Acridine orange-DNA complex in actinic keratosis.

Electron microscopy processing and staining of nuclei were used to localize reaction products of acridine orange staining in actinic keratosis of human skin. Electron-dense granules about 10-100 nm in diameter were seen exclusively in the euchromatin portion of the nucleus. Almost all tumor cells had granules (mean = 65; SD = 26). These granules were also occasionally observed in the dermal connective tissue cells in the lesion. However, the mean number of 10 granules seen in these cells was definitely less than that of tumor cells. Normal skin controls did not have granules except occasionally in the basal cells of the epidermis.

Colloid-amyloid bodies in PUVA-treated human psoriatic patients.

In 4 cases of PUVA-treated psoriatic patients a number of colloid bodies and amyloid-like deposits were seen at the dermo-epidermal junction, in a very similar fashion the colloid bodies and the amyloid substances are deposited in lichen planus and in primary cutaneous anyloidoses respectively. In some instances such a deposition was found within the epidermis and contained remnants of cellular debris. Serial sections revealed that a part of the intraepidermal deposit is located in the dermis. Some deposition is partially composed of typical amyloid filaments and partially of nonamyloidal filaments. The latter were identical to those found in colloid or Civatte bodies of lichen planus and other conditions. Follow-up biopsies done in 1 patient several months after the cessation of the treatment still showed these substances. It was suggested that the amyloid-like substances are the product of special degeneration (apoptosis) of epidermal cells as colloid substances: Initally these cells undergo filamentous or colloid degeneration and they drop off into the dermis; where some of the characteristics of filaments are modified and connective tissue elements are added to produce such filamentous and amorphous components as seen in the amyloid island, whereas others remain as colloid bodies. Absorption or elimination of these substances seem to be extremely slow.

Histogenesis of primary localized cutaneous amyloidosis: sequential change of epidermal keratinocytes to amyloid via filamentous degeneration.

Two cases of lichen amyloidosus and 8 cases of macular amyloidosis were examined by electron microscopy. Epidermal keratinocytes showed variable degrees of focal degeneration in the basal or lower Malpighian layer. The primary change was seen in cells which contain fibrillar (30 nm in thickness) cytoplasmic inclusion. The following developments seemed to lead to filamentous degeneration (colloid bodies): (1) aggregation of tonofilaments within the granular or fibrillar cytoplasm, (2) filamentous cells, which are composed of bundles of 7-nm thick filaments surrounded by cell membrane and desmosomes, and (3) filamentous masses composed of bundles or whorls of tightly packed 7-nm thick filaments in the intercellular spaces. At the dermo-epidermal junction, some of the filamentous masses were surrounded by the basal lamina of the epidermis and others were dropping into the dermis. Occasionally, loosened filaments (similar to amyloid filaments) were about to drop into the dermis. Early formation of amyloid islands consisted of electron-dense and electron-light parts. They were located directly beneath the epidermis. In the deeper postion of the papillary dermis and in the upper reticular dermis, the majority were electron-light masses. Electron dense parts were the densely packed 7-nm thick filaments, whereas electron-light parts were the typical straight amyloid filaments. Small tubular filaments were seen in common in the filamentous cells, filamentous masses, and amyloid islands. It is concluded that some of the amyloid substance in primary localized forms of cutaneous amyloidoses derive from the epidermal cells through filamentous degeneration.

Intranuclear tonofilaments in verruca vulgaris.

Electron microscopic examination of a verruca vulgaris has revealed the intranuclear inclusion of tonofibrils. Although many intracellular inclusions may be related directly to the virus, nonviral inclusions may also be present.

Biologic changes due to long-wave ultraviolet irradiation on human skin: ultrastructural study.

