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The morphology of mast cells in the skin lesion from a case of urticaria pigmentosa was observed using light and electron microscopy. Furthermore, the IgE deposition on mast cells in the lesion was studied with immunoelectron microscopy. Diaminobenzidine reaction and 4-chloro-naphthol reaction were used to detect endogenous peroxidase activity in mast cells in the lesion. Ultracytochemical techniques were applied to clarify the localization of peroxidase activity in the mast cells. Morphologically, neither blast-like cells nor intermediate cells between basophils and mast cells were found in the lesion. No mast cells in the lesion did exhibit IgE deposition on their plasma membrane. By light microscopy, granular stains of diaminobenzidine reaction and 4-chloro-naphthol reaction were observed in the cytoplasm of mast cells in the lesion. By electron microscopy, a finely stippled peroxidase staining was observed in some granules of the mast cells. Perinuclear cisterna and endoplasmic reticulum appeared devoid of enzymatic activity. These findings suggest that the infiltrating mast cells in this case were immature.

近年,黒色真菌感染例に対して局所温熱療法が用いられ,有効な結果が得られているが,その作用機序については未だ明確にされていない.我々は,病原性黒色真菌5種25株について,その発育に及ぼす温度の影響を観察するとともに,懐炉圧抵時の表面皮膚温および深部温を測定し,自験7例のクロモミコーシスの病理組織標本における菌要素の存在部位を比較検討した.その結果,1)上限発育温度はFonsecaea pedrosoi 39℃, Exophiala jeanselmei 36～38℃, Exophiala dermatitidis 40～42℃(ただし顆粒形は38℃),Phialophora verrucosa 37℃, Cladosporium trichoides 45℃以上であり,41℃の環境下にて,F.pedrosoiま25日目,E.jeanselmeiは5日目,P.verrucosaは5日目に死滅していたが,E. dermatitidis, C. trichoidesは25日後も死滅しなかった.2)懐炉貼布にて皮表42℃のとき深さ3～4mmの部位は40.5℃まで上昇した.3)局面型を呈する病変組織内において,黒色真菌の菌要素は皮表から1.5mm,炎症性細胞浸潤は4mm以内に認められた.以上の結果よりクロモミコーシスにおいては,原因菌がF. pedrosoi, E. jeanselmei, P. verrucosaであれば局所温熱療法が有効であることが示されたが,温熱効果には菌に対する直接効果以外にも宿主側の要因も考えられるので,更に検討が望まれる.

Using complement immunofluorescence, the capability of complement fixation by bound antibodies in pemphigus lesional skin was compared with circulating intercellular antibodies of complement-fixing type in the same patient. Five lesional skin samples from 6 patients, containing bound IgG, gave positive binding of complement (C3) supplied from fresh guinea pig serum, while only one serum sample showed complement-fixing pemphigus antibodies in the circulation. These results support the view that complement-fixing pemphigus antibodies are rapidly localized to the skin and play a role on the production of acantholysis.

Normal human skin was sliced into five horizontal layers and distribution of type I and type III collagens, glycosaminoglycans, and non-collagenous glycoprotein(s) among the five layers was analyzed. No remarkable differences in the relative contents of type I and type III collagens and noncollagenous glycoprotein(s) were detected among the five layers. However, glycosaminoglycan content was higher in the upper layers than in the lower and the ratio of hyaluronic acid to dermatan sulfate was also higher in the upper layers. These results are compared and discussed with our previous data on calf skin.

Complement-fixing intercellular antibodies were detected in 10 of 17 sera from untreated pemphigus patients. The role of complement in the organ culture system was investigated using these sera. Ten sera possessing complement-fixing intercellular antibodies showed IgG binding to the intercellular substance in the organ-cultured skin and acantholysis-like changes were observed in eight cases. C3 deposition was not seen in any case. However, after treatment of the sections of cultured skin with fresh normal human serum, complement fixation of the intercellular substance by bound IgG was revealed in all the ten cases. No significant differences in the grade of acantholysis-like changes between the complement-depleted system and the complement-supplied system were observed. Complement does not appear to be necessary in the acantholytic process in the in vitro organ culture system, even though we considered the presence of complement-fixing intercellular antibodies.

