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Genetic differences are involved in the development of lupus erythematosus (LE). Skin lesions are influenced by environmental triggers such as ultraviolet light, temperature, and chemical stresses, and the patterns of skin lesion are variable in cutaneous LE such as systemic LE (SLE), chronic discoid LE (CDLE), subacute cutaneous LE (SCLE), and LE tumidus (LET). Although there are a few conflicting reports, many Japanese dermatologists feel there are photosensitivity differences in lupus erythematosus between Asian and Caucasian subjects with SCLE and LET. HLA studies in Japanese subjects revealed that HLA-DRB1*1501 association was with both CDLE and SLE. The association between HLA-Cw6 and CDLE was first reported in Japanese population, and a HLA-A33-B44-DRB1*1302 haplotype showed a positive association in CDLE. However, these results are not compatible with those from Caucasian subjects. There are no significant associations among HLA studies, photosensitivity, and anti-Ro/SS-A antibodies in Japanese CLE patients. Photosensitivity will be a key factor to dissolve multi-factorial complexes of LE etiopathogenesis. Our present understanding is that an axis of photosensitivity, anti-Ro/SS-A antibodies and apoptosis via TNF are the best (markers) to verify the contribution of genetics in SCLE, LET, and other CLEs. The incidence and photosensitivity of SCLE and LET are much lower in Japanese than in Caucasian subjects. However, this discrepancy may open the window for investigating CLE pathogenesis through global collaborations. For this purpose and goal, a new and more conventional method should be developed for the examination of so-called photosensitivity.

Neuropeptides mediate stress-induced cuta- neous inflammation such as atopic dermatitis. The ef- fect of substance P on proliferation and cytokine mRNA expression of peripheral blood mononuclear cells in re - sponse to Dermatophagoides farinae (Der f) was stud- ied in atopic dermatitis patients with positive RAST scores to Der f. Upon stimulation with Der f peripheral blood mononuclear cells from patients proliferated in a B7-dependent (CD80- and CD86-dependent) man- ner, while those from the patients with negative scores, nonatopic eczematous dermatitis patients or normal individuals, did not. Based on the reactivity of normal individuals, atopic dermatitis patients with a stimula- tion index greater than 1.8 were tentatively defined as high responders, who comprised two-thirds of the pa- tients. Proliferation in high responders was associated with upregulation of IL-2 mRNA expression and in- duction of IL-5 mRNA expression. Substance P at 10 -10 to 10 -8 M promoted the Der f-induced prolifera- tion when added at the start of culture and upregu- lated IL-10 mRNA expression while downregulating IL-5 mRNA expression. Our results suggest that sub- stance P modifies immune responses of atopic T cells to Der f by promoting proliferation and altering cy- tokine profiles, and thus modulates the clinical mani- festations of atopic dermatitis.

The effects of FK506, a new immunosuppressive agent, on the development of lupus dermatoses were investigated in the autoimmune-prone MRL/Mp-lpr/lpr (MRL/lpr) mouse, which is an animal model for the spontaneous development of skin lesions similar to those of human lupus erythematosus (LE). FK506 reduced the incidence of skin lesions, lupus nephritis, the titre of serum anti-double-stranded DNA antibodies and the massive T cell proliferation. The incidence and magnitude of IgG deposition at the dermoepidermal junction were not changed. These results suggest that FK506 is a promising immunosuppressive agent for the control of autoimmune skin diseases.

The distribution of lipocortin I, a steroid-induced inhibitory protein of phospholipase A2, was examined in normal and psoriatic human skin. Using immunoblotting analysis with specific antibody against human lipocortin I purified from human placenta, lipocortin I was detected as a 37 kDa protein in cultured epidermal cells, whole skin and epidermis. In the dermis and stratum corneum, lipocortin I was only weakly detectable by Western blotting. In contrast to normal skin, much less lipocortin I was detected by Western blotting analysis in psoriatic skin. Using immunoperoxidase immunohistochemical analysis, lipocortin I was demonstrated in the cytoplasm of keratinocytes in the upper and middle layers of the epidermis and in some infiltrating cells in the dermis in normal skin. In involved psoriatic skin, by contrast, lipocortin I was almost undetectable in the epidermis, although it was demonstrated in some infiltrating cells in the dermis. No immunostaining of lipocortin I was observed in the stratum corneum of normal or psoriatic skin. These results, together with the finding that phospholipase A2 activity is higher in psoriatic epidermis than in normal epidermis, suggest that lipocortin I plays an important role in the regulation of differentiation and proliferation of epidermal keratinocytes.

