Plasmacytoid dendritic cells suppress Th2 responses induced by epicutaneous sensitization.

Epicutaneous (EC) sensitization with protein allergens is the most important sensitization route for atopic dermatitis. Plasmacytoid dendritic cells (pDCs) are characterized by massive secretion of interferon-α (IFNα). B6 mice are T helper type 1 (Th1)-prone and are representative of non-atopic humans, whereas BALB/c mice are Th2-prone and are representative of atopic humans. Here, we show that naïve BALB/c mice contain a greater number of nonactivated pDCs in peripheral lymph nodes (LNs) than do naïve B6 mice. Naïve BALB/c mice also have more of the CD8α- subset in LNs than naïve B6 mice. Moreover, in vivo depletion of pDCs during EC sensitization results in enhanced Th2 responses in BALB/c mice, but not in B6 mice. Mechanistically, when BALB/c mice undergo EC sensitization, there is an increase in pDCs entering draining LNs. These cells exhibit modest activation including comparable costimulation expression but increased cytokine expression compared with those of naïve mice. In vivo depletion of pDCs during EC sensitization significantly increases the activation of dermal dendritic cells (dDCs) suggesting a regulatory effect on these cells. To this end, a suppressive effect of pDCs on conventional dendritic cells was also demonstrated in vitro. Further, in vivo blockade of IFNα by an anti-IFNAR antibody (Ab) or in vivo reduction of IFNα production of pDCs by anti-siglec-H Ab both resulted in enhanced activation of dDCs. Collectively, our results demonstrate that pDCs suppress Th2 responses induced by EC sensitization via IFNα-mediated regulation of dDCs.

miR-519d Promotes Melanoma Progression by Downregulating EphA4.

Increasing evidence suggests that there is a unique cell subpopulation in melanoma that can form nonadherent melanospheres in serum-free stem cell medium, mimicking aggressive malignancy. Using melanospheres as a model to investigate progression mechanisms, we found that miR-519d overexpression was sufficient to promote cell proliferation, migration, invasion, and adhesion in vitro and lung metastatic capability in vivo The cell adhesion receptor EphA4 was determined to be a direct target of miR-519d. Forced expression of EphA4 reversed the effects of miR-519d overexpression, whereas silencing of EphA4 phenocopied the effect of miR-519d. Malignant progression phenotypes were also affected at the level of epithelial-to-mesenchymal transition and the ERK1/2 signaling pathway inversely affected by miR-519d or EphA4 expression. In clinical specimens of metastatic melanoma, we observed significant upregulation of miR-519d and downregulation of EphA4, in the latter case correlated inversely with overall survival. Taken together, our results suggest a significant functional role for miR-519d in determining EphA4 expression and melanoma progression.Significance: These results suggest a significant role for miR-519d in determining expression of a pivotal cell adhesion molecule that may impact risks of malignant progression in many cancers. Cancer Res; 78(1); 216-29. ©2017 AACR.

A clinicopathological analysis of 153 acral melanomas and the relevance of mechanical stress.

The pathogenesis of melanomas emerging in plantar surfaces remains unclear; however, mechanical stress has been reported to increase the formation of melanomas. In this study, we conducted a multicenter retrospective analysis of 153 acral melanomas diagnosed between 2000 and 2015 in Taiwan. The male-to-female ratio of the patients in question was 1:1.28, and the mean age at diagnosis was 68 years. We examined whether melanomas which developed in different areas of the patients' soles differed in their associations with various clinicopathological characteristics and survival. Testing by goodness of fit indicated that stress-bearing areas were significantly more conducive to the generation of melanomas than non-stress-bearing areas (P < 0.0001). More specifically, compared to the arch, the rear of the foot and front of the foot were significantly more conducive to the generation of melanomas (P < 0.0001 and P < 0.0001, respectively). The distribution pattern was not associated with differences in age, gender, right/left foot involvement, ulceration, mitosis, lymph node metastasis, tumor thickness, or stage. The overall, distant metastasis-free, and recurrence-free survival rates did not differ significantly between the stress-bearing and non-stress-bearing areas. Furthermore, while acral melanomas tended to develop on stress-bearing areas, the distribution pattern was not associated with the prognostic index or survival.

Clinicopathological features and prognosis of patients with de novo versus nevus-associated melanoma in Taiwan.