23歳男性のchromoblastomycosisより分離されたExophiala jeanselmeiの絨毛形と穎粒形をE. jeanselmeiの保存株8株およびE. dermatitidisの保存株7株 (うち1株は顆粒形) と, 形態学的あるいは発育温度の面から, さらには生理学的, 血清学的に比較検討した.その結果, 本菌株の絨毛形はE. jeanselmeiとして矛盾するところがなかった.また, 本菌株の顆粒形はE. dermatitidisの顆粒形とは集落の外観, 顕微鏡的所見, 血清学的所見に若干の差違が見出された.また, E. jeanselmeiとE. dermatitidisとの比較検討の結果, 両者の分生子形成細胞に環紋 (annellation) が認められ, ともにExophiala属としてその近縁性は示唆されるものの, 集落の形態, 発育温度等が異なり, 鑑別可能と思われるので, 現時点では両者を独立した菌種とする見解に従うべきと思われた.

We report a case of angiosarcoma and dermal melanocytosis occurring simultaneously in the same lesion. We examined the primary and 2 metastatic lesions. Histopathologically, the anaplastic angiosarcoma cells had a tendency to form irregularly shaped small cavities. Immunohistochemically, they were strongly reactive with Ulex europaeus agglutinin (UEAI) stain. The mid and deep dermis of the same lesion had spindle or elongated slender melanocytes containing melanin granules. The melanocytes were positive with S-100 protein stain. Ultrastructurally, pinocytotic vesicles, fine filaments, and Weibel-Palade body-like dense granules were observed in the cytoplasm of angiosarcoma cells. Dermal melanocytes had external lamina and melanosomes in various stages. The melanocytes showed no similarity to the neoplastic tumor cells and there was no apparent intermediate form between the 2 kinds of cells. The etiological implications of dermal melanocytosis with a tumor of vascular origin are discussed.

DNA synthesis of WF-1 fibroblasts derived from a patient with Werner's syndrome was stimulated by fetal calf serum and adult human serum but not by various mitogens including epidermal growth factor, platelet-derived growth factor (PDGF), fibroblast growth factor, insulin and 12-O- tetradecanoylphorbol-13-acetate (TPA). To clarify the cause of nonrespon-siveness to these mitogens, we compared the rate of protein phosphorylation in normal fibroblasts HF-O and Werner's WF-1 cells. PDGF and TPA enhanced the phosphorylation of a Mr 80 K protein, which is known to be a substrate for protein kinase C, both in HF-O and WF-1 cells. This indicates that the pathway involving PDGF receptor, phosphatidylinositol turnover and protein kinase C activation is operational in WF-1 cells. Several species of phosphoproteins of Mr 250 K, 135 K, 110 K, 78 K and 42 K were detected in normal HF-O cells by immunoprecipitation using an anti-phosphotyrosine antibody. The same species of phosphoproteins were detected in Werner's WF-1 cells at passage 6, but only when treated with various mitogens and were not detected in WF-1 cells at passage 10 even after the PDGF- or TPA-treatment. These results sug-gest that the reduction of phosphorylation of these target proteins may be in part responsible for the diminished mitogenic responsiveness of Werner's fibroblasts.

Biochemical analysis of involved and uninvolved skin of a 16-year-old female with acrogeria showed that hyaluronic acid and collagen contents were decreased only in involved skin. Explant cultures from both involved and uninvolved skin synthesized mainly hyaluronic acid in similar amounts. Since the glycosaminoglycans and hydroxyproline excreted in the urine were not increased, we speculate that a localized rather than a systemic abnormality may be present in acrogeria. Decreased collagen and hyaluronic acid contents in the patient are discussed in relation to Werner’s syndrome and type IV Ehlers-Danlos syndrome.

ポビドンヨード (Povidone-iodine, PVP-I) の病原性酵母に対する抗菌効果を検討した結果, PVP-Iは7種のCandida, Torulopsis glabrata, Cryptococcus neoformansの保存株およびC. albicansの新鮮分離株10株に対してほぼ等しい殺菌効果を示した. 血清の混入により効力が減少する傾向があるが, 有効濃度においては1～3分以内に殺菌作用が発現することが示された. またC. albicansによつて汚染された器具や皮膚もPVP-Iによつて強力かつ迅速に消毒されることが示された.