Alteration of the skin induced by single and repeated long-wave ultraviolet (UVA) exposures was studied. Following a single exposure to relatively large doses of UVA, pronounced dermal damage was observed. In the papillary dermis, superficial dermal vessels showed widely open endothelial gaps and extravasation of blood cells. Marked changes of fibroblasts were also seen in the superficial dermis. In the reticular dermis, extravascular fibrin deposition was seen. After repeated exposures to UVA the formation of cross-banded filamentous aggregations ("Zebra bodies") was observed in the superficial and reticular dermis. These were often found in amorphous masses surrounding the blood vessels. These striking dermal alterations were absent in skin irradiated by solar stimulating radiation and in control skin. Dyskeratotic "sunburn cells" were occasionally seen in the epidermis after single as well as repeated exposures to UVA. The number of these cells was less than that seen after a single exposure to solar simulating radiation.

The specialized junctions between Merkel cell and neurite: an electron microscopic study.

Longitudinal serial sections of one half of the entire sinus hair of a mouse were examined by the electron microscope. Three neurites entering the outer root sheath from the perifollicular blood sinus were encountered. These were separate nerve trunks from those connected with perifollicular tactile nerve endings and exclusively innervated intrafollicular Merkel cells. Two types of specialized junctions were observed at the contact regions between Merkel cell plasma membrane and neurite plasma membrane: (i) desmosome-like structures in which small clear vesicles and/or the large cored vesicles of neurite and thicker membrane (post-synaptic?) of apposed Merkel cell were found ant (ii) synapse-like structures in which Merkel cell granules were concentrated near the plasma membrane and the membrane of the apposed neurite was usually thicker (post-synaptic?). In some of the synapse-like junctions the limiting membrane of Merkel cell granules fused with the Merkel cell plasma membrane and its content seemed to be discharged into the intercellular space. This suggested actual exocytotic secretion of Merkel cell granules. Juxtaposition of 2 types of junctions, i.e. (i) and (ii) above, was also found. This suggested the possibility that the reciprocal synapse would be present between Merkel cells and afferent neurites.

Presence of basal lamina-like substance with anchoring fibrils within the amyloid deposits of primary localized cutaneous amyloidosis.

The dermal-epidermal (DE) junction areas of skin specimens obtained from 16 patients with either lichen amyloidosis or macular amyloidosis were studied. In the dermal papillae where amyloid was deposited, elastic fibers frequently were absent, but periodic acid-Schiff reaction after diastase digestion was homogenously positive. Ultrastructural studies revealed that a basal lamina-like substance with anchoring fibrils was present between and within amyloid deposits. By indirect immunofluorescence technique using an anti-basement membrane zone antiserum obtained from a patient with bullous pemphigoid, specific linear fluorescence occurred at the DE junction, and in a reticular pattern in dermal papillae. It seemed that apoptotic keratinocytes of the epidermis brought down basal lamina and fine fibrous components attached to it when these cells dropped down to the papillary dermis and became the source of amyloid. These findings support the hypothesis that epidermal keratinocyte degeneration plays an important role in the histogenesis of cutaneous amyloidoses.

The ultrastructural findings of Charcot-Leyden crystals in stroma of mastocytoma.

Charcot-Leyden crystals (CLCs) have been found in many conditions associated with eosinophilia, but their occurrence in skin diseases is very rare. We report ultrastructural observations on the presence of CLCs in the cutaneous lesions of two cases of mastocytoma. Electron microscopy documented CLCs located in phagosomes of morphologically activated macrophages as well as free CLCs in the stromal tissue, close association between CLCs formation and damaged and lysed eosinophils was present. These findings provided evidence that the formation of CLCs in mastocytoma implicated the individual and interrelated biology of mast cells, eosinophils and macrophages. Phagosomes probably acted as the site of CLCs formation. The clinic and pathologic role of CLCs in mastocytoma deserves further investigation.

The ultrastructural characteristics of eccrine sweat glands in a Fabry disease patient with hypohidrosis.

