Iwao Takiuchi

Isolation of an extracellular proteinase (keratinase) from Microsporum canis.

Proteolytic activity was demonstrated when a strain of Microsporum canis was cultured in broth with human hair, but not in the medium without hair. The extracellular proteinase (keratinase) was isolated and purified by chromatography. Disc electrophoresis showed one protein band of extracellular proteinase, and the antibody against this enzyme gave a single precipitin line in agar diffusion. The IgG fraction completely neutralized the proteinase activity. The proteinase of M. canis may play a role in infection caused by this fungus, by affecting keratinized tissue such as stratum corneum and hair.

Partial characterization of the extracellular keratinase from Microsporum canis

The extracellular keratinase of Microsporum canis released peptides from alpha-type fibrous protein and the membranous fraction isolated from human stratum corneum. Inhibition of the enzyme by phenylmethyl-sulfonylfluoride and its weak inhibition by N-ethylmaleimide and etheneglycol tetra-acetic acid indicated that it is probably a serine proteinase.

Experimental penetration of Trichophyton mentagrophytes into human stratum corneum

We present confirmation of the experimental penetration of Trichophyton mentagrophytes into human stratum corneum under designated conditions of temperature and humidity. When stratum corneum, obtained from healthy human heel region, was incubated at 100% humidity, mycelium was observed in the corneum layer on day 2 at 35 °C and 27 °C, and on day 4 at 15 °C. At 90% humidity, the hyphae penetrated into the stratum corneum on day 4 at 35 °C, and on day 6 at 27 °C. Whereas, at 80% humidity, no fungal elements were observed in the stratum corneum at both 27 °C and 35 °C for up to 7 day. These data suggested that humidity was a more important environmental factor for penetration than temperature, and that at least 90% humidity is necessary for dermatophytes to penetrate into the stratum corneum within a few days. Mean humidity in the interdigital space between the fourth and fifth toes was found to be approximately 98%.

A comparative clinical study between 2 weeks of luliconazole 1% cream treatment and 4 weeks of bifonazole 1% cream treatment for tinea pedis

The aim of the study was to compare the efficacy and safety of luliconazole 1% cream and bifonazole 1% cream as applied in the treatment of tinea pedis (interdigital‐type and plantar‐type). A multi‐clinic, randomised single‐blind, parallel group study with 34 hospitals and 11 clinics formed the study design. Five hundred and eleven patients with mycologically confirmed tinea pedis were included. Of the 489 evaluable patients, 247 were randomised to luliconazole, and 242 to bifonazole. Luliconazole 1% cream applied once a day for 2 weeks, followed by a placebo cream for 2 weeks, thereafter. Bifonazole 1% cream applied once a day for 4 weeks. Mycological effect (negative result on microscopy) and improvement of skin lesions were measured at weeks 1, 2, 3 and 4. Safety frequency and severity of adverse reactions were also measured. The improvement of skin lesions after 4 weeks was comparably good with rates of 91.5% vs. 91.7% (luliconazole vs. bifonazole). The mycological effect was characterised by high negative rates of 76.1% vs. 75.9% (luliconazole vs. bifonazole). The progression of tinea‐related signs and symptom scores differed insignificantly between evaluated luliconazole and bifonazole treatment groups comprising a total of 500 patients. Both substances appeared to be comparably safe and well‐tolerated.

Seborrhoeic Dermatitis: Treatment with Anti‐Mycotic Agents

In order to elucidate the effectiveness of anti‐mycotics in treating seborrhoeic dermatitis, an attempt was made to isolate Malassezia from seborrhoeic lesions of patients of seborrhoeic dermatitis. The results revealed that, in male patients, 46/49 cases were positive for Malassezia furfur on the face and 30/48 cases were positive for M. furfur on the scalp. In female patients, 7/13 cases were positive for M. furfur on the face, and 6/17 cases were positive for M. furfur on the scalp. Anti‐mycotic agents were excellent in 50% and good in 31% of the spore‐positive cases, yielding an overall efficacy rate of 81%. In contrast, the treatment of the face with vehicle alone showed only one excellent result out of 8 cases. Although clinical improvement was rapid on the side treated with a topical corticosteroid in the half‐side‐test, numerous fungal elements remained. While the improvement with anti‐mycotic agents was slower than that with the corticosteroid, clinical improvement became evident by the third week of administration and fungal elements disappeared.