Sporothrix schenckii の菌糸形および酵母形の菌体および培養濾液より蛋白質を抽出後, SDS電気泳動にて分離し, 各菌形における蛋白質を比較検討した. その結果, 酵母形菌体に特異的な分子量15万前後の蛋白質が認められた. この蛋白質はアミノ酸分析の結果, グリシン, グルタミン酸, セリン, アラニン等を多く含むことがわかつた. また, 酵母形および菌糸形の培養濾液中にそれぞれ特異的な蛋白質がいくつか認められた. 以上の結果より, これらの蛋白質と S. schenchii の二形性との関連性につき若干の考察を加えた.

The tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA) induces DNA synthesis in quiescent 3T3-L1 cells but not in its variant VT-1 cells. A lambda gt10 cDNA library was constructed using poly(A)+ RNA from 3T3-L1 cells that were stimulated by TPA for 20 min. Radioactive cDNA probes were prepared from mRNAs of TPA-treated 3T3-L1 and VT-1 cells and used for screening of the 3T3-L1 cDNA library by differential hybridization. Nine of 6000 phase plaques hybridized only to the 3T3-L1 cDNA probe. Analysis of the nucleotide sequence of five of these clones indicated a high degree of homology with human or mouse type I and type III collagen genes. Three other independent clones showed no homology with any known DNA sequences. These isolated clones of TPA-inducible early (TIE) genes may be useful to study the signal transduction pathway of phorbol esters.

A 40-year-old Japanese male with diabetic scleredema was reported. Histochemical examination revealed that the alcian blue and colloidal iron strains were positive in the dermis, especially in the lower dermis. The biochemical analyses of the 3 horizontally sliced layers of scleredema skin revealed that (1) glycosaminoglycan content was low in the upper and middle dermis, which was chiefly due to the decrease in hyaluronic acid, (2) dermatan sulfate content was constant among the 3 layers, (3) the content of collagen in the upper and middle dermis was higher, and (4) the ratio of type I/type III collagen in the whole involved skin was normal. From these results, we speculate that the increase in collagen content in the upper and middle dermis may be essential for the pathophysiology of the disease.

Summary Skin biopsy was performed in five cases of pemphigus; direct immunofluorescence (IF) studies demonstrated intercellularly bound IgG and C3 in each case. Sections of lesional skin, containing in vivo bound IgG were, in two cases, capable of further binding C3in vitro from normal human serum, an effect most marked in relation to areas of acantholysis. This work provides additional evidence that the complement system is involved in the process of pemphigus acantholysis.

Prurigo pigmentosa is an inflammatory dermatosis characterized by pruritic, reddish, papular lesions and gross reticular pigmentation that occurs mainly on the trunk. Nearly 100 cases have been reported in Japan to date. We describe a patient with this condition who responded well to dapsone. An electron microscopic study of the reddish papular lesion showed marked intercellular edema and evidence of cellular injury of the basal cells. Direct immunofluorescence was negative. We stress that this condition is a distinct clinical entity, histologically characterized by a lichenoid tissue reaction as proposed by Pinkus, although it is little known outside Japan.

To solve the problems of the multiplication and invasion of Candida albicans in the alimentary tract, germ-free mice were orally inoculated with C. albicans and the antagonistic effect of superinfection with Escherichia coli was examined. C. albicans could easily be established in the alimentary tract of germ-free mice by inoculation with less than 10 organisms, whereas in the alimentary tract of the SPF mice, having normal bacterial flora, C. albicans could not establish even after inoculation with an overwhelming dose. In germ-free mice large numbers of the inoculated Candida were excreted in the feces throughout the observation period of 130 days, without affecting the conditions of the mice. By histopathological examination of these mice only one mouse showed microabscesses with leukocytic infiltration accompanied by hyperkeratosis and acanthosis in the epithelium of the forestomach. However even in this mouse no invasion of Candida cells into the acanthotic squamous cell layer was seen. After inoculation of the mice who had already E. coli in their gut as a monocontaminant with C. albicans, or even after inoculation of E. coli to the mice harboring Candida as a monocontaminant, E. coli always outnumbered C. albicans. In the former case Candida were even completely eliminated from the mice within few days. Thus, it appeared that in the alimentary tract of the mice, E. coli have the capacity to antagonize to C. albicans.

