Lineage-specific transcriptional regulation of DICER by MITF in melanocytes.

DICER is a central regulator of microRNA maturation. However, little is known about mechanisms regulating its expression in development or disease. While profiling miRNA expression in differentiating melanocytes, two populations were observed: some upregulated at the pre-miRNA stage, and others upregulated as mature miRNAs (with stable pre-miRNA levels). Conversion of pre-miRNAs to fully processed miRNAs appeared to be dependent upon stimulation of DICER expression--an event found to occur via direct transcriptional targeting of DICER by the melanocyte master transcriptional regulator MITF. MITF binds and activates a conserved regulatory element upstream of DICER's transcriptional start site upon melanocyte differentiation. Targeted KO of DICER is lethal to melanocytes, at least partly via DICER-dependent processing of the pre-miRNA-17 approximately 92 cluster thus targeting BIM, a known proapoptotic regulator of melanocyte survival. These observations highlight a central mechanism underlying lineage-specific miRNA regulation which could exist for other cell types during development.

A fundamental study of cryoablation on normal bone: diagnostic imaging and histopathology.

Cryoablation is a minimally invasive cancer treatment. In this study, the effects of cryoablation on normal rabbit bone were evaluated using imaging and histopathological examinations. Cryoablation was performed using a Cryo-Hit (Galil Medical, Yokneam, Israel). Under anesthesia, one cryoablation needle was inserted at the center of the femur (day 0). To create an ice ball (2 x 3 cm), two 10-min freeze cycles were performed, separated by a 5-min thaw cycle. During cryoablation, changes in the bone and regional tissue were monitored using magnetic resonance imaging (MRI). MRI scans, computed tomography (CT) scans, and collections from the femur (for histopathological evaluation) were performed on days 7, 14, 28, and 56. In terms of the all rabbits' general conditions, we did not observe lameness, decreased appetite, or any other side effects during the experimental periods. Histopathological evaluations of the femur were performed using hematoxylin and eosin staining. MRI indicated inflammation around the ice ball on day 7. Subsequently, the area of inflammation gradually decreased from days 14 to 56. In the histopathological examination, necrosis of bone marrow cells and endosteum were observed from days 7 to 56. No regeneration of bone marrow cells was observed during the experimental period. On the other hand, cryoablation did not influence osteoblasts. Furthermore, there was no pathologic fracture during the experimental period. Our results suggest that cryoablation does not induce severe adverse effects on normal bone, and therefore has potential as a therapeutic option for bone tumors, including metastatic tumors to bone.

S100C/A11 is a key mediator of Ca(2+)-induced growth inhibition of human epidermal keratinocytes.

An increase in extracellular Ca2+ induces growth arrest and differentiation of human keratinocytes in culture. We examined possible involvement of S100C/A11 in this growth regulation. On exposure of the cells to high Ca2+, S100C/A11 was specifically phosphorylated at 10Thr and 94Ser. Phosphorylation facilitated the binding of S100C/A11 to nucleolin, resulting in nuclear translocation of S100C/A11. In nuclei, S100C/A11 liberated Sp1/3 from nucleolin. The resulting free Sp1/3 transcriptionally activated p21CIP1/WAF1, a representative negative regulator of cell growth. Introduction of anti-S100C/A11 antibody into the cells largely abolished the growth inhibition induced by Ca2+ and the induction of p21CIP1/WAF1. In the human epidermis, S100C/A11 was detected in nuclei of differentiating cells in the suprabasal layers, but not in nuclei of proliferating cells in the basal layer. These results indicate that S100C/A11 is a key mediator of the Ca(2+)-induced growth inhibition of human keratinocytes in culture, and that it may be possibly involved in the growth regulation in vivo as well.

A novel fusion gene of collagen type I alpha 1 (exon 31) and platelet-derived growth factor B-chain (exon 2) in dermatofibrosarcoma protuberans.