Polyarteritis nodosa (PN) is a classical collagen disease with poor prognosis that demonstrates systemic necrotizing vasculitis of small and medium-sized arteries. Cutaneous symptoms are observed in 25-60% of PN patients. On other hand, cutaneous polyarteritis nodosa (CPN) is designated for the cutaneous limited form of PN and demonstrates benign prognosis. However, there has been much debate on whether or not CPN can progress to PN. Although CPN lesions are fundamentally limited to skin, some CPN cases show extracutaneous symptoms such as peripheral neuropathy and myalgia. According to PN diagnostic criteria, which were established by the Ministry of Health, Labour and Welfare of Japan, a disease with both cutaneous and at least one extracutaneous symptom with appropriate histopathological findings can be diagnosed as PN. The same is true according to diagnostic criteria established by the American College of Rheumatology. In addition, there are no specific diagnostic criteria for CPN. In this study, CPN cases were retrospectively collected from multiple Japanese clinics, and analyzed for detailed clinical and histopathological manifestations, in order to redefine the clinical entity of CPN and to propose appropriate diagnostic criteria for CPN and PN. According to the CPN description in Rook's Textbook of Dermatology, we collected 22 cases with appropriate histopathological findings. Of the 22 cases, none progressed to PN or death during the follow-up period, 32% had peripheral neuropathy and 27% had myalgia. Regarding extracutaneous symptoms with CPN, 17 dermatological specialists in vasculitis sustained the opinion that CPN can be accompanied by peripheral neuropathy and myalgia but these symptoms are limited to the same area as skin lesions. Based on these results, we devised new drafts for CPN and PN diagnostic criteria. Our study shows the efficacy of these criteria and most dermatologists recognized that our new diagnostic criteria for CPN and PN are appropriate at the present time. In conclusion, this study suggests that CPN does not progress to PN, and introduces new drafts for CPN and PN diagnostic criteria.

The structure and function of the epidermis is maintained by cell renewal based on epidermal turnover. Epidermal turnover is delayed by aging, and it is thought that the delay of the epidermal turnover is a cause of aging alternation of skin. The epidermal turnover is related to the energy metabolism of epidermal basal cells. Adenosine 5′-triphosphate (ATP) is needed for cell renewal: cell division, and adenosine 5′-monophosphate (AMP) increases the amount of intracellular ATP. These findings suggest that AMP accelerates the epidermal turnover delayed by aging. This study investigated whether AMP and adenosine 5′-monophosphate disodium salt (AMP2Na) accelerates the epidermal turnover. An effect of AMP2Na on cell proliferation was examined by our counting of keratinocytes. An effect of AMP2Na on cell cycle was examined by our counting of basal cells in DNA synthetic period of hairless rats. The effects of AMP2Na (or AMP) on the epidermal turnover were examined by our measuring stratum corneum transit time by use of guinea pigs, and by our measuring stratum corneum surface area by use of hairless rats and in a clinical pharmacological study. The AMP2Na showed two different profiles on the proliferation of primary cultured keratinocytes. At a low concentration it induced cell growth, whereas at a high concentration it inhibited cell growth. The number of basal cells in the DNA synthetic period of AMP2Na was significantly higher than that of the vehicle in hairless rats. The stratum corneum transit time of AMP2Na was significantly shorter than that of the vehicle in guinea pigs. The corneocyte surface area of emulsion containing AMP2Na was significantly smaller than that of the vehicle in volunteers. We conclude that AMP promotes the cell proliferation and the cell cycle progression of epidermal basal cells and accelerates epidermal turnover safely. In addition, AMP is useful for skin rejuvenation in dermatology and aesthetic dermatology.

