Yuko Takeuchi

MFG-E8 Regulates Angiogenesis in Cutaneous Wound Healing

Our research group recently demonstrated that pericytes are major sources of the secreted glycoprotein and integrin ligand lactadherin (MFG-E8) in B16 melanoma tumors, and that MFG-E8 promotes angiogenesis via enhanced PDGF-PDGFRβ signaling mediated by integrin-growth factor receptor crosstalk. However, sources of MFG-E8 and its possible roles in skin physiology are not well characterized. The objective of this study was to characterize the involvement of MFG-E8 in skin wound healing. In the dermis of normal murine and human skin, accumulations of MFG-E8 were found around CD31(+) blood vessels, and MFG-E8 colocalized with PDGFRβ(+), αSMA(+), and NG2(+) pericytes. MFG-E8 protein and mRNA levels were elevated in the dermis during full-thickness wound healing in mice. MFG-E8 was diffusely present in granulation tissue and was localized around blood vessels. Wound healing was delayed in MFG-E8 knockout mice, compared with the wild type, and myofibroblast and vessel numbers in wound areas were significantly reduced in knockout mice. Inhibition of MFG-E8 production with siRNA attenuated the formation of capillary-like structures in vitro. Expression of MFG-E8 in fibrous human granulation tissue with scant blood vessels was less than that in granulation tissue with many blood vessels. These findings suggest that MFG-E8 promotes cutaneous wound healing by enhancing angiogenesis.

Immunohistochemical analyses of p63 expression in cutaneous tumours

1 Belardi MG, Maglione MA, Vighi S et al. Syringoma of the vulva. A case report. J Reprod Med 1994; 12:957–9. 2 Matsumoto K, Satou K, Inamura M et al. A case of eruptive syringoma. Rinsho Derma 2002; 44:87–9 (in Japanese). 3 Shimada Y, Akimoto S, Warita S et al. A case of localized vulval syringoma. Jpn J Clin Dermatol 2003; 57:917–19 (in Japanese). 4 Komatsu H, Kojima M, Tsutsumi N et al. Study of the mechanism of inhibitory action of tranilast on chemical mediator release. Jpn J Pharmacol 1988; 46:43–51. 5 Yamada H, Tajima S, Nishikawa T et al. Tranilast, a selective inhibitor of collagen synthesis in human skin fibroblasts. J Biochem (Tokyo) 1994; 116:892–7. 6 Huang YH, Chuang YH, Kuo TT et al. Vulvar syringoma: a clinicopathologic and immunohistologic study of 18 patients and results of treatment. J Am Acad Dermatol 2003; 48:735–9. 7 Reitamo S, Anttila HS, Didierjean L et al. Immunohistochemical identification of interleukin 1 alpha and beta in human eccrine sweat-gland apparatus. Br J Dermatol 1990; 122:315–23. 8 Suzawa H, Kikuchi S, Ichikawa K et al. Inhibitory action of tranilast, an anti-allergic drug, on the release of cytokines and PGE2 from human monocytes-macrophages. Jpn J Pharmacol 1992; 60:85–90. 9 Seifert HW. Multiple Syringome mit Vermehrung von Mastzellen unter dem klinischen Bild einer Urticaria pigmentosa. Z Hautkr 1981; 56:303–6. 10 Kameyoshi Y, Morita E, Tanaka T et al. Interleukin-1 alpha enhances mast cell growth by a fibroblast-dependent mechanism. Arch Dermatol Res 2000; 292:240–7.

The prevalence of Merkel cell polyomavirus in Japanese patients with Merkel cell carcinoma

BACKGROUND A novel polyomavirus, the Merkel cell polyomavirus (MCPyV) has been implicated in the pathogenesis of Merkel cell carcinoma (MCC); however, the prevalence of MCPyV in Japan has not been extensively investigated. OBJECTIVE To clarify the prevalence of MCPyV in Japanese patients with MCC. METHODS MCPyV DNA was examined by polymerase chain reaction (PCR) in formalin-fixed paraffin-embedded (FFPE) or frozen tissue samples from 26 patients with MCC diagnosed in four medical centers in Japan. Immunohistochemistry was simultaneously performed using a monoclonal antibody against the viral large T (LT) antigen. FFPE samples from basal cell carcinoma (BCC) and squamous cell carcinoma (SCC) were also analyzed as controls. RESULTS Twenty-three out of 26 cases (88.5%) were positive for MCPyV DNA by PCR. The amplified products harbored 4 patterns of mutations. Phylogenetic analysis demonstrated that one of our strains was closely related to the other Japanese strains previously reported. The LT antigen was expressed in various degrees in 20 of 26 cases (76.9%) by immunohistochemistry. Histological type had little relation to CM2B4 positivity, whereas 3 of 5 trabecular-type tumors showed no staining. The immunoreactivity for CM2B4 did not correlate with the relative viral DNA load. In BCC and SCC, the LT antigen was immunohistochemically positive, but MCPyV DNA was not detected by PCR. The cells around some MCC and non-MCC tumors were stained with CM2B4 with a distribution similar to CD20- and CD45RO- (especially CD8-) positive lymphocytes. CONCLUSION MCPyV was highly positive in Japanese patients with MCC. It is of note that the positive rate differs depending upon the detection method.