There was no macroscopic or microscopic ulcer found in the stomach of ICR germ-free mice even when they were treated with relatively large doses of cortisone. When the germ-free mice were orally inoculated with C. albicans combined with subcutaneous administration of cortisone acetate with the doses of more than 5mg, shallow ulcer developed at the site of the junction between forestomach and glandularstomach in the majority of the cases. In the base of ulcer, candida showed not only hyphal form but also other various forms. Polymorphonuclear leukocytic infiltration with candida organisms was observed at the site of the junction of the forestomach and glandularstomach shortly after oral inoculation with C. albicans. It was considered that the development of ulcer was secondary to the affection of candida. These results indicate that the lowering of the host resistance play an important role on the developement of systemic candida infection.

1. A case of 33-year-old house wife with Senear-Usher syndrome was reported.2. Antiepithelial antibody in the sera of the patient was demonstrated by indirect IF staining technique. Bound IgG was present in the intercellular space of the prickle cell layer of the skin.3. IgG was significantly high in the cantharidin blister fluid induced from uninvolved skin of the patient while IgA and IgM remained slightly higher levels.4. Several immunological tests for lupus erythematosus gave negative results.5. These data indicate that the Senear-Usher syndrome should be classified as a form of pemphigus. However, more cases should be further accumulated and immunologically investigated as an aid to clarify the relationship between lupus erythematosus and this syndrome and to elucidate the pathogenesis of the syndrome.

Paraneoplastic pemphigus (PNP) is an autoimmune blistering disease that occurs in association with underlying neoplasms. PNP patients develop characteristic autoantibodies directed against multiple antigens, mostly identified as members of the plakin family of cytoplasmic proteins (desmoplakin I and II, bullous pemphigoid antigen I, envoplakin, and periplakin). HD1/plectin, another member of the plakin family, has not previously been detected in the characteristic PNP antigen complex, which may relate to practical difficulties associated with its large size (molecular weight approximately 500 kDa). In this study, a combination of immunoprecipitation and immunoblot is used to demonstrate that HD1/plectin is also recognized by sera from PNP patients. Thirteen of 16 PNP sera tested were positive for HD1/plectin compared with none of 43 control sera (11 pemphigus vulgaris, 11 pemphigus foliaceus, 11 bullous pemphigoid, and 10 normal individuals). Combined with our recent finding that desmoglein 3 and desmoglein 1 are cell surface target antigens in PNP, this demonstration of plectin/HD1 as another component of the antigen complex in PNP confirms that PNP is an autoimmune disease against desmoglein and plakin family molecules.

Pemphigus vulgaris is an autoimmune blistering disease caused by autoantibodies against desmoglein 3, a member of the desmosomal cadherin family. These autoantibodies recognize conformation-dependent epitopes on desmoglein 3. In this study we attempted to map the conformational epitopes of desmoglein 3 in pemphigus vulgaris using recombinant desmoglein 3 produced by the baculovirus expression system. We developed a series of domain-swapped molecules between desmoglein 3 and desmoglein 1, which have similar structures but distinct epitopes. These were developed by substituting deleted segmental regions of desmoglein 3 by the corresponding desmoglein 1. Thus domain-swapped molecules containing desmoglein 3 residues 1-403, 1-161, 163-566, and 405-566 were constructed and used as competitors for competition enzyme-linked immunosorbent assay against the entire extracellular domain of desmoglein 3 with 25 pemphigus vulgaris sera. Considering more than 50% absorption as significant, residues 1-403 and 1-161 showed significant absorption in 24 out of 25 (96%) and 18 out of 25 (72%) pemphigus vulgaris sera, respectively, whereas only one serum and no sera showed significant absorption by residues 163-566 and 405-566, respectively. Furthermore, no apparent change in their major epitopes was seen during the time course in four pemphigus vulgaris cases tested. These findings indicate that the domain-swapping approach is useful for conformational epitope mapping in pemphigus and that amino-terminal residues 1-161, which are considered to include a region essential for cell-cell adhesion in cadherins, contain the critical residues of the conformational epitope of desmoglein 3 recognized by the autoantibodies in pemphigus vulgaris sera.