Trichloroacetic acid (TCA) is one of the most widely used peeling agents, and induces full necrosis of the whole epidermis, followed by reconstitution of the epidermis and the matrix of the papillary dermis. The cytotoxic effects of TCA, such as suppressing proliferation of keratinocytes and fibroblasts and protein synthesis by fibroblasts, have already been reported. However, the entire biological mechanism responsible for TCA peeling has yet to be determined. Hypothetical activation effects of TCA treatment on epidermal cells to induce production of growth factors and cytokines are examined, and are compared with its cytotoxic effects in terms of time course and applied TCA concentrations. After various periods of incubation with TCA, viability of Pam212 murine keratinocytes was investigated with MTT assay and dye exclusion assay, and production of growth factors and cytokines with reverse transcription-polymerase chain reaction (RT-PCR). Changes in platelet-derived growth factor (PDGF)-B mRNA expression and protein production in the human skin specimens after TCA application were then examined by RT-PCR and immunohistochemistry, respectively. Incubation with TCA showed cytotoxicity and induced death of Pam212 cells, depending on the incubation period and the TCA concentration. In addition, expressions of PDGF-B, tumor growth factor (TGF)-alpha, TGF- beta1 and vascular endothelial growth factor, which are the growth factors reportedly secreted from keratinocytes during wound healing, were all detected in Pam212 cells after short-term treatment with TCA. Expressions of inflammatory cytokines such as interleukin (IL)-1 and IL-10 were also induced. In TCA-treated NIH-3T3 fibroblasts, in contrast, observed was upregulation of only keratinocyte growth factor, which is reportedly secreted from fibroblasts, as well as the similar cytotoxic effect. In human skin, PDGF-B mRNA expression became significantly upregulated after TCA application, and then immediately downregulated. Immunoreactive PDGF-B in the cytoplasm of keratinocytes became detectable throughout the epidermis after TCA application, reached maximum after the peak of mRNA expression, and then declined significantly over 24 h when the epidermis became completely necrotic. The TCA-treated epidermis acts as a major source of growth factors, including PDGF-B, before undergoing full necrosis. This effect might contribute to a promotion of re-epithelialization and dermal regeneration without wound contraction and scarring.

We investigated the in vitro responses to bacterial superantigens of peripheral blood mononuclear cells taken from patients with psoriasis (one arthropathic, two guttate and four chronic plaque type). We also analysed the relationship between the magnitude of the responses of peripheral blood mononuclear cells to bacterial exotoxins and the number of circulating T cells bearing V beta 2 and V beta 3 regions. The proliferative response of peripheral blood mononuclear cells to Staphylococcal enterotoxin B and toxic shock syndrome toxin-1 was significantly higher in patients with active psoriasis than in normal subjects. An improvement in skin eruption was associated with a decrease in the lymphocyte response to one-half or one-third that of the active phase. There was no significant difference between patients with psoriasis and normal subjects in the percentage of V beta 2- and V beta 3-positive circulating T cells. The percentages of V beta 2-positive and V beta 3-positive cells were not correlated with the levels of responsiveness to toxic shock syndrome toxin-1 and to Staphylococcal enterotoxin B, respectively. These findings suggest that the magnitude of responses of peripheral blood mononuclear cells to bacterial toxins does not depend on the number of T cells reactive with the relevant superantigen, but depends on the extent of skin lesions in psoriasis.

Ceramide and sphingosine have been suggested to be intracellular modulators of cell growth and differentiation. The effects of these sphingolipids on the growth and differentiation of keratinocytes were examined using cultured human keratinocytes (the squamous cell carcinoma cell line, DJM-1). The synthetic short-chain cell-permeant analogues of ceramides, N-acetylsphingosine, N-hexanoylsphingosine and N-octanoylsphingosine, significantly promoted differentiation as confirmed by upregulation of cornified envelope formation, synthesis of involucrin and increased transglutaminase activity, and inhibited proliferation as shown by a reduction in cell numbers, DNA amount and thymidine incorporation. Generally, these activities were greater the longer the N-acyl carbon chain. On the other hand, sphingosine at an appropriate concentration modestly stimulated the proliferation of cultured cells. Our results suggest the possibility that the growth and differentiation of keratinocytes are at least partially regulated by ceramide and sphingosine.