Studies surveying melanomas associated with melanocytic nevi in Asia are rare. In this study, we examined whether nevus-associated melanomas differ from de novo melanomas in terms of their associations with clinical factors, histologic characteristics, and patient survival in Taiwan. Using data on cancer cases obtained from the Department of Pathology archives and the Cancer Registry of National Taiwan University Hospital, we conducted a retrospective analysis of 103 consecutive melanoma patients who were diagnosed between 2010 and 2015 and received follow-up through November 2016. Approximately 17.5% of the melanomas in question were associated with a nevus. In patients under 65 years of age, non-acral lentiginous melanomas were significantly associated with a higher percentage of nevus-associated melanomas. The superficial spreading subtype, younger patient age, thinner tumor, intermittent solar exposure, and early stage were significant predictors of a melanoma being histologically associated with a nevus. The appearance of a nevus associated with a melanoma predicted better recurrence-free survival compared with de novo melanomas. Although acral lentiginous melanomas (70.9%) constituted the most common histologic subtype, only 9.6% of the acral lentiginous melanomas were associated with a nevus. Furthermore, there was no statistically significant difference between the nevus-associated and de novo acral lentiginous melanomas with regard to clinicopathological factors and survival. In conclusion, nevus-associated melanomas were uncommon among acral lentiginous melanomas. Relatedly, because over half of all melanomas in Asians are acral lentiginous melanomas, Asians are less likely than Caucasians to have nevus-associated melanomas.

Segmentation of nucleus and cytoplasm of a single cell in three-dimensional tomogram using optical coherence tomography.

A random rayburst sampling (RRBS) framework was developed to detect the nucleus and cell membrane boundaries in three-dimensional (3-D) space. Raw images were acquired through a full-field optical coherence tomography system with submicron resolution­i.e., 0.8 ?? ? m in lateral and 0.9 ?? ? m in axial directions. The near-isometric resolution enables 3-D segmentation of a nucleus and cell membrane for determining the volumetric nuclear-to-cytoplasmic (N/C) ratio of a single cell. The RRBS framework was insensitive to the selection of seeds and image pixel noise. The robustness of the RRBS framework was verified through the convergence of the N/C ratio searching algorithm. The relative standard deviation of the N/C ratio between different randomly selected seed sets was only 2%. This technique is useful for various in vitro assays on single-cell analyses.

Longitudinal Melanonychia: Differences in Etiology Are Associated with Patient Age at Diagnosis.

BACKGROUND: Longitudinal melanonychia (LM) may occur as a result of nail apparatus melanoma. Knowledge of etiology plays an important role in the management of LM. OBJECTIVES: The study is aimed to compare the diagnosis of LM in different age groups. METHODS: We collected 63 cases (45 adults and 18 children) with LM who underwent nail matrix biopsy or excision in a 21-year cohort and assessed their clinicopathological features. RESULTS: Melanomas in adults and children were 40% and none, while nevi accounted for 15.6% in adults and 94.4% in children. There was a statistically significant difference between the average age at diagnosis for melanoma (54.5 ± 13.3 years) and nevus (15.2 ± 18.5 years). Logistic regression related the occurrence of melanoma to older ages with a relative risk of 1.2 compared to nevus, but no cutoffs between age groups could be defined between LM-associated nevus and melanoma. CONCLUSION: The adult group has a significantly higher risk of melanoma, while children with LM show mostly nonmelanoma etiologies. Tissue proof is more warranted in adult cases, and it is needed in selected cases of children with LM.

Differential activation behavior of dermal dendritic cells underlies the strain-specific Th1 responses to single epicutaneous immunization.

BACKGROUND: Epicutaneous immunization with allergens is an important sensitization route for atopic dermatitis. We recently showed in addition to the Th2 response following single epicutaneous immunization, a remarkable Th1 response is induced in B6 mice, but not in BALB/c mice, mimicking the immune response to allergens in human non-atopics and atopics. OBJECTIVE: We investigated the underlying mechanisms driving this differential Th1 response between BALB/c and B6 mice. METHODS: We characterized dermal dendritic cells by flow cytometric analysis. We measured the induced Th1/Th2 responses by measuring the IFN-γ/IL-13 contents of supernatants of antigen reactivation cultures of lymph node cells. RESULTS: We demonstrate that more dermal dendritic cells with higher activation status migrate into draining lymph nodes of B6 mice compared to BALB/c mice. Dermal dendritic cells of B6 mice have a greater ability to capture protein antigen than those of BALB/c mice. Moreover, increasing the activation status or amount of captured antigen in dermal dendritic cells induced a Th1 response in BALB/c mice. Further, differential activation behavior, but not antigen-capturing ability of dermal dendritic cells between BALB/c and B6 mice is dendritic cell-intrinsic. CONCLUSION: These results show that the differential activation behavior of dermal dendritic cells underlies the strain-specific Th1 responses following single epicutaneous immunization. Furthermore, our findings highlight the potential differences between human atopics and non-atopics and provide useful information for the prediction and prevention of atopic diseases.

Secondary neoplasms arising from nevus sebaceus: A retrospective study of 450 cases in Taiwan.

