Sakuhei Fujiwara

Dermatopontin, a Novel Player in the Biology of the Extracellular Matrix

Dermatopontin is a widely distributed small molecular weight protein in the extracellular matrix (ECM) and today its homologues are known in five mammals and several invertebrates. The structures of these homologues are relatively well conserved among the species. In the skin, dermatopontin is located mainly on the surface of the collagen fibers. It is found in the conditioned medium and also in the cytoplasm of cultured fibroblasts. Early studies focused on ECM assembly (collagen fibrillogenesis) and interactions (with the proteoglycan decorin). Subsequently, a targeted disruption of dermatopontin resulted in a phenotype similar to Ehlers-Danlos syndrome. In addition, a cell adhesion activity of this protein for dermal fibroblasts and several other cells was found, and this activity might suggest this proteins involvement in wound healing. The expression of dermatopontin around an infarct zone of experimental myocardial infarction may support this possibility. In invertebrates, dermatopontin homologues act mainly as adhesion/agglutination molecules. In addition, we found that transforming growth factor-β1 interacts with dermatopontin and the function of this cytokine is modified by dermatopontin. Recently, the involvement of this protein in cell proliferation has been indicated. In this review we describe the reported functions of this protein and speculate on the multiple roles of this largely uncharacterized matrix molecule.

Elimination of Epiplakin by Gene Targeting Results in Acceleration of Keratinocyte Migration in Mice

ABSTRACT Epiplakin (EPPK) was originally identified as a human epidermal autoantigen. To identify the function of epiplakin, we generated epiplakin “knockout” mice. These mice developed normally, with apparently normal epidermis and hair. Electron microscopy after immunostaining revealed the presence of EPPK adjacent to keratin filaments in wild-type mice, suggesting that epiplakin might associate with keratin. The appearance and localization of keratin bundles in intact epidermal keratinocytes of EPPK−/− mice were similar to those in wild-type mice. Wounds on the backs of EPPK−/− mice closed more rapidly than those on the backs of wild-type and heterozygous mice. The outgrowth of keratinocytes from skin explants from knockout mice was enhanced compared to outgrowth from explants from wild-type mice, even in the presence of mitomycin C, suggesting that the difference in keratinocyte outgrowth might be due to a difference in the speed of migration of keratinocytes. At wound edges in wild-type mice, EPPK was expressed in proliferating keratinocytes in conjunction with keratin 6. In EPPK−/− mice, no similar proliferating keratinocytes were observed, but migrating keratinocytes weakly expressed keratin 6. EPPK was coexpressed with keratin 6 in some keratinocytes in explant cultures from wild mice. We propose that EPPK might be linked functionally with keratin 6.

A vesicular bullous pemphigoid with an autoantibody against plectin.

q 2000 British Association of Dermatologists, British Journal of Dermatology, 142, 812±851 remains uncertain. The exfoliative erythroderma in our patient is consistent with a pathogenic role for this autoantibody as it can be seen in patients with PF. In contrast, it has recently been demonstrated that the pathogenic effect of PNP sera containing both anti-Dsg1 and anti-Dsg3 antibodies was abolished by specific adsorption of anti-Dsg3 antibodies, suggesting that the remaining anti-Dsg1 antibodies present in the PNP sera were not pathogenic. The presence of mucosal lesions in our patient and the histological suprabasilar cleavage suggested the presence of pathogenic anti-Dsg3 antibodies. Such an antibody population could not be detected in the patients serum by immunoblotting. However, such anti-Dsg3 antibodies are not consistently detected in PNP sera by immunoblotting, whereas they are consistently detected by ELISA. Finally, the demonstration of anti-Dsg3 and/or anti-Dsg1 antibodies in sera from patients with PNP, PV and PF suggests a common mechanism of breakdown of immunological tolerance against desmogleins in these types of pemphigus, and confirms the role of antiDsg3 and possibly of anti-Dsg1 antibodies in the pathogenesis of PNP.

Interactions between epiplakin and intermediate filaments

Epiplakin, a cytoskeletal linker protein, was originally identified as an autoantigen in a serum specimen obtained from a patient with subepidermal blistering disease. To examine the binding ability of epiplakin with intermediate filaments (IF), we performed slot‐blot assays using fusion proteins that included various domains and subdomains of epiplakin. At least two of the 4.6 copies in the B domains of epiplakin were necessary for the binding of fusion proteins to keratin. The repeated structures of linker domains also played an important role in the binding of epiplakin to keratin in these assays while also increasing the repeated structure in the linker domain of epiplakin which is involved in the increased binding to IF. A similar but weaker binding to vimentin and desmin was also detected. These observations indicated that the highly repeated structures of epiplakin in both the B and the linker domains, which is the unique feature of this molecule in the plakin family, play an essential role in the functioning of this molecule.

Successful treatment by double‐filtration plasmapheresis of a patient with bullous pemphigoid: effects in vivo on transcripts of several genes for chemokines and cytokines in peripheral blood mononuclear cells

Summary The involvement of various cytokines and chemokines has been reported in the pathogenesis of bullous pemphigoid (BP). Double‐filtration plasmapheresis (DFPP) is an effective treatment for BP but the mechanism of action remains unclear. Using semiquantitative reverse transcription–polymerase chain reaction, we examined levels of transcripts for various cytokines and chemokines in freshly isolated peripheral blood mononuclear cells in a patient with BP before and after DFPP treatment. DFPP was performed four times. Relative levels of transcripts for interleukin (IL)‐8, macrophage inflammatory protein (MIP)‐1α and IL‐5, and the ratio of relative levels of transcripts for IL‐4 and interferon (IFN)‐γ, were higher, before treatment, than in healthy controls, and decreased when the extent of the lesions was reduced. Relative levels of transcripts for tumour necrosis factor (TNF)‐α and IL‐4 also decreased with regression of lesions, although they were similar to or lower than the corresponding levels in healthy individuals. When eruptions recurred, relative levels of transcripts for IL‐8, MIP‐1α, RANTES (regulated upon activation normal T cell expressed and secreted), IL‐2, IFN‐γ and TNF‐α were very much higher than those prior to the recurrence, while relative levels of mRNAs for IL‐4 and IL‐5 did not increase. Relative levels of transcripts for IL‐8, MIP‐1α, TNF‐α and IL‐2 were lower at the end of each individual DFPP and after the four treatments than at the beginning of treatment. Our observations suggest that cytokines and chemokines produced in mononuclear cells play important roles in the pathogenesis of BP and that regulation of their expression might be involved in the therapeutic effects of DFPP in BP.

A New Bullous Pemphigoid Antigen

Previously we reported a case of subepidermal blistering disease, which resembled bullous pemphigoid clinically. An immunoblot analysis revealed that the patients serum reacted exclusively with a 450-kD epidermal polypeptide. The 450-kD antigen was also expressed in human keratinocytes and a human squamous carcinoma (HO 1-u-1 or A431) cell line. Using the patients serum, cDNA clones in a bacteriophage lambda gt11 library derived from human foreskin keratinocytes were immune-screened. One positive clone was isolated from 2 million clones. Northern blot analysis showed that the inserted part of this clone hybridized with a more than 12-kb mRNA species derived from HO 1-u-1 cells. The deduced amino acid sequence had no homology with any other human component.