The deposition of immunoglobulin (Ig) at the dermo-epidermal junction (DEJ) of the skin, frequently observed in autoimmune mouse strains, is similar to that seen in patients with systemic lupus erythematosus (SLE). MRL/Mp-lpr/lpr (MRL/lpr) mice have an autosomal recessive mutant gene, lpr, which produces massive T-cell proliferation and accelerates the onset of autoimmune diseases. MRL/Mp- +/+ (MRL/n) mice lack the lpr gene, and do not develop autoimmune disease during the first year after birth under pathogen-free conditions. To verify the mechanisms of subepidermal Ig deposition in the skin of LE, we designed an experiment in which we could induce Ig deposition in the control MRL/n mice. Intraperitoneal injection of lymphoproliferative cells of aged MRL/lpr mice induced splenomegaly and splenic granulomatous angitis in the control MRL/n mice. Lipopolysaccharide (LPS), a polyclonal B-cell activator, induced slight splenomegaly and relatively high levels of serum Ig. Dermatopathological investigation revealed mild lymphocyte infiltration without positive Ig deposition at the DEJ of MRL/n mice treated with proliferative T cells. Injection of both proliferative T cells and LPS induced 50% positivity of subepidermal Ig deposition, and high levels of serum immunoglobulins and anti-double stranded DNA (anti-dsDNA) antibodies. These changes were not observed in MRL/n mice injected with thymocytes of newborn MRL/lpr mice. Skin lesions and lupus nephritis were not demonstrated in any of the mice tested. This study suggest that both the mild inflammatory reaction and the presence of anti-dsDNA antibodies are required for the induction Ig deposition at the DEJ in the skin of LE patients.

Expression of proliferating cell nuclear antigen/cyclin (PCNA/cyclin) in skin tissue specimens and cultured keratinocytes was studied using a monospecific antibody, obtained from a patient with systemic lupus erythematosus, and a monoclonal antibody. Indirect immunofluorescent staining revealed that cultured keratinocytes obtained from human foreskins expressed PCNA/cyclin as variable nuclear patterns in 15–30% of the cells. In normal human skin tissue specimens, PCNA/cyclin was demonstrated in only a few basal cells. Interestingly, PCNA/cyclin was expressed strongly in almost all the cells of the lowest layer of the epidermis adjacent to squamous cell carcinomas, whereas the tumor aggregates themselves had no positive staining. In contrast, no such characteristic staining was demonstrated in specimens of basal cell carcinoma. The staining pattern of PCNA/cyclin was different from that of Ki-67 in the skin tissue specimens. Our results suggest that PCNA/cyclin could be a useful marker of cell proliferation.

Expression of E (epithelia) and P (placental) cadherin cell adhesion molecules was examined immunohistochemically using human developing skin. In adult skin, E-cadherin was expressed on cell surfaces of whole epidermal layers including skin appendages, whereas P-cadherin was expressed only on those of basal layers and the outer layers of skin appendages, which was consistent with the compartment of proliferating cells. In fetal skin, while the patterns of E- and P-cadherin expression were generally similar to those in the adult, P-cadherin temporarily showed a unique spatiotemporal expression pattern in developing sweat ducts. During this stage, the expression of P-cadherin accumulated in the epidermal ridges and showed a discrepancy with the compartment of proliferating cells. These results suggest that the expression of P-cadherin is spatiotemporally controlled, and may be closely related to the segregation of basal layers as well as to the arrangement of epidermal cells into eccrine sweat ducts, but is not closely related to cell proliferation.

We examined the time-dependent dynamics of epidermal Langerhans' cells (LC) in human systemic lupus erythematosus (SLE), in MRL/Mp-lpr/lpr (MRL/lpr) mice, and in various experimental cutaneous inflammations, such as the Arthus reaction, dinitrochlorobenzene allergic dermatitis, and croton oil primary irritant dermatitis, in order to clarify the pathomechanisms of lupus skin lesions. The numbers of LC in untreated SLE patients with newly developed skin lesions decreased in the central lesional sites and increased significantly in the peripheral lesional sites. In MRL/lpr mice, the number of LC increased significantly in the central lesional sites during the initial stage and increased in the peripheral lesional sites and decreased in the central lesional sites 2–4 weeks after the onset of skin lesions. In contrast, with experimental cutaneous inflammations of guinea pigs, the increase in numbers of LC in the peripheral lesional sites was not significant during the time course of the reaction. These results suggest that the increased numbers of LC during the active and early stages of skin lesions in human SLE and MRL/lpr mice are closely related to the specific spontaneous development of skin lesions, unlike the dynamics of LC in experimental cutaneous inflammations.