As an investigation of the pathogenetic mechanism of diminished sweating in Fabry disease, an electron microscopy ultrastructural study was conducted on specimens of eccrine sweat glands from a typical patient with Fabry disease who had hypohidrosis, a low skin moisture content, and diminished thermoregulation ability. Numerous characteristic cytoplasmic inclusions were observed in the eccrine sweat glands, the lamellar pattern of which was considerably variable in various types of gland cells. Large vacuolar inclusions predominated in clear cells of secretory coil; lesser vacuoles were also seen in the coiled duct, and the basal cells of the straight duct toward the coiled duct displayed mulberry-like figures. There were some clear cells showing cell damage and necrosis in the secretory coil. Lamellated inclusions were noted in the unmyelinated axons innervating the eccrine sweat glands. The small blood vessels around the eccrine glands were narrowed by swollen endothelial cells with heavy inclusions. These intracytoplasmic deposits may be responsible for the decreased sweating ability in Fabry disease. The factors related to hypohidrosis are also discussed.

Expression of granulocyte colony-stimulating factor and its receptor in epithelial skin tumors.

Granulocyte colony-stimulating factor receptors (G-CSFR) have been observed on the surface of not only hematopoietic cells but also several cancer cells. In the present study, we investigated the expression of G-CSFR or G-CSF in epithelial skin tumors by immunohistochemical staining. The assessments were defined by the percentage of G-CSFR or G-CSF positive cells and expressed as G-CSFR and G-CSF scores. The G-CSFR score in SCC (77.6+/-20.0%) was significantly higher than that in Bowen's disease (BD) (51.0+/-35.6%), actinic keratosis (AK) (49.3+/-34.6%) or normal skin (30.0+/-32.1%) (P=0.0004, P=0.0003, P<0.0001, respectively). The mean G-CSF score in SCC (56.7+/-27.4%) or in BD (44.1+/-31.4%) was higher than that in normal skin (24.9+/-25.8%) (P=0.0075, P<0.001, respectively). G-CSF expression in AK (29.8+/-31.2%) was lower than that in SCC (P=0.0037). There was significant positive correlation between the G-CSFR score and the G-CSF score (gamma=0.274, P=0.0107) in skin tumors. These findings suggested that the assessment of G-CSFR expression might be associated with carcinogenesis of skin tumors.

Nylon brush macular amyloidosis.

Long-term use of a nylon brush for back scratching by a 53-year-old white woman was associated with the development of typical macular amyloidosis. EKH4 monoclonal antikeratin antibody, which recognizes 50-kd neutral and acidic keratin species, labeled this amyloid. Confirmation of amyloid substance in the lesion included positive staining with Dylon and thioflavin T; immunohistochemical reactions with monoclonal and polyclonal antibodies against elastic fiber microfibrils (NKH1 and anti-P component), immunoglobulins (IgG, IgM, and IgA), and complement (C3); and electron microscopic identification of 6- to 10-nm straight filaments. Type IV collagen staining demonstrated a breakage and/or thickening of the dermoepidermal basement membrane above the amyloid deposition in the papillary dermis. Electron microscopic findings confirmed this phenomenon.

Psoralen-UVA-treated psoriatic lesions. Ultrastructural changes.

Psoralen-ultraviolet light (PUVA)-treated psoriatic lesions were studied for ultrastructural changes. In early stages of treatment, sunburn cells in the epidermis and bizarre giant cells in the dermis were more frequently observed. When clinical improvement was apparent, these changes had subsided. Dermal abnormality in long-term therapy consisted of a thick perivascular coat of amorphous substance. No abnormality was found in the epidermal keratinocytes in long-term therapy, except a clustering and giant cell formation of melanocytes, a heavy melanization of keratinocytes, and hyperkeratosis. Low-dose initiation and slow increment of both 8-methoxypsoralen and UVA is probably a reasonable regimen for benign dermatoses such as psoriasis because it will allow enough time for the skin to become more protected, while the therapeutic results are as satisfactory as in a high-dose schedule.

Photodynamic therapy based on combined use of 5-aminolevulinic acid with a pheophorbide-a derivative for murine tumors.