Dermatofibrosarcoma protuberans (DFSP) is an uncommon, slow growing, sarcoma of dermal and subcutaneous tissue with an infiltrative growth pattern. Although DFSP has a high rate of local recurrence, it rarely metastasizes. DFSP is characterized by a chromosomal translocation involving the collagen type I a 1 (COL1A1) gene on chromosome 17 and the platelet-derived growth factor B-chain (PDGFB) gene on chromosome 22. Various exons of the COL1A1 gene have been reported to be involved in the fusion with exon 2 of the PDGFB gene. In this study, we examined the COL1A1-PDGFB fusion transcripts using frozen specimens from three patients with DFSP. The molecular biology study with reverse transcriptase polymerase chain reaction (RT- PCR) and sequencing showed that the ends of exons 25, 31, and 45 in the COL1A1 gene were fused with PDGFB. The exon 2 of the PDGFB gene fused with exon 31 of the COL1A1 gene was a novel fusion gene.

Melanocytic nevi clinically simulating melanoma.

Melanoma and other benign or malignant pigmented skin tumors can significantly overlap in their clinical and dermoscopical presentations. Thus, pigmented skin lesions may be misdiagnosed in a large number of cases. An extensive review of the published work provides numerous examples of benign lesions mimicking melanoma. Although a number of melanocytic nevi may have been identified as melanomas, information about their clinical appearance is limited. In this report, we present the clinical appearances of two melanocytic nevi on the vulva and the upper extremity that were difficult to diagnose clinically. Detecting melanoma at an early stage is of the utmost importance. However, more attention should be given to the diagnostic accuracy of benign pigmented skin lesions, which otherwise may be diagnosed and treated as melanoma.

A randomized double-blind trial of intravenous immunoglobulin for bullous pemphigoid.

BACKGROUND: Patients with steroid-resistant bullous pemphigoid (BP) require an appropriate treatment option. OBJECTIVE: A multicenter, randomized, placebo-controlled, double-blind trial was conducted to investigate the therapeutic effect of high-dose intravenous immunoglobulin (IVIG; 400mg/kg/day for 5days) in BP patients who showed no symptomatic improvement with prednisolone (≥0.4mg/kg/day) administered. METHODS: We evaluated the efficacy using the disease activity score on day15 (DAS15) as a primary endpoint, and changes in the DAS over time, the anti-BP180 antibody titer, and safety for a period of 57days as secondary endpoints. RESULTS: We enrolled 56 patients in this study. The DAS15 was 12.5 points lower in the IVIG group than in the placebo group (p=0.089). The mean DAS of the IVIG group was constantly lower than that of the placebo group throughout the course of observation, and a post hoc analysis of covariance revealed a significant difference (p=0.041). Furthermore, when analyzed only in severe cases (DAS≥40), the DAS15 differed significantly (p=0.046). The anti-BP180 antibody titers showed no difference between the two groups. CONCLUSION: IVIG provides a beneficial therapeutic outcome for patients with BP who are resistant to steroid therapy.

Cutaneous angiolipoleiomyoma.

We describe a girl with cutaneous angiolipoleiomyoma on the buttock. The 16-year-old girl had a 2.5- x 1.5-cm subcutaneous tumor on the right buttock, which was slightly tender. The tumor appeared to be vascular and was, therefore, surgically excised. Histologically, the lesion was poorly circumscribed and was composed of differently sized blood vessels, smooth-muscle bundles, and mature adipose tissue. These histologic findings were consistent with those of angiomyolipoma, which commonly occurs in the kidney. Cutaneous angiomyolipoma, which is also known as cutaneous angiolipoleiomyoma, is a rare benign mesenchymal tumor. To our knowledge, only 16 cases have been reported in the English-language literature. In our report, we review the clinical features of 17 cases, including the current one. We point out the differences between the cutaneous and renal forms of angiomyolipoma, and conclude that the cutaneous lesion is distinct from a renal lesion in several aspects, including tuberous sclerosis complex association and immunoreactivity to both HMB-45 and MART-1.