Desmoglein 3 is the autoimmune target of pemphigus vulgaris. Most, if not all, pathogenic autoantibodies are raised against conformational epitopes on desmoglein 3. In this study, we examined whether posttranslational modification in endoplasmic reticulum is required for the proper three-dimensional structure formation of recombinant pemphigus vulgaris antigen. Previously, we have produced by baculovirus expression a secreted form of desmoglein 3, PVIg, which represents equivalent conformational epitopes of the native antigen and showed that PVIg is able to immunoadsorb heterogeneous autoantibodies from pemphigus vulgaris patients' sera. To elucidate the role of proteolytic processing, we constructed a mutant PVIg molecule, PVIg-fXa, whose putative endoproteolytic cleavage site was replaced by the recognition sequence of serum coagulation factor Xa. PVIg-fXa was produced without proteolytic processing; however, exogenous treatment of PVIg-fXa with factor Xa resulted in cleavage of the prosequence. Interestingly, not only the processed PVIg-fXa, but also the unprocessed form, showed the immunoadsorptive activity. Furthermore, to elucidate the role of endoplasmic reticulum signal peptide at the amino terminus, we constructed another mutant, PVIg-delta sig, which lacks the signal peptide and prosequence. PVIg-delta sig was not secreted and accumulated in the cytosol. PVIg-delta sig failed to show the immunoadsorption. Together with our previous finding on the role of glycosylation, these observations indicate that the conformational epitopes of the recombinant pemphigus antigen are not affected either by glycosylation or proteolytic processing, although they need to be formed in the endoplasmic reticulum, and emphasize the importance of conformation of the antigen in pathogenic autoantibody binding.

Pemphigus vulgaris is an autoimmune blistering disease of the skin and mucous membranes that is caused by anti-desmoglein 3 IgG autoantibodies. Recently, we generated an active disease mouse model for pemphigus vulgaris by adoptive transfer of splenocytes from immunized desmoglein 3 –/– mice to Rag2 –/– mice. In this study, we performed immunologic and histopathologic studies using this pemphigus vulgaris model in mice and compared the gross and microscopic phenotypes of pemphigus vulgaris model mice and desmoglein 3 –/– mice. Pemphigus vulgaris model mice showed strong in vivo IgG, and weak IgA deposition on keratinocyte cell surfaces in stratified squamous epithelia, and produced circulating anti-desmoglein 3 IgG antibodies without apparent cross-reactivity to desmoglein 1, in enzyme-linked immunosorbent assays. The predominant IgG subclass was IgG1. Pemphigus vulgaris model mice and desmoglein 3 –/– mice were almost indistinguishable in terms of both gross and microscopic findings. Both types of mice showed suprabasilar acantholysis in the stratified squamous epithelia, including the oral mucous membranes and traumatized skin around the snout or paws; however, some pemphigus vulgaris model mice demonstrated a more severe phenotype than desmoglein 3 –/– mice. The esophagus and forestomach were affected in some pemphigus vulgaris model mice, but not in desmoglein 3 –/– mice. Furthermore, eosinophilic spongiosis, which is found in early pemphigus vulgaris lesions in patients, was observed in pemphigus vulgaris model mice but not in desmoglein 3 –/– mice. Pemphigus vulgaris model mice reflect several of the histopathologic and immunologic features seen in pemphigus vulgaris patients, and provide a valuable tool to investigate the pathophysiologic mechanisms of pemphigus vulgaris.

Mutations within the gene encoding the anchoring fibril protein type VII collagen (COL7A1) have recently been established as the pathogenetic basis for the inherited blistering skin disorder, dystrophic epidermolysis bullosa. We report a patient with a moderately severe phenotype of recessive dystrophic epidermolysis bullosa. We report a patient with a moderately severe phenotype of recessive dystrophic epidermolysis bullosa, in whom COL7A1 mutations have been identified on both alleles. The patient is a 5-y-old Japanese male of nonconsanguineous parents, with clinical features including generalized trauma-induced blistering since birth, complete loss of nails, and partial fusion of the fingers and toes. Immunofluorescence microscopy examination of the dermal-epidermal junction in the patient's skin revealed near-normal intensity staining with an antitype VII collagen antibody (LH7:2). Transmission electron microscopy showed a reduced number of thin, poorly-formed anchoring fibrils. PCR amplification of genomic DNA, followed by heteroduplex analysis, and nucleotide sequencing demonstrated that the patient was a compound heterozygote for a nonsense mutation (E2858X) within the NC-2 domain of type VII collagen and a missense mutation (G2576R) within the type VII collagen triple helix. Both mutations were verified by restriction endonuclease digestion. Information about these mutations advances our understanding of genotype-phenotype correlations in dystrophic epidermolysis bullosa, and further delineates the mechanisms involved in dermal-epidermal dysadhesion.

