Ayumi Adachi

Cloning and characterization of human MCM7 promoter

MCM7 is a member of the MCM protein family which has been implicated in the regulatory machinery allowing DNA to replicate only once during S phase. In quiescent cells, human MCM7 (hMCM7) mRNA is almost undetectable. Stimulation of cells to enter the cell cycle results in induction of hMCM7 expression. Here, we report cloning and characterization of the hMCM7 promoter. We isolated and sequenced a 0.5 kb genomic fragment that contains putative transcription factor binding sites including three E2F sites, three GC boxes and an E box. Several transcription start sites, which were used upon growth stimulation, were identified. The minimal promoter region required for transcription of a luciferase reporter gene was delineated, and it contained an E box and one E2F site, which were important for promoter activity. Interestingly, the cloned sequence appears to act as a promoter for mu-adaptin-related protein 2 (mu-ARP2) gene in the opposite orientation.

Immunolocalization of hCDC47 protein in normal and neoplastic human tissues and its relation to growth

hCDC47 is a human member of the MCM family, which has been implicated in the regulatory machinery causing DNA to replicate once per cell cycle. We examined its protein expression and localization in normal human tissues, using immunostaining with polyclonal antibodies. Positive nuclei were found in the proliferative components of lymph nodes, bone marrow, epidermis and mucosa. Immunohistochemical analysis was also performed for 3 types of cutaneous keratinocytic tumor originating from same cell type but showing different grades of malignancy. In seborrheic keratosis, a benign condition, cells with hCDC47‐positive nuclei were located in the outermost layers of the tumor lobules, while in Bowens disease, carcinomas in situ and squamous‐cell carcinomas, they were present throughout the lesions. The percentages of hCDC47‐positive cells were 65.4% in squamous‐cell carcinomas, 60.9% in Bowens disease, 12.6% in seborrheic keratosis and 3.9% in normal epidermis (n = 5 in all cases). Further expansion of the analysis to include malignant tumors from several other organs revealed that all malignant lesions tested contained more nuclear hCDC47‐positive cells than their normal counterparts. Our findings indicate that hCDC47 plays a role in normal and neoplastic cell growth in vivo and that hCDC47 immunolocalization could be used as an index of cell proliferation in tissue sections.Int. J. Cancer 74:180–184, 1997.

Human papillomaviruses of the mucosal type are present in some cases of extragenital Bowen's disease

Background  Bowens disease in the genital area is generally considered to be caused by mucosal high‐risk human papillomaviruses (HPVs). However, the detection rate and spectrum of HPVs in extragenital Bowens disease are various and it is not clear to what extent HPV is involved in its pathogenesis.

Detection of human papillomavirus type 56 DNA, belonging to a mucous high‐risk group, in hair follicles in the genital area of a woman no longer suffering from viral warts

Background  Human papillomaviruses (HPVs) parasitize human epithelium, but it is not clear where they reside when they do not cause apparent infection. Hair follicles are important candidates as reservoirs.

The role of polymorphonuclear leukocyte infiltration in herpes simplex virus infection of murine skin.

Abstract We undertook the present study to investigate the role of polymorphonuclear leukocytes (PMN) in defending skin against herpes simplex virus (HSV) infection. For this purpose, we established a mouse model of cutaneous HSV infection. The hind limb footpad skin of 4-week-old ICR mice was abraded linearly once with a feather edge file and infected with various strains of HSV with different virulence. In uninfected control mice, PMN appeared at the abraded skin lesion within 24 h, and were eliminated from the epidermis after 3 days. Mice inducted with a highly virulent strain of HSV demonstrated wide and severe erythematous lesions of the footpad skin and histologically, virus antigen-positive ballooning degenerated keratinocytes were observed. However, in infections with attenuated strains of HSV, the epidermis was regenerated and a viral antigen was discharged within 5 days, together with any infiltrated PMN. Macrophages and NK cells numbered less than PMN. In mice treated with anti-PMN antiserum before HSV infection, PMN infiltration was significantly suppressed 1 day after infection, and these animals developed a severe cutaneous disease even if infected with an attenuated virus. These results indicate the importance of PMN in the control of HSV cutaneous infections, especially in the primary infectious phase.

Genome organization and taxonomic position of human papillomavirus type 47 inferred from its DNA sequence.

The complete nucleotide sequence of human papillomavirus type 47 (HPV-47) DNA isolated from the lesion of epidermodysplasia verruciformis (EV) was determined. The computer-aided comparison of HPV-47 with other EV-associated viruses using the available sequence data on them revealed that HPV-47 resembles both HPV-5 and HPV-8 as much as HPV-5 and HPV-8 resemble each other, and it led us to regard these three viruses as one cluster and HPV-19 and HPV-25 as another. The conclusion implies that HPV-47 as well as HPV-5 and HPV-8 is associated with the cancer occurrence in EV. Two sets of splicing donor and acceptor sequences in HPV-47, which were previously shown to work in vivo, are also conserved in HPV-5 and HPV-8. One of them allows formation of an ORF predicted to encode an E1/E4 fused protein.