In the sera of patients with lepromatous leprosy anti-DNP antibodies were detected in order to determine the mode of polyclonal B-cell activation. Anti-DNP antibodies were found in 30% of the patients with active lepromatous leprosy and in 8% of those with inactive lepromatous leprosy. The level of anti-DNP antibodies in active patients was significantly higher than the level in inactive patients and control. However, the presence of anti-DNP antibodies was unrelated to the production of circulating immune complexes and antinuclear antibodies. These results suggest that polyclonal B-cell activation might occur but that the B-cell clones stimulated by M. leprae are different from patient to patient.

The skin of New Zealand, MRL and BXSB mice was immunohistopathologically examined in order to study the appearance of skin immunoglobulin (Ig) deposition and its correlation with the occurrence of anti-single-stranded (ss) DNA antibodies in sera. Our studies revealed Ig deposition at the dermoepidermal junction (DEJ) in non-lesional skin and a significant age-related correlation between skin Ig deposition and serum anti-ssDNA antibodies. However, immunofluorescent study of autoimmune mice using anti-ultraviolet-irradiated DNA antiserum failed to demonstrate DNA antigens at the DEJ.

The skin of New Zealand, MRL and BXSB mice was immunohistopathologically examined in order to study the appearance of skin immunologublin (Ig) deposition and its correlation with the occurrence of anti-single-stranded (ss) DNA antibodies in sera. Our studies revealed Ig deposition at the dermoepidermal junction (DEJ) in non-lesional skin and a significant age-related correlation between skin Ig deposition and serum anti-ssDNA antibodies. However, immunofluorescent study of autoimmune mice using anti-ultraviolet-irradiated DNA antiserum failed to demonstrate DNA antigens at the DEJ.

The MRL-lpr/lpr(MRL/l) mouse is a new animal model for human systemic lupus erythematosus (SLE) and skin lesions with hair loss and scab formation are one of the characteristic manifestations in this mouse. We investigated the histopathology of the skin lesions in MRL/l mice and studied the related autoimmune phenomenon. Light microscopical observations revealed hyperkeratosis, acanthosis, hypergranulosis, liquefaction, vasodilation in the dermis and T-cell infiltration into the dermis at the age of 5 months (mo). Immunohistological studies showed the presence of immunoglobulins and/or complement depositions at the dermal-epidermal junction (DEJ). In some mice there was deposition of immunoglobulin at the DEJ at 2 mo and in 90%-100% of MRL/l mice at over 5 mo. Temporal relationship was present among cutaneous immunoglobulin depositions, the occurrence of anti-DNA antibodies and proteinuria. These findings suggest that MRL/l mice might provide a new aid for studying the biological mechanisms of the development of skin lesions in human SLE.

Of several murine autoimmune models, MRL/Mp-lpr/lpr (MRL/l) mice are the most attractive from a dermatopathological point of view, because their skin lesions resemble the erythematous lesions of systemic lupus erythematosus (SLE). In order to clarify the pathogenesis of lupus dermatoses, histamine and its metabolizing activities in the skin of MRL/l mice were investigated. The specific activities of histamine-N-methyltransferase (HMT) in the skin of MRL/l mice were significantly lower than those in the skin of control mice i.e., MRL/Mp-+/+ (MRL/n), C57BL/6J, and BALB/c mice. In the dorsal lesional skin of MRL/l mice, HMT activities were markedly lower than those in normal abdominal skin. In addition, age-related analysis of HMT levels in the dorsal skin of MRL/l mice revealed that HMT activities reached their maximum at the age of 2 or 3 months and then decreased at 4 or 5 months when skin manifestation appeared: however, HMT activities in the abdominal skin increased almost linearly with age. There were no significant differences in histamine content in these mice, and diamine-oxidase activities were not detected in any skin specimens. From these results, it is suggested that impaired histamine metabolism is a particular biochemical feature of the skin of MRL/l mice.