We Investigated the antigen molecules for six clinically typical cases of paraneoplastic pemphigus (PNP) using immunofluorescence, immunoprecipitation, and immunoblotting. All the PNP sera showed a clear reactivity with transitional epithelia of rat urinary bladder and immunoprecipitated the 250-kD, 230- kD 210-kD, 190-kD, and 170-kD proteins hi various combinations, confirming the diagnosis of PNP. Immunoblot analysis demonstrated slightly different reactivity from that of immunoprecipitation. With immunoblotting of normal human epidermal extracts bovine desmosome preparation, and extract of cultured squamous cell carcinoma cells, all the PNP sera reacted with a characteristic doublet of the 210-kD and 190-kD proteins. However, inununoblotting detected the 250-kD desmoplakin I and the 230-kD bullous pemphigoid antigen less frequently and did not detect the 170-kD protein. Further immunoblot studies indicated that the 210-kD protein is different from desmoplakin II and that the 190-kD protein is most frequently detected by PNP sera, Two of the six PNP sera specifically reacted with the extracellular domain of recombinant pemphigus vulgaris antigen protein, indicating that pemphigus vulgaris antigen may be involved in PNP, In future studies to unravel the complex mechanisms of the PNP antigens, the immunoblot technique may be a useful tool.

Pemphigus vulgaris (PV) is an autoantibody-mediated bullous disease, but the role of natural killer (NK) cells in its pathogenic process has never been examined in detail. Circulating CD56 + CD3 − NK cells as well as CD69 + -activated NK cells were increased in PV patients compared with healthy controls and patients with other autoantibody-mediated autoimmune diseases, including immune thrombocytopenic purpura and myasthenia gravis. Gene expression analysis of highly purified NK cells demonstrated an increased expression of IL-10 and decreased expression of IL-12Rβ2 , perforin , and granzyme B ex vivo in PV patients versus healthy controls. The NK cells from PV patients also showed impaired signal transducer and activator of transduction4 phosphorylation upon in vitro IL-12 stimulation. Moreover, NK cells from PV patients exhibited reduced IL-10 production in response to in vitro stimulation with IL-2/IL-12. Finally, IL-5 expression in NK cells was exclusively detected ex vivo in PV patients with active disease, and was lost in subsequent analyses performed during disease remission. Together these findings suggest that NK cells contribute to a T helper type 2-biased immune response in PV patients through impaired IL-12 signaling and an upregulation of IL-10 and IL-5.

Desmoyokin, a high-molecular-weight protein of 680 kD with a 170-nm-long dumbbell shape, was originally thought to be localized to the desmosomal attachment plaque and to work as a kind of stabilizer of desmosomes. Recently, desmoyokin was shown to be widely detected in many types of cells that do not possess desmosomes. The purpose of the present study was to elucidate the precise localization and possible function of desmoyokin in human epidermis. In 0.2-micron ultrathin cryosections of human skin for immunofluorescence, anti-desmoyokin antibody showed a ladder-like staining pattern along the cell surface, whereas anti-desmocollin and anti-desmoplakin antibodies as controls showed a discontinuous dotted staining pattern, indicating their distinct localization. Post-embedding immunoelectron microscopy with cryofixation and cryosubstitution revealed that desmoyokin was localized mainly along the non-desmosomal and non-hemidesmosomal plasma membrane, but not to the desmosomes and hemidesmosomes themselves. This localization was further confirmed by double-labeling immunoelectron microscopy with antibodies against desmocollin, desmoplakin, or bullous pemphigoid antigen. Results indicate that desmoyokin was not localized to the desmosomes themselves as previously considered. Desmoyokin was localized to the non-desmosomal and non-hemidesmosomal epidermal keratinocyte cell surface as a plasma membrane-associated protein, and might play a role in cell adhesion that is not directly associated with desmosomes or hemidesmosomes.

By in vitro complement immunofluorescence, 6 sera from pemphigus with intercellular antibodies were tested for their capability to fix C1q, C4, C3, and properdin. All 6 serum samples yielded positive reaction for C3 staining. Three serum samples gave positive staining for C1q, 5 serum samples for C4, and 3 serum samples for properdin, respectively. Substitution of C2 deficient serum as a complement source inhibited C3 and properdin staining but not positive C1q and C4 staining. These results are best explained by the concept that complement activation in vitro by intercellular antibodies occurs via the classical pathway followed by assembly of the C3 amplification mechanism.