IMMUNOHISTOCHEMICAL DISTRIBUTION OF EPIMORPHIN IN HUMAN AND MOUSE TISSUES

A novel protein epimorphin has been identified as a mesenchymal signal factor. We reported previously ubiquitous expression of epimorphin in normal skin and a significant increased expression in diseased human skin. The present immunofluorescence study was conducted to determine systematically the distribution of epimorphin in adult human organs with an anti-epimorphin monoclonal antibody. Epimorphin was found to be widely distributed in all human organs examined. It was present in the connective tissue adjacent to or around various epithelial tissues, muscles and vessels. In particular, strong staining was present on the endomysium of muscles, the adventitia of blood vessels, along the sinusoidal lining of hepatocytes and connective tissue around epithelial cells, exocrine and endocrine glands. The results suggest that epimorphin may play a key role in maintaining normal tissue structure and interaction between mesenchymal tissue and epithelial tissue in vivo. ©; 1998 Chapman & Hall

MFG-E8 Drives Melanoma Growth by Stimulating Mesenchymal Stromal Cell–Induced Angiogenesis and M2 Polarization of Tumor-Associated Macrophages

Secretion of the powerful angiogenic factor MFG-E8 by pericytes can bypass the therapeutic effects of anti-VEGF therapy, but the mechanisms by which MFG-E8 acts are not fully understood. In this study, we investigated how this factor acts to promote the growth of melanomas that express it. We found that mouse bone marrow-derived mesenchymal stromal cells (MSC) expressed a substantial amount of MFG-E8. To assess its expression from this cell type, we implanted melanoma cells and MSC derived from wild type (WT) or MFG-E8 deficient [knockout (KO)] into mice and monitored tumor growth. Tumor growth and M2 macrophages were each attenuated in subjects coimplanted with KO-MSC compared with WT-MSC. In both xenograft tumors and clinical specimens of melanoma, we found that MFG-E8 expression was heightened near blood vessels where MSC could be found. Through in vitro assays, we confirmed that WT-MSC-conditioned medium was more potent at inducing M2 macrophage polarization, compared with KO-MSC-conditioned medium. VEGF and ET-1 expression in KO-MSC was significantly lower than in WT-MSC, correlating in vivo with reduced tumor growth and numbers of pericytes and M2 macrophages within tumors. Overall, our results suggested that MFG-E8 acts at two levels, by increasing VEGF and ET-1 expression in MSC and by enhancing M2 polarization of macrophages, to increase tumor angiogenesis. Cancer Res; 76(14); 4283-92. ©2016 AACR.

The Comparative Study of Anti-Double Stranded DNA Antibody Levels Measured by Radioimmunoassay and Enzyme-Linked Immunosorbent Assay in Systemic Lupus Erythematosus

Anti‐double stranded (ds) DNA antibody is an autoantibody specific for systemic lupus erythematosus (SLE). For the measurement of this antibody, radioimmunoassay (RIA) or enzyme‐linked immunosorbent assay (ELISA) is widely used in Japan. We studied the correlation of anti‐dsDNA levels in 158 sera from 53 SLE patients between RIA and ELISA. The correlation coefficient between anti‐dsDNA antibody levels measured by RIA and ELISA was 0.736 (p<0.001). The concordance rate of sera with both ELISA/RIA‐positive or both ELISA/RIA‐negative results was 82.3% (130/158 sera). Twenty sera (12.7%) of 14 patients were ELISA‐positive but RIA‐positive, and 8 sera (5.0%) of 6 patients were ELISA‐negative but RIA‐positive. Our results suggest that both methods may be equally reliable for the detection of anti‐dsDNA antibodies and that ELISA may be more sensitive than RIA. ELISA may be preferable because of its simplicity and convenience of the measurement procedure. The correlation coefficient between anti‐ssDNA levels measured by ELISA and anti‐dsDNA levels measured by RIA was 0.322 (p<0.01), and the correlation coefficient between anti‐ssDNA and anti‐dsDNA antibody levels measured by ELISA was slightly higher, 0.515 (p<0.001). These results may reflect the predominance of anti‐DNA antibodies reactive with both ss and dsDNA in SLE sera. Since the clinical significance of anti‐ssDNA antibodies remains unclear, further analysis of accumulated cases is required.