BACKGROUND: 5-Aminolevulinic acid (5-ALA)-based photodynamic therapy (PDT) appears to be a promising cancer treatment modality. Here, we investigated whether enhancement of 5-ALA-PDT by combining another photosensitizer, a pheophorbide-a derivative (PH-1126), is an option. MATERIALS AND METHODS: PH-1126 (2.5, 5 or 10 mg/kg.bw) and 5-ALA (168 mg/kg.bw) were injected i.p. into C3H/HeN mice bearing squamous cell carcinoma (SCC) or BALB/c nude mice bearing L5178Y lymphoma. Afterwards, these mice received laser irradiations (630 nm for 5-ALA and 650 nm for PH-1126) with a total dose of 88 J/cm2. The results showed that PDT with 5-ALA plus PH-1126 at a low dose (2.5 mg/kg.bw) were well tolerated by both animal models, with resultant synergistically enhanced inhibition of tumor growth and/or survival advantage for the treated animals. CONCLUSION: This study demonstrated the usefulness of the combination of a low dose PH-1126 with 5-ALA for PDT of experimental tumors in vivo.

Clonal evolution and progression of 20-methylcholanthrene-induced squamous cell carcinoma of mouse epidermis as revealed by DNA instability and other malignancy markers.

We examined the clonal evolution of skin malignant lesions by repeated topical applications of 20-methylcholanthrene (20-MC) to the skin, which induces hyperplastic epidermis, papillomatous lesion and invasive carcinoma in mice. The lesions were examined histologically and immunohistochemically with anti-single-stranded DNA after acid hydrolysis (DNA-instability test), p53, VEGF, DFF45, PCNA and AgNORs parameters analyses. Multiple clones with increased DNA instability comparable to that of invasive carcinoma were noted in early-stage (2-6 weeks) hyperplastic epidermis, and their number increased in middle (7-11 weeks), and late-stages (12-25 weeks) of hyperplastic epidermis, indicating that they belong to the malignancy category. All papillomatous lesions and invasive carcinomas showed a positive DNA-instability test. Positive immunostaining for various biomarkers and AgNORs parameters appeared in clones with a positive DNA-instability test in early-or middle-stage hyperplastic epidermis, and markedly increased in late-stage hyperplastic epidermis, papillomatous lesions and invasive carcinomas. The percentage of PCNA-positive vascular endothelial cells was significantly higher in VEGF-positive lesions with a positive DNA-instability test and became higher toward the late-stage of progression. Cut-woundings were made to papillomatous and invasive carcinoma lesions, and the regeneration activity of vascular endothelial cells was determined by using flash labeling with tritiated thymidine (3H-TdR). In small papillomatous lesions, vascular endothelial cells showed regenerative response, but the response was weak in large lesions. No such response was noted in invasive carcinomas; rather, cut-wounding induced collapse of blood vessels, which in turn induced massive coagulative necrosis of cancer cells. These responses can be interpreted to reflect exhausted vascular growth activity due to excessive stimulation by VEGF-overexpression, which was persistently seen from hyperplastic epidermis to invasive carcinoma.

Comparative histochemical study of Bowen's disease and actinic keratosis: preserved normal basal cells in Bowen's disease.

The degree of DNA-instability as revealed by immunohistochemical staining with anti-cytidine antibody after acid hydrolysis (DNA-instability test) has been recently used as a marker of malignancy. This technique was applied to examine 17 skin tissue samples of Bowen's disease, 47 of actinic keratosis, 15 of squamous cell carcinoma, 5 of seborrheic keratosis, and 10 of normal skin. All benign neoplastic cells of seborrheic keratosis and normal epidermal cells were negative. On the other hand, all cancer cells were positive with the DNA-instability test, indicating their malignancy, but all basal cells in Bowen's disease were completely negative. Compatible with this result, the basal cells in Bowen's disease were characteristically normal as evident in other histochemical examinations. Thus, they were negative with p53 immunohistochemistry, with normal signals of chromosome 17 in situ hybridisation and argyrophilic nucleolar organiser region, and showed slightly enhanced proliferative activity as revealed by proliferating cell nuclear antigen immunohistochemistry. Immunohistochemical staining with 34 beta E12 (monoclonal antibody against cytokeratins 1, 5, 10, and 14), which stains all normal epidermal keratinocytes including basal cells, showed that only the basal cells of Bowen's disease stained strongly and homogeneously, while all cancer cells in the upper layers of Bowen's disease and all layers of actinic keratosis were only sporadically or weakly stained. Staining with 34 beta B4 (monoclonal antibody against cytokeratin 1), which recognises the whole epidermis except for the basal layer in the normal epidermis, showed that the basal cells in the Bowen's disease were completely negative, and lower layer cells in the actinic keratosis and upper layer cells in Bowen's disease were only sporadically stained positive, although the superficial layer cells in actinic keratosis stained strongly and homogeneously. Our findings clearly indicate that the basal cells in Bowen's disease are normal. In support of this conclusion, the same cells showed normal morphology on electron microscopy with preserved basement membrane, although the latter was often damaged in actinic keratosis.