Congenital atrophic dermatofibrosarcoma protuberans detected by COL1A1-PDGFB rearrangement.

BACKGROUND: Atrophic variant of dermatofibrosarcoma protuberans (DFSP) is a distinct form of DFSP. CASE PRESENTATION: Here, we report the case of a 19-year-old woman with a small congenital atrophic plaque on the right precordium. The lesion remained atrophic for more than 10 years. Several years earlier, a portion of the plaque became tuberous and enlarged. Physical examination revealed a 25 × 30 mm erythematous atrophic plaque surrounded by three hard, smooth, and orange-colored nodules of varying sizes on the right precordium, along with visible subcutaneous adipose tissue and cutaneous veins. Biopsy of the nodule and atrophic plaque revealed dense proliferation of spindle-shaped tumor cells from the dermis to the subcutaneous adipose tissue, and positive immunostaining for CD34 and vimentin in addition to negative staining for factor XIIIa and α-smooth muscle actin. Reverse transcription polymerase chain reaction (RT-PCR) of the tumor tissue revealed the presence of a COL1A1-PDGFB fusion gene. Thus, congenital atrophic dermatofibrosarcoma protuberans was diagnosed. No metastasis to the lungs or regional lymph nodes was found on magnetic resonance imaging. Wide local excision and split-thickness skin grafting was performed and neither recurrence nor metastasis has been observed for 5 years and 8 months since the surgery. CONCLUSION: This case indicates that a congenital atrophic lesion could represent a quiescent phase of DFSP. Awareness of this rare condition can aid with early diagnosis and thereby improve the prognosis of DFSP.

Introduction of an N-terminal peptide of S100C/A11 into human cells induces apoptotic cell death.

S100 proteins belong to the EF-hand Ca2+-binding protein family and are involved in the regulation of a variety of cellular processes. Individual S100 proteins are expressed in cell- and tissue-specific manners, and functional deterioration of S100 proteins leads to a number of human diseases, including cancer. We previously demonstrated that S100C/A11 was translocated to nuclei and inhibited DNA synthesis in human keratinocytes when exposed to high Ca2+. In the present study we examined the effects of synthetic partial peptides of S100C/A11 on human carcinoma cell lines. Only an N-terminal peptide with 19 amino acid residues (MAK19) showed cytotoxicity to the cell lines in dose- and time-dependent manners when introduced into cells by flanking the HIV-TAT protein transduction domain (TAT-MAK19). Pulse field electrophoresis revealed that DNA of the treated cells was partially degradated. Annexin V, a marker of cellular apoptosis, was detected in the cells treated with TAT-MAK19 by immunostaining and flow cytometry. The induction of apoptotic cell death was apparently independent of p53, p21WAF1/CIP1, and caspase activity, but treatment with TAT-MAK19 resulted in partial translocation of apoptosis-inducing factor (AIF) from the cytoplasm to nuclei. These results indicate that MAK19 induces apoptosis in human cell lines and may therefore lead to the establishment of a new molecular target for the treatment of human cancer.

Inflammatory stage of disseminated superficial porokeratosis.

Disseminated superficial porokeratosis (DSP) is a keratinization disorder characterized by multiple small lesions with a slightly elevated, sharply defined ridge over the whole body. Unusual DSP cases with acute exacerbation of their lesions accompanied by severe pruritus have been reported and designated as "eruptive pruritic papular porokeratosis" or "inflammatory DSP". Histologically, the pruritic lesions in the majority of these unusual DSP cases had a dense infiltration of eosinophils and lymphocytes in the vicinity of blood vessels in the upper dermis. In this report, we describe an additional case of DSP with a similar clinical course and histopathological findings. A review of the literature showed that the pruritic condition in these unusual DSP cases can be transient and is not necessarily related to spontaneous regression. We propose the term "Inflammatory stage of DSP" for describing this unusual variant of DSP.

