of cells. The cellular content of sodium, magnesium, phosphorus, sulfur, chloride, potassium and calcium can be indicative for proliferation, cell membrane damage or cell death. X-ray microanalysis is a method for measuring the intracellular elemental content. In the present study this technique has been used for in vivo and in vitro studies of human epidermal keratinocytes under different physiological conditions. Focus has been on irritant reactions, proliferation/hyperplasia and on methodological aspects. As a part of this, studies on the effects of local anaesthesia on normal skin and the effects of calcipotriol on psoriatic skin were included Irritant reactions were induced with sodium lauryl sulfate and non-anoic acid. In studies on skin invivo, X-ray microanalysis was combined with reverse transcriptase-polymerase chain reaction to determine the cytokine profile and with analysis with the monoclonal antibody Ki67 to determine the proliferation following application of the two irritants. By studying keratinocytes in cell cultures, it is possible to eliminate the influence from other cells in the skin and to perform more controlled studies. In the last part of the study the influence of methodological factors, cell type and exposure to sodium lauryl sulfate on the cellular elemental content were evaluated in cultured keratinocytes.Following application of the irritants to the skin it was possible to identify an early response at 6-24 hours with elemental changes compatible with a cell membrane damage followed by a proliferation with a maximum at 24-48 hours later. It was also possible to detect a difference in the response between the two applied irritants. The elemental changes found in the proliferative phase were similar to those found in psoriatic skin. Calcipotriol has an anti-proliferative effect and this was also reflected in the elemental content of the keratinocytes aftercalcipotriol treatment The in vitro analysis of cultured cells revealed that the elemental content of the cells differed between cell types, preparation techniques and with the age of the cell culture. It was also demonstrated that exposure to sodium lauryl sulfate caused time, and dose dependent variations similar to those found at irritant reactions in human skin. In conclusion, X-ray microanalysis was demonstrated to be a suitable technique for studies on physiological changes in the epidermal keratinocytes at irritant reactions. Furthermore it was shown that the technique can be used to evaluate effects on the keratinocytes in cell cultures and thus be utilized as a method to predict damaging effects of different chemicals.