Dose‐finding comparative study of 2 weeks of luliconazole cream treatment for tinea pedis – comparison between three groups (1%, 0.5%, 0.1%) by a multi‐center randomised double‐blind study

Luliconazole is a newly developed imidazolyl antifungal agent. A randomised double‐blind comparative study was designed to assess the efficacy and safety of 1% luliconazole cream (group A), 0.5% cream (group B) and 0.1% cream (group C), in tinea pedis (interdigital type and plantar type), when used once daily for 2 weeks. Follow‐ups were performed at 4 weeks after the end of topical treatment. A total of 241 patients were enrolled and 213 patients were evaluated for efficacy. Rates of improvement of skin lesions in the A, B and C groups assessed at week 4 were 90.5%, 91.0% and 95.8%, respectively. Rates of mycological cure (negative result of microscopy) in the A, B and C groups assessed at week 4 were 79.7%, 76.1%, 72.2% and at week 6 (at 4 weeks after the end of topical treatment) were 87.7%, 94%, 88.9%, respectively. For the mycological effect on tinea pedis of the interdigital type at 2 weeks, the negative conversion of fungi showed a concentration‐dependent relationship and indicated a difference in tendency statistically 81.1% (1%– treatment), 62.9% (0.5%– treatment), 58.3% (0.1%– treatment) (Fishers exact test, P = 0.079) and there was a trend between three groups by Cochran–Mantel–Haenszel method (P = 0.038). The incidence of adverse events in which a causal relationship to this drug could not be ruled out was low (2.6%). All of the adverse events were mild in severity and insignificant clinically.

IMMUNOLOGICAL STUDIES OF AN EXTRACELLULAR KERATINASE

The extracellular keratinase from M. canis was purified using ion exchange chromatography and gel filtration. The purified enzyme showed a single protein band on sodium dodecyl sulfate‐polyacrilamide gel electrophoresis. A molecular weight of approximately 45,000 was determined by SDS‐electrophoresis.

Bullous Pemphigoid Complicated with Pemphigus vulgaris

A 67-year-old housewife had polymorphous eruptions with tense bullae on the extremities and upper trunk. Erosions were noted on the oral and esophageal mucosa. A skin biopsy showed subepidermal blistering with linear deposition of IgG and C3 at the basement membrane zone. In addition, intercellular deposition was observed at the lower part of the epidermis. On the basis of clinical and histological features, a diagnosis of bullous pemphigoid (BP) was made. After she was treated with prednisolone, the whole-skin lesions soon disappeared. An esophageal biopsy showed intercellular deposition of IgG and C3. Indirect immunofluorescence testing using esophageal mucosa of guinea pigs showed the same deposition at the titer of 1:20. Immunoblottings with this patients serum revealed no reactions with either BP or pemphigus antigens. Her serum reacted strongly with a 190-kD protein, the nature of which was unknown. A treatment with ciclosporin was added to this steroid therapy, and the mucosal lesions improved gradually.

Staphylococcal Scalded Skin Syndrome in an Adult

We report a case of a 77‐year‐old man with staphylococcal scalded skin syndrome (SSSS). The patient, who had no physical abnormalities other than herpes zoster, showed intact humoral and cellular immunity. A 3‐ and 1‐phage group strain of Staphylococcus aureus was isolated from an unruptured pustule of the herpes zoster lesion, but not from the throat. Inoculation of the isolated bacterium into newborn mice produced a characteristic Nikolsky sign. Histological diagnosis of SSSS was established by findings of intraepithelial cleavage through the stratum granulosum and a lack of inflammatory cells in both dermis and epidermis. After a ten‐day course of oral cefaclor (250 mg every 8 hours) and topical triamcinolone acetonide containing fradiomycin and gramycidin J, the patient recovered completely. The patient left the hospital without any scars other than those of herpes zoster.

Histological identification of prolonged survival of a skin allograft on an extensively burned patient

We applied skin allografts obtained from 38 unrelated volunteers to a very extensively burned female patient. Seven months after skin transplantation, one allograft from a male still survived clinically. In order to ascertain that the graft site resulted from growth of the male donor skin, the allografted area and patients own unburned normal area were examined by the fluorescent staining of Y-body (Y-chromosomal fluorescence). Y-body was detected in 57 per cent and 65 per cent of cells from the grafted area, whereas no fluorescence was detected in the patients own unburned normal area. These observations suggest that one allograft from an unrelated donor survived for 221 days on a very extensively burned subject.