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The presence and tissue localization of macrophage migration inhibitory factor (MIF) in human skin were examined. Reverse transcription-polymerasc chain reaction analysis revealed that MIF mRNA was expressed in both surgically obtained normal human epidermis and primary cultured human keratinocytes. The expression of MIF was further conf'wmed by Western blot analysis, which demonstrated a single band at about 12.5 kDa using a polyelonal antibody against human recombinant MIF. Immunohistochemical studies showed that MIF existed in human epidermis, especially in the basal layer. The pathophysiological role of MIF in human skin remains undefined; however, the present results indicate that MIF may play an important role in immunity, inflammation and cellular differentiation of epidermal cells.

After the cDNA of human macrophage migration inhibitory factor (MIF) was cloned in 1989, this protein has been re-evaluated as a pro-inflammatory cytokine, pituitary hormone and glucocorticoid-induced immunoregulatory protein. We previously reported the expression of MIF in the basal cell layers of the epidermis, but its pathophysiological function in the skin has not been well understood. In this study, we examined the expression of MIF during the wound healing of rat skin injured by excision. Reverse transcription-polymerase chain reaction in combination with Southern blot analysis revealed that the increase of MIF mRNA expression was biphasic. The maximum peaks were observed at 3 and 24 h after the injury. Similarly, maximal increases of the serum MIF level were observed at 3 and 24 h after the injury. Immunohistochemical analysis at 12 h after injury demonstrated enhanced expression of MIF protein in the whole epidermal lesion of the wound tissue. By the Boyden chamber assay, we demonstrated that MIF had a chemotactic effect on freshly prepared keratinocytes from rat skin. Additionally, cultured fibroblasts from the skin wound lesion secreted a higher amount of MIF in response to lipopolysaccharide compared to those of the normal skin. Furthermore, administration of anti-MIF antibodies induced a delay of wound healing in vivo. Taken together, these results suggest that MIF contributes to the wound healing process of skin tissue.

The Fas antigen is a cell surface protein that can mediate apoptosis in many cell types. Although its physiological function is still unclear, recent evidence indicates that this surface molecule is involved in apoptosis in the immune system and the liver. The epidermis is an organ that undergoes terminal differentiation with the eventual death of keratinocytes, and it has been suggested that this is a specialized form of apoptosis. In the present study, we examined whether or not the Fas antigen is involved in keratinocyte apoptosis. Immunoreactivity for the Fas antigen was found throughout the epidermis in normal human skin sections and cultured normal human keratinocytes, and mRNA for the Fas antigen was found to be constitutively expressed in normal epidermis and cultured normal keratinocytes by RT-PCR analysis. To determine whether the Fas antigen in keratinocytes is functional, we used a cytotoxic monoclonal antibody (mAb) against the Fas antigen to induce apoptosis. This antibody did not induce apoptosis of cultured keratinocytes even though they expressed the Fas antigen. We then tested the ability of several cytokines (TGF beta, TNF alpha and IFN gamma) to induce Fas-mediated keratinocyte apoptosis. Only pretreatment with IFN gamma followed by the addition of the anti-Fas mAb induced apoptosis, as assessed by cell viability, morphological changes and ultrastructural characteristics, suggesting that constitutive expression of the Fas antigen is not sufficient to induce apoptosis in keratinocytes and that keratinocyte apoptosis via the Fas antigen-mediated mechanism may require the activation of keratinocytes by IFN gamma, which is thought to be produced by activated T cells.(ABSTRACT TRUNCATED AT 250 WORDS)