Localization of S100C immunoreactivity in various human tissues.

Using 2-dimensional gel electrophoresis, we previously demonstrated that the S100C protein remarkably decreased after immortalization of normal human fibroblasts, and that this protein caused growth inhibition of human tumor cells when forcibly expressed in these cells, suggesting that S100C plays a significant role in tumor suppression. The present study was carried out to determine what type of human tissues express S100C protein, and, subsequently, whether the S100C content in these tissues changes after normal cells have been transformed into cancer cells. We found that ductal cells in various tissues were positively stained with the S100C protein. In comparison, epithelial cells in digestive organs such as the stomach, small intestine, and colon were not stained as strongly. When 14 pairs of human normal and cancerous tissues were stained with the antibody, decreases in the staining levels of S100C were observed in 6 kinds of cancerous tissues--from the bronchus, mammary duct, renal tubule, prostate, uterus, and testis--in comparison with staining in their normal counterparts. These results suggest that S100C is a new tumor marker protein, the expression of which significantly decreases after malignant transformation of human tissues.

The structure of S100A11 fragment explains a local structural change induced by phosphorylation.

S100A11 protein is a member of the S100 family containing two EF-hand motifs. It undergoes phosphorylation on residue T10 after cell stimulation such as an increase in Ca(2+) concentration. Phosphorylated S100A11 can be recognized by its target protein, nucleolin. Although S100A11 is initially expressed in the cytoplasm, it is transported to the nucleus by the action of nucleolin. In the nucleus, S100A11 suppresses the growth of keratinocytes through p21(CIP1/WAF1) activation and induces cell differentiation. Interestingly, the N-terminal fragment of S100A11 has the same activity as the full-length protein; i.e. it is phosphorylated in vivo and binds to nucleolin. In addition, this fragment leads to the arrest of cultured keratinocyte growth. We examined the solution structure of this fragment peptide and explored its structural properties before and after phosphorylation. In a trifluoroethanol solution, the peptide adopts the alpha-helical structure just as the corresponding region of the full-length S100A11. Phosphorylation induces a disruption of the N-capping conformation of the alpha-helix, and has a tendency to perturb its surrounding structure. Therefore, the phosphorylated threonine lies in the N-terminal edge of the alpha-helix. This local structural change can reasonably explain why the phosphorylation of a residue that is initially buried in the interior of protein allows it to be recognized by the binding partner.

Onycholemmal carcinoma.

A 70-year-old Japanese man presented with a 5-year history of refractory indolent onycholysis of the little finger of the right hand. Roentgenograms did not show involvement of the bone. Histological examination revealed an epithelial tumor consisting of lobular masses varying in size. The tumor was composed of keratinocytes varying in atypicality and showed infiltrative growth into the dermis but not into the phalangeal bone. The tumor had cystic structures composed of eosinophilic amorphous keratin and a surrounding thin layer of keratinocytes. Characteristically, the epithelium in the center of the tumor abruptly changed into amorphous keratin without the formation of intervening keratohyaline granules. From these findings, the mass was diagnosed as onycholemmal carcinoma. Immunohistochemically, the tumor showed a keratin profile comparable to that of the nail bed epithelium and a smaller number of Ki-67-positive proliferating tumor cells compared with those of a previous case of onycholemmal carcinoma.

Minimal change nephrotic syndrome developing during postoperative interferon-beta therapy for malignant melanoma.

A 64-year-old man presented with proteinuria during postoperative interferon (IFN)-beta therapy against malignant melanoma. Renal pathologic findings were consistent with minimal change nephrotic syndrome (MCNS) showing extensive foot process effacement of visceral glomerular epithelial cells (podocyte). Nephrotic range proteinuria gradually regressed after stoppage of local injection of IFN-beta without glucocorticoid treatment. To our knowledge this is the first report that demonstrates histological abnormalities of the glomerulus associated with postoperative IFN-beta therapy for the malignant melanoma.