Eruptive steatocystoma multiplex on the scalp.

We report a 71-year-old Japanese woman who noticed approximately 120 small yellowish cysts on the scalp from nine months before her first visit. An incision made into the cysts discharged oily yellow material. Biopsy of a lesion disclosed that the cysts had typical features of steatocystoma multiplex histologically. This case was a unique eruptive variety of steatocystoma multiplex on the scalp.

Localized form of multiple glomus tumors: report of the first case showing partial involution.

We recently examined a boy with relatively large multiple glomus tumors on the left scapular region. Histologic examination revealed a typical non-capsulated glomangioma in the middle and deep dermis; by electron microscopic examination, the tumor cells were seen to exhibit a characteristic smooth muscle cell structure. Thermographic examination revealed the higher temperature of the tumor. Within a year, the tumor showed a partial involution. We think this is the first report of multiple glomus tumor showing a partial involution.

A case of epidermotropic metastatic mammary carcinoma.

A special type of metastatic carcinoma to the skin, epidermotropic carcinoma, is thought to be a parasitic occupation of malignant tumors in the epidermis. A patient, a 56-year-old Japanese woman who had had a mastectomy because of mammary carcinoma thirteen months earlier, developed a metastatic skin tumor in the operation wound and another area of mammary skin. Histologically, almost only the epidermis was occupied by multifocal metastatic tumor nests, so the tumor was a type of epidermotropic metastatic carcinoma. Moreover, some tumor nests were found to be eliminated from the epidermis. Like other types of metastatic skin tumor, nevus pigmentosus, and amyloidosis, epidermotropic metastatic carcinoma can show transepithelial elimination.

Papillary eccrine adenoma with a pomegranate-like appearance.

Papillary eccrine adenoma (PEA) is a rare cutaneous tumor which histopathologically presents numerous intradermal tubular structures with inward papillary projections. Only a few cases of PEA have been reported recently. We report a case of PEA of a 58-year-old Japanese man. The marked hyperkeratosis and pits gave the tumor the clinical appearance of a burst-open pomegranate. Compact hyperkeratosis within proliferated epidermis contained spiral ducts mimicking intraepidermal eccrine sweat ducts histopathologically. These keratinous structures were thought to correspond to the pores. Several tubular structures running up to the overlying thickened epidermis were observed in the upper dermis. With these findings and with immunohistochemical studies, we proposed that this tumor originated from eccrine sweat ducts.

Adenoid cystic carcinoma of the skin--an immunohistochemical and ultrastructural study.

Adenoid cystic carcinoma of the skin was studied. Histologically, tumor cells were arranged in a tubular and a cribriform pattern, mainly int he reticular dermis. Immunohistochemically, epithelial membrane antigen was reactive with the tumor cells, but S-100 protein, vimentin, and carcino-embryonic antigen were not. On electron microscopy, we confirmed the findings of previous reports; tumor cells were arranged to form luminal structures; most of them were pseudolumina containing fine mucin granules, basal laminae, and collagen fibers, but some were true lumina with numerous microvilli and junctional complexes. New findings of this study were bizarre-shaped, electron-dense, net-like structures within the true lumina which were considered to be a type of mucin.