Skin tissue and explant culture were used to investigate glycosaminoglycan (GAG) metabolism in the atrophied skin of a patient treated with long-term oral corticosteroids. Total GAGs per dry weight were almost the same in the atrophied and control skin. Hyaluronic acid was increased 1.5 times above the control values. Dermatan sulfate decreased to 3% of the total GAGs in the atrophied skin; in the controls dermatan sulfate content was 30%. In the skin explant culture experiment, the synthetic activity of dermatan sulfate in the atrophied skin was higher than in the controls. These results suggest that the synthesis of hyaluronic acid and dermatan sulfate in atrophied skin is modulated by corticosteroid, which causes thinning, fragile skin.

There are four main subclasses of human IgG: IgG1, IgG2, IgG3, and IgG4, among which IgG1-IgG3 activate complement, but IgG4 does not. We studied the IgG subclasses of anti-intercellular (IC) antibodies in pemphigus patients and anti-basement membrane zone (BMZ) antibodies in bullous pemphigoid (BP) patients by immunofluorescent staining using mouse monoclonal antibodies against human IgG1-IgG4. At the same time, the capability of complement fixation of each serum was determined by means of complement immunofluorescence. In both pemphigus and BP autoantibodies, various distributions of IgG subclass were shown, but specific patterns were not observed. In BP, all of the complement fixing antibodies had at least one of IgG1-IgG3 subclasses, while noncomplement fixing antibodies only possessed IgG4. This result agreed well with the biologic characteristics of IgG subclasses in respect of complement fixing capability. On the contrary, in pemphigus, the circulating antibodies showed a distribution of IgG subclass which did not correlate with the biologic characteristics in terms of complement activation. This discrepancy may further dispute the roles of the complement system on the bulla formation in pemphigus.

Eight out of 12 serum samples from patients with bullous pemphigoid having basement membrane zone antibodies gave positive binding not only to the basement membrane zone but also to the basal cell membrane and/or cytoplasm as observed by complement immunofluorescence. Reaction of fluorescein labeled pemphigus vulgaris gamma-globulins, binding mainly to the lower intercellular spaces of the epidermis, was greatly reduced by the prior incubation of high-titered bullous pemphigoid sera having the reactivity to the basal cells, while that of fluorescein labeled pemphigus foliaceus gamma-globulins binding mainly to the upper and middle intercellular spaces, was not influenced by the prior application of these bullous pemphigoid sera. These results indicate that bullous pemphigoid antibodies are heterogenous and can be classified into 2 types and that some cross reaction is present between pemphigus antibodies and bullous pemphigoid antibodies having the reactivity to the basal cells.

Lesional skin specimens from scleroderma patients were sliced horizontally into three layers. The ratio of hyaluronic acid to dermatan sulfate in each layer was analyzed. The ratios in each layer of scleroderma skin were lower than those in the corresponding layers of normal skin and the lowest value was found in the bottom layer of scleroderma skin. The possible involvement of glycosaminoglycan metabolism in the pathophysiology of scleroderma is discussed.

• The serum of a patient with a typical case of pemphigus vulgaris contained complement-fixing intercellular autoantibodies (pemphigus antibodies). Complement-fixing pemphigus antibodies were demonstrated only during the untreated active stage of the disease and titrated lower than corresponding IgG autoantibodies, which paralleled well with the disease activity. Pemphigus lesional skin, which contained in-vivo-bound IgG, showed the capability of further binding C3 in vitro from normal human serum. It was suggested from these findings that the complement system may play an active role in pemphigus acantholysis through complement-fixing pemphigus antibodies. ( Arch Dermatol 114:1191-1192, 1978)

• An immunohistochemical and ultrastructural study of an atypical case of multiple non-X histiocytoma was done. There was involvement of the skin, lungs, and liver in a 3-month-old male infant. Microscopic examination of the cutaneous tumors revealed a dense infiltration of cells with polymorphous large nuclei and abundant eosin-ophilic cytoplasm in the entire dermis. Both immunohistochemical and electron microscopic studies suggested that the tumors were non-X histiocytomas. The patient's condition deteriorated with dyspnea due to rapid enlargement of tumor masses in the liver and lungs. However, at 5 months of age, the cutaneous nodules and pulmonary and hepatic lesions showed a tendency to involute. Furthermore, at 12 months of age, they were no longer detectable. The patient at 24 months of age was well with normal development. To date, no recurrence of the disease has been observed. (Arch Dermatol1988;124:1254-1257)

