Natsuko Okada

Separation of the epidermal sheet by dispase

Dispase is a bacterial neutral protease which is obtained from the culture filtrate of Bacillus polymyxa, After 24‐h treatment of the human skin with 500 and 1000 U/ml dispase, the epidermal sheet was easily peeled from the dermis, and its undersurface retained rete ridges. Electron microscopic observation showed that the basal surface was composed of cells with numerous slender villi and cytoplasmic projections. Although the intercellular spaces of the spinous as well as the basal layers were wide, all desmosomes were intact with their accompanying tonofilaments. An epidermal cell suspension was obtained by incubating the dispase‐separated epidermal sheet in trypsin solution for 15 min and the viability of the cells was more than 95% as measured by the trypan blue dye exclusion test. A culture of keratinocytes was established by seeding the dissociated cells in the usual way.

Skin pigmentation associated with minocycline therapy

A patient on long‐term minocycline therapy developed blue‐black discoloration on the legs. Skin biopsy specimens from the pigmented areas were examined by light and electron microscopy and energy‐dispersive X‐ray microanalysis. Pigmented granules were present at all levels of the dermis and subcutaneous fat tissues. Ultrastructural examination showed electron‐dense graular material within the cytoplasm of dermal macrophages and energy‐dispersive X‐ray microanalysis indicated that the granules contained iron. Thyroid tissue obtained by aspiration biopsy showed the presence of fine brown granules within the cytoplasm of the follicular epithelial cells. An extract of skin from the pigmented areas was subjected to high performance liquid chromatography and minocycline was detected.

Epstein-Barr virus—associated large granular lymphocyte leukemia with cutaneous infiltration

A 36-year-old man had a high titer of antibody to Epstein-Barr virus (EBV) and recurrent necrotizing papules and nodules on his face and oral mucosa. The disease was diagnosed as CD3+4- 8+ large granular lymphocyte leukemia of T-cell origin. Southern blot analysis demonstrated that EBV DNA was present in CD8+ lymphocytes; EBV antigens were also observed in these lymphocytes. These findings demonstrated that EBV latently infected the leukemic cells and may have played a role in the pathogenesis of this disease. This is the first report of an association between EBV and large granular lymphocyte leukemia of T-cell origin.

A case of calcinosis universalis successfully treated with low-dose warfarin.

A 27‐year‐old male patient with calcinosis universalis resulting from dermatomyositis was successfully treated with low‐dose warfarin. On his trunk and extremities, there were many subcutaneous calcified nodules, and knee flexion was difficult. After oral warfarin therapy for three years, the calcified nodules became smaller, and the knee mobility improved. His serum vitamin K level was abnormally high, decreased just after starting warfarin therapy, and then remained within the normal range. Since vitamin K has been known to play an important role in the Ca2+ binding process in bones or tissues, we suggest that this therapy is effective in reducing subcutaneous calcification through the vitamin K cycle.

Bone Mineral Density of the Lumbar Spine in Psoriatic Patients with Long Term Etretinate Therapy

Etretinate has been known to produce a variety of skeletal manifestations when administered for prolonged periods. To study whether osteoporotic changes occurred during the treatment, bone mineral density (BMD) was measured by dual energy X‐ray absorptiometry in 13 psoriatic patients who received long term etretinate therapy (average 3.7 yrs). The values of BMD of the lumbar spine in these patients were decreased from those of age‐matched controls (92.3 ± 9.7%, p<0.01), while those in psoriatic patients who received topical corticosteroid applications alone showed no significant decrease. Our findings suggest that long term etretinate intake induces an increased risk of osteoporotic changes.

Salivary Gland MALT Lymphoma Associated with Helicobacter pylori Infection in a Patient with Sjögren's Syndrome

We report a case of salivary gland MALT lymphoma in Sjögrens syndrome associated with localized H. pylori infection. A 76–year‐old woman had a history of bilateral cheek masses for two years. Histologically, the parotid glands were invaded by numerous centrocyte‐like cells to form lymphoepithelial lesions. The tumor cells showed immunohistological differentiation into B cells. Southern blotting demonstrated immunoglobulin gene rearrangement. These results indicated that the tumors were MALT lymphoma. H. pylori, as assessed by the urease test (CLO test; BML Ltd., Tokyo, Japan), was positive in the tumor specimen. After wide local excision of the tumors followed by radio therapy and oral administration of antibiotics and proton pump inhibitor, no evidence of recurrence was found during the 24–months of follow up. H. pylori infection in the salivary gland is rare, although the source of infection and transmission of H. pylori organisms has been thought to be the oral cavity. We discussed the association between H. pylori infection and salivary gland MALT lymphoma. The microorganism may play a role as an additional antigenic stimulus for the development of salivary gland MALT lymphoma as well as for the development of gastric MALT lymphoma. This means that H. pylori can play a role in lymphoma progression as booster of B cell lymphoproliferation.

Characterization of pigmented granules in minocycline-induced cutaneous pigmentation: observations using fluorescence microscopy and high-performance liquid chromatography

We describe a technique to facilitate histopathological detection and quantitative measurement of trace amounts of tissue minocycline.

Isolation of a viable eccrine sweat gland by dispase

SummaryDispase was used to obtain viable eccrine sweat glands from human skin in an intact shape. The full thickness of human skin was soaked in a solution of dispase in Eagles minimum essential medium at a concentration of 500 units/ml and kept in a refrigerator at 4°C for 24h. The epidermal sheet with its appendages could then be easily separated from the dermis by lifting the epidermis with fine forceps. Electronmicroscopic observation revealed that the eccrine sweat gland was completely separated from the dermis at the basement membrane zone. The isolated epidermal sheet was scarcely dissociated by mechanical agitation in the presence of Ca2+ and Mg2+ ions. The eccrine sweat gland was cut away from the epidermis by using microscissors under a stereomicroscope. A cell suspension of the isolated eccrine sweat glands was obtained after trypsinization. The cells remained more than 90% viable up to 48 h in the culture medium. The obtained viable eccrine sweat glands will be useful for the study of the biology of sweating.

Metabolism of an aromatic retinoid Ro 10-9359 by cultured human epidermal keratinocytes

SummaryAn aromatic retinoid Ro 10-9359 is metabolized after absorption from intestine to form Ro 10-1670 which is an active therapeutic compound. The epidermal keratinocytes, a main target tissue of retinoid therapy in dermatology, was examined in the capacity to metabolize the retinoid. The culture of human epidermal keratinocytes was treated with 10-6 M Ro 10-9359 and the metabolites released in the medium was analyzed by HPLC. The HPLC profile showed a distinct peak of Ro 10-1670. The human skin fibroblasts, HeLa cells, Chang liver cells and 3T3 cells were less active in metabolizing Ro 10-9359, and only a small amount of Ro 10-1670 was detected in the culture of human skin fibroblasts treated with 10-6 M Ro 10-9359 for 2 days. When these cells were disrupted by a glass homogenizer, and incubated with Ro 10-9359, no Ro 10-1670 formation was detected.

Cutis laxa with ultrastructural abnormalities of elastic fiber

A case of a congenital, autosomal recessive form of generalized cutis laxa is reported. The patient was a 27-month-old boy with generalized flaccid skin and short stature. Radiologic examination revealed that the age of the bones of the wrist was compatible with a chronologic age of only 1 year. Elastic fibers were diminished throughout the dermis, and results of electron microscopic study showed globular and unstained elastin and relatively large amounts of the microfibrillar components of elastic fibers.