Specific distribution patterns of hCDC47 expression in cutaneous diseases

hCDC47 is a human member of the MCM family, which has been implicated to be concerned with the regulatory machinery causing DNA to replicate once per cell cycle. In a previous paper, we described hCDC47 expression as being localized in the proliferative component of normal tissues, and showed greater expression in squamous cell skin carcinomas than in seborrheic keratosis. In the present study, we compared its expression in another type of skin tumor and variotis non‐neoplastic cutaneous proliferative diseases. Two patterns of distribution of hCDC47‐positive cells were observed. Keratoacanthomas showed a peripheral pattern in which only the cells located the basal cell layers were positive. Psoriasis vulgaris also showed this peripheral type of location, with the cells in the suprabasal layers also occasionally expressing hCDC47. Verruca vulgaris demonstrated a diffuse pattern, with positive epithelial cells distributed throughout the layers. Basal cell carcinomas also showed a similar pattern. The keratoacanthomas showed the highest hCDC47 positive cell rate (43.1%), followed by verruca vulgaris (42.5%). psoriasis vulgaris (23.4%), and basal cell carcinoma (17.3%). Our results suggest participation of hCDC47 in these proliferative disorders involving keratinocytes.

The successful use of topical tacrolimus treatment for a chronic actinic dermatitis patient with complications of idiopathic leukopenia.

Chronic actinic dermatitis (CAD) is a photosensitivity disorder marked by severe eczematous lesions on exposed areas. Although associations with contact dermatitis, atopic dermatitis, and human immunodeficiency virus (HIV) have been suggested, its pathogenesis remains unknown. CAD is often refractory, and systemic administration of cyclosporin A has been the treatment of choice. Recently, topical tacrolimus therapy has been reported to be effective. We report the efficacy of topical tacrolimus treatment in a CAD patient who also had the complication of idiopathic leukopenia. A phototest showed marked suppression of erythema formation in the skin pre‐treated with tacrolimus before UVB radiation but not in the skin treated after the irradiation. Therefore, it is suggested that tacrolimus may prevent UV‐B induced erythema by suppressing a very early phase of the inflammatory process in CAD.

Two cases of protothecosis in Nagoya, Japan

Two cases of protothecosis caused by Prototheca wickerhamii have been reported from Nagoya in a 12 year period. In both cases the infection presented on the cheeks of otherwise healthy women. Biopsies showed numerous PAS positive staining organisms with the distinctive mulberry‐like endosporulation in the dermis. Prototheca wickerhamii was identified on sugar assimilation tests of colonies isolated from tissue on Sabouraud agar. Case 1 responded to 11 months of oral ketoconazole therapy. Case 2 might not respond to itraconazole. The source of the infections has not been identified.

Successful treatment of warts with a combination of maxacalcitol ointment and salicylic acid sticking plaster

Dear Editor, Warts, or verrucae vulgares, are benign proliferations of the skin and mucosa caused by infection with human papillomaviruses (HPV). They are usually treated by traditional destructive modalities such as cryotherapy.1,2 However, unfavorable side effects of cryotherapy include pain, especially in children, and sometimes scarring or pigmentation after treatment. Here we report an open pilot study of a combination of maxacalcitol ointment and salicylic acid sticking plaster for the treatment of warts on the palms and soles in 22 patients. After obtaining informed consent, 22 patients (mean age, 22 years; range, 3–62) with recalcitrant or untreated warts on the palms and soles were included in our trial. The diagnosis of warts was based on the clinical appearance. The mean duration of the disease was 4.7 months (0.2–17), and nine of the patients had previously received other therapies. The warts were treated with topical maxacalcitol (25 μg/g ointment, Oxarol ointment, Maruho, Osaka, Japan), and then covered with salicylic acid plaster (35.7 mg/cm2, 1 mm thickness, SPEEL-KO M, Nichiban, Tokyo, Japan). Patients treated the warts this way once a day by themselves or their parents did it for them. They applied no other medicine that could affect the warts during the trial. The treatment was regarded as ineffective in patients who had no response 12 weeks after the treatment was started. Patients came to our hospital every 2 weeks, and the changes were evaluated in terms of the number and sizes of the warts. We examined the patients for adverse effects on the skin at every visit and evaluated them according to the following four-grade system: cleared (C), clearing of the wart; improvement (I), the wart decreased or became smaller in size without disappearance; no change (NC); or worse (W), the wart increased or became larger in size. Of the 22 patients, four were lost to follow up, and therefore the data from only 18 patients (10 women and eight men) were evaluated. The results are summarized in Table 1. Thirteen patients (13/22, 59%) showed complete regression. Clinical improvement of the lesions was evident in all 18 patients within 3 months. The mean term of treatment was 45 days (10–90). No warts became worse or were unchanged. Warts located from the nail fold to the bed, which are usually resistant to any treatment, had completely disappeared (Fig. 1C,D), although more time was required for these than for other warts. In cases with multiple warts, only those treated had regressed (Fig. 1E,F). After stopping the treatment, the warts re-grew gradually in patient 14. This indicated to us that the present treatment had certainly been effective in improving the warts. Only in patient 11 did one of the eleven warts relapse 5 months after complete regression, and he was re-treated with the combination therapy for 1 month until a second disappearance. In the other patients, no worsening or recurrence was found for 4 months after this trial. No patient was withdrawn from the study because of clinically apparent adverse events during the medication. Maxacalcitol, an active vitamin D3 analog, has been used for the treatment of psoriasis vulgaris as well as palmoplantar keratosis.3 It is well known that vitamin D3 analogs have some biological actions in epidermal cells, such as the regulation of cell proliferation and differentiation and the modulation of cytokine production.4 Recent reports have suggested that vitamin D3 analogs have anti-tumor effects and may be effective in treating seborrheic keratosis and warts.5,6 According to these reports, this efficacy may depend on the dose of the analogs, and the effect is increased by occlusive dressing therapy (ODT), whereas a simple application has a