Using a sensitive and modified solid-phase radioimmunoassay for detecting anti-cardiolipin antibodies, sera of 45 patients with lepromatous leprosy were examined. Nine of the 45 (20%) showed positive levels of anti-cardiolipin antibodies. Inhibition tests revealed that these antibodies significantly cross-reacted with double-stranded (ds) DNA, but not with single-stranded (ss) DNA or extractable nuclear antigens (ENA). We describe the unique pattern of antibody cross-reactivity with cardiolipin and dsDNA in sera of patients with lepromatous leprosy.

MRL/Mp-lpr/lpr (MRL/l) and MRL/Mp-+/+ mice were irradiated with middle-wavelength ultraviolet light (UVB), and the development of skin lesions, skin lupus band test (LBT), anti-DNA antibodies in sera, and the extent of glomerulonephritis of the kidney were examined. Long-term exposure to low doses of UVB irradiation accelerated the development of skin lesions and enhanced the intensity of the positive skin lupus band in MRL/l mice. The contents of anti-DNA antibodies in sera, the incidence of positive findings in LBT, and the extent of glomerulonephritis were not influenced by the UVB irradiation. The promotion of the development of the skin lesions in MRL/l mice by the UVB exposure was not considered to be associated with acceleration of systemic autoimmune phenomena.

The MRL-lpr/lpr (MRL/1) mouse, a new animal model for the study of human systemic lupus erythematosus (SLE), shows characteristic skin manifestations in addition to several systemic autoimmune phenomena. The ultrastructural changes observed in the dermoepidermal junction (DEJ) and in the uppermost dermis were: (1) infolding of the DEJ; (2) deformities of the basal lamina, — partial disappearance, thickening, hanging down, duplication, and separation from the basal cell membrane; (3) basal laminalike dense material in the uppermost dermis and increased anchoring fibrils; (4) particles composed of circulated half-desmosomes between the basal cells and the basal lamina, and in the uppermost dermis with or without an enclosing basal lamina; (5) cell processes of the basal cells; and (6) invagination of the basal lamina in the basal cells. Most of these findings were similar to the ultrastructural changes observed in the skin lesions of human SLE. The skin eruptions of MRL/l mice might be a new aid in the investigation of the pathogenesis of the skin lesions of human SLE.

Regulation of the distribution of concanavalin A (Con A)/receptor complexes by the cytoskeletal contracture system was studied in guinea pig epidermal cells in suspension and culture using the fluorescence double staining method. After treatment with 100 micrograms/ml of Con A at 37 degrees C for 30 min lectin/receptor complexes were endocytosed by the less-differentiated cells in suspension and by the adherent cells in 1- and 3-day cultures that represent a growing cell fraction. The same treatment resulted in diffuse surface distribution of the complexes in the well-differentiated cells in suspension. Colchicine (10(-5) and 10(-6) M) inhibited internalization of the complexes with resultant diffuse distribution in 60% of the adherent cells in culture. Cytochalasin B (5 and 10 micrograms/ml) not only inhibited endocytosis but promoted formation of surface patchy clumps of the complexes in suspended, less-differentiated cells and cultured adherent cells. The distribution profile was not influenced by these drug treatments in the well-differentiated cells. SDS polyacrylamide gel electrophoresis and autoradiography of 125I-labelled epidermal membranes revealed several Con A-reactive polypeptides common to the cells at various differentiation steps. The progressive decrease in endocytosis and mobility of Con A/receptor complexes was suggested to occur with differentiation. In the germinative cells the distribution of lectin/receptor complexes seemed to be regulated by microfilaments and microtubules.

In 79 patients with leprosy a significant increase of anti-extractable nuclear antigen (ENA) antibodies and circulating immune complexes (CIC) was found. No correlation between CIC and anti-ENA antibodies was demonstrable. Since such a correlation is known from antinuclear antibodies and CIC in patients with systemic lupus erythematosus, it appears likely that anti-ENA antibodies do not play a causative role in CIC-mediated pathogenesis of leprosy.