Association of antiphosphatidylserine/prothrombin antibodies with neuropsychiatric systemic lupus erythematosus

Neuropsychiatric manifestations in patients with systemic lupus erythematosus (SLE) are well-recognized symptoms although the pathophysiology of neuropsychiatric SLE (NPSLE) is unclear. Since an association with antiphospholipid antibodies has been reported, we examined the prevalence of antiphosphatidylserine–prothrombin antibodies (anti-PS/PT Abs), lupus anticoagulant (LA), anticardiolipin/β2-glycoprotein I antibodies (anti-β2-GPI Abs), and antiribosomal P protein antibodies (antiribosomal P Abs) in 68 SLE patients and analyzed their associations with neuropsychiatric manifestations. The prevalence of LA was significantly higher in the patients with neuropsychiatric (NP) features than those without NP features (P < 0.02). The levels of anti-PS/PT antibody were also significantly higher in the patients with NP features than those without NP features (P < 0.01). The results indicate that LA positivity and higher levels of anti-PS/PT antibody can be predictive markers for NPSLE.

Protective Effect of MFG-E8 after Cutaneous Ischemia–Reperfusion Injury

We recently demonstrated that the secreted glycoprotein and integrin-ligand MFG-E8 promotes cutaneous wound healing by enhancing angiogenesis. Several studies have identified potential roles for MFG-E8 in regulation of ischemia-reperfusion (I/R) injury in the brain, kidney, and liver. Our objective was to assess the role of MFG-E8 in the formation of skin ulcers using a murine model of cutaneous I/R injury-cutaneous pressure ulcers. Cutaneous I/R was performed by trapping the dorsal skin between two magnetic plates for 12 hours, followed by plate removal. Expression of MFG-E8 increased in the dermis during ischemia, and then decreased after reperfusion. Administration of recombinant (r)MFG-E8 in I/R areas at the beginning of reperfusion significantly inhibited the formation of cutaneous pressure ulcers, and the number of CD31(+) vessel and NG2(+) pericytes in wounds were increased in I/R mice treated with rMFG-E8. The number of M1 macrophages and the amount of proinflammatory mediators monocyte chemotactic protein-1,induced nitric oxide synthase, IL-6, tumor necrosis factor-α, and IL-1β in the wound area were reduced by the administration of rMFG-E8. We conclude that MFG-E8 may inhibit the formation of pressure ulcers induced by cutaneous I/R injury by regulating angiogenesis and inflammation. Exogenous application of MFG-E8 might have therapeutic potential for cutaneous I/R injuries, including decubitus ulcers and Raynauds phenomenon-induced digital ulcers.

Amalgam tattoo of the oral mucosa mimics malignant melanoma

Figure 1. Clinical features: A blue-black macule was seen on the floor of the oral cavity adjacent to a restored tooth with dental filling. Dear Editor, Amalgam pigmentation, generally called amalgam tattoo, is a pigmented lesion on the oral mucosa. Amalgam is the most commonly used dental restorative material for dental fillings because it is inexpensive and relatively easy to manipulate during placement. Amalgam is characterized by its initial pliability and it can be easily packed to fill any irregular space. It then forms a hard, durable compound with bacteriostatic effects. Amalgam fillings are made of a mixture of several metallic particles, especially mercury, silver and tin. Clinically, amalgam tattoos appear as blue, black or dark gray asymptomatic, flat macules on the oral mucosa, located adjacent to a restored tooth, most commonly on the gingival surfaces. A pigmented mucosal lesion requires an extensive differential diagnosis, particularly with malignant melanoma. A 63-year-old woman had received treatment to her teeth many years ago. When she received dental scaling 2 months prior, an asymptomatic pigmented lesion on the oral mucosa was pointed out by a dentist. Over the next 2 months, the lesion considerably enhanced. She was referred for evaluation of an asymptomatic pigmented lesion on the oral mucosa. Physical examination revealed a 9 mm · 6 mm blueblack macule, with irregular borders on the left floor of the mouth (Fig. 1). Regional lymphadenopathy was not noted. Adjacent to the pigmented lesion, a tooth restored with a dental filling was confirmed (Fig. 1). The results of routine laboratory tests including hematology, liver and renal functions as well as C-reactive protein levels were within normal limits. X-rays did not show the presence of metallic particles in the mucosa. With the main differential diagnoses of malignant melanoma and amalgam tattoo, the lesion was completely excised.

A possible inhibitory action of diaminodiphenyl sulfone on tumour necrosis factor-α production from activated mononuclear cells on cutaneous lupus erythematosus

Background.  We have previously reported that diaminodiphenyl sulfone (DDS; dapsone) can bring about significant clinical improvement in patients with cutaneous lupus erythematosus (CLE). However, the pharmacological actions of DDS in the skin lesion of patients with CLE are still unknown.