In order to elucidate the mechanism underlying deposition of the substance observed mainly in the intercellular space of parakeratotic psoriatic horny layer, 6 cases of psoriasis vulgaris and, in comparison with these, 3 cases of pemphigus vulgaris were examined by using direct immunofluorescence technique with or without pretreatment with acid citrate buffer (pH 3.0) or digestion by fibrinolysin. The substance in the intercellular space of parakeratotic psoriatic horny layer showed positive fluorescence for IgG, IgA, IgM, C3 and fibrinogen. Positive fluorescence for IgG and fibrinogen was not significantly affected by preincubation of tissue sections in acid citrate buffer, but almost completely disappeared by digestion with fibrinolysin. On the contrary, positive fluorescence for IgG in the intercellular space of the prickle cell layer of pemphigus skin lesions almost completely disappeared by acid citrate buffer, but was not significantly affected by fibrinolysin. From these findings, it was suggested that the deposition of the substance in the intercellular space of parakeratotic psoriatic horny layer is rather the result of coagulation of plasma protein than that of antigen-antibody reactions.

A little-known dermatosis, prurigo pigmentosa, is characterized by itchy reddish lesions and gross reticular pigmentation which occurs mainly on the trunk. Histologically, it is characterized by lichenoid tissue reaction. We present a 23-year-old male patient who showed typical eruption not only on the trunk but also on the forehead. Although the etiology remains to be explained, this unusual distribution of the eruption might help to know the real cause of this peculiar disorder.

We investigated the genetic expression of bullous pemphigoid (BP) antigen (230 kD) synthesized by Pam cells using an in vitro cell-free translation assay followed by immunoprecipitation. Prior to the study, we showed that (1) the 230 kD polypeptide does not undergo further processing after it is produced by cells in the short pulse experiment; (2) the 230 kD polypeptide is not degraded at least within the 18 h of the chase experiment; and (3) with tunicamycin treatment the underglycosylated BP antigen still interacts specifically with BP serum. The polypeptide of molecular size slightly greater than 230 kD was identified in the translated proteins directed by ribonucleic acid (RNA) isolated from Pam cells. The size of the mRNA was estimated to be approximately 34S–40S (7.7–10.9 kilobase) using the fractionation method with sucrose density gradient ultracentrifugation of RNA. These results indicate that bullous pemphigoid antigen (230 kD) is a primarily translated product, genetically expressed by Pam cells.

Using an immunoelectron microscopic technique we have compared the ultrastructural localization of IgG and complement (C3) in the basement membrane zone of three typical cases of bullous pemphigoid and three atypical variant cases (nodular, polymorphic and localized pemphigoid, respectively). There was no significant difference in the in vivo ultrastructural localization of these immunoreactants between the typical and the atypical cases of pemphigoid. In all cases IgG and C3 were detected between the basilar surface of the basal keratinocytes and the basal lamina. Moreover, the in vitro binding sites of circulating anti-basement membrane zone antibodies were identical in the case of nodular pemphigoid and a typical pemphigoid as control. We also observed that immunoreactants were not deposited beneath melanocytes.

• In a patient whose clinical features resembled subcorneal pustular dermatosis, IgA deposits in the intercellular space of the upper epidermis were found on direct immunofluorescence study. Furthermore, by indirect immunofluorescence, IgA autoantibodies against the same area of the epidermis were demonstrated in the patient's serum. ( Arch Dermatol 1987;123:1062-1065)

The capability of complement fixation of pemphigus antibodies was tested using combined in vitro complement immunofluorescent (IF) staining methods. Three sera out of 25 serum samples from 22 pemphigus patients revealed positive reactions, while all other sera gave negative results. Specificity control tests confirmed the positive reactions to be specific for complement staining. Complement fixing pemphigus antibodies were titrated lower than corresponding IgG antibodies and were demonstrable only in the extensive stage of the disease. Thus, the present work supplied evidence that pemphigus antibodies fix complement in vitro. However, the discrepancy still remains between the in vivo deposition of complement in most cases of pemphigus and in vitro capability of complement fixation in only few cases. More investigations should be needed to explain the exact role of complement in pemphigus acantholysis.