Nevus sebaceus is frequently associated with the development of secondary neoplasms. Incidences of malignant transformation vary among different reports and few data is available regarding Asian populations. We aimed to determine the characteristics of secondary tumors developing from nevus sebaceus in a Taiwanese population and to review the published work. Patients with clinically and histologically confirmed nevus sebaceus were identified from 1992 to 2012 in a medical center. Among the 450 cases of nevus sebaceus, 38 secondary neoplasms were noted, accounting for 8.5% of all cases. Benign tumors represented more than 80% of all tumors. Syringocystadenoma papilliferum (2.7%) was the most common benign tumor, followed by trichoblastoma (1.6%) and trichilemmoma (1.6%) whereas basal cell carcinoma (0.9%) was the most frequent malignant tumor on nevus sebaceus and its clinical features were not typical. All the malignant tumors on nevus sebaceus were noted only in adulthood and the mean age of those with basal cell carcinoma was significantly older than that of trichoblastoma (P = 0.028). Our study concludes that malignant transformation is rare in nevus sebaceus and occurs uniquely in adulthood. On the basis of the findings, prophylactic excision of nevus sebaceus can be elective during childhood but is strongly advocated at puberty due to the increased risk of malignant transformation with time.

An alternatively spliced IL-15 isoform modulates abrasion-induced keratinocyte activation.

In a routine phenotype-driven screen, we identified a point mutation in exon 7 of the IL-15 gene in Pedigree 191 (deficient memory (DM)) of N-ethyl-N-nitrosourea mutagenized mice. The DM epidermis expressed an alternatively spliced IL-15 mRNA isoform, IL-15ΔE7, and a wild-type (WT) IL-15 isoform at comparable levels. Mechanical stimulation of DM skin or DM skin graft transplanted onto the WT host resulted in reduced keratinocyte activation and inhibition of neutrophil infiltration into the dermis, demonstrating that DM keratinocytes produced less inflammatory response to external stimulation. Ectopic expression of IL-15ΔE7 in WT skin prevented abrasion-induced epidermal thickening, blocked the accumulation of nuclear antigen Ki67(+) cells in the basal and the suprabasal cell layers, increased loricrin expression, and also increased keratinocyte CXCL1 and G-CSF production. IL-15ΔE7 also profoundly blocked neutrophil infiltration in SDS- or immiquimod (IMQ)-treated WT skin. Recombinant IL-15ΔE7 failed to activate STAT-5 and its downstream target bcl-2 expression. Our study points to IL-15ΔE7 as a potential therapeutic agent for treating neutrophilia-associated inflammatory skin disorders.

Dermal dendritic cells, but not Langerhans cells, are critical in murine single epicutaneous sensitization.

A murine repeated protein-patch model has been established to study epicutaneous sensitization in atopic dermatitis. This model has shown a predominant Th2 and a weak Th1 response in both BALB/c and C57BL/6 mice. However, Th responses induced in the repeated model are not consistent with the generally accepted theory that BALB/c and C57BL/6 mice are Th2 and Th1 prone and are representatives of human atopy and non-atopy, respectively. In this study, a single protein-patch model was established, which showed in addition to the Th2 response, a remarkable Th1 response in C57BL/6 mice, but not in BALB/c mice. Moreover, using muLangerin-DTR mice, we demonstrated that dermal dendritic cells, but not Langerhans cells, are critical in single epicutaneous sensitization in both strains of mice.

Differential diagnosis of nonmelanoma pigmented skin lesions based on harmonic generation microscopy.

In vivo harmonic generation microscopy (HGM) has been applied successfully in healthy human skin and can achieve a submicron resolution, similar to histopathologic examination, even at a penetration depth up to 270 µm. This study aims to investigate the clinical applicability of HGM imaging for differential diagnosis of nonmelanoma pigmented skin lesions. A total of 42 pigmented skin tumors, including pigmented basal cell carcinoma, melanocytic nevus, and seborrheic keratosis were evaluated by HGM ex vivo or in vivo. Based on the standard histopathologic characteristics, we established the corresponding HGM imaging criteria for each pigmented tumor. Diagnostic performance of HGM for differentiating nonmelanoma pigmented skin tumors was evaluated through the observers' direct general assessment (overall evaluation) or the presence of two imaging criteria with the highest sensitivity and specificity (major criteria evaluation). Our results show that, based on the direct general assessment, the sensitivity is 92% [95% confidence interval (CI): 67 to 97%] and the specificity is 96% (95% CI: 83 to 99%); by major criteria evaluation, 94% sensitivity (95% CI: 70 to 99%) and 100% specificity (95% CI: 87 to 100%) are achieved. Our study indicates that HGM serves as a promising histopathological examination tool for noninvasive differential diagnostics of nonmelanoma pigmented skin tumors.

Concurrent exposure to a dectin-1 agonist suppresses the Th2 response to epicutaneously introduced antigen in mice.

BACKGROUND: Epicutaneous sensitization with protein allergen that induces predominant Th2 responses is an important sensitization route in atopic dermatitis. Fungal components have been shown to modulate Th cell differentiation. However, the effects of fungal components on epicutaneous sensitization are unclear. RESULTS: In this study, we showed that co-administration of curdlan, a dectin-1 agonist, during epicutaneous ovalbumin sensitization of BALB/c mice decreased the IL-5 and IL-13 levels in supernatants of lymph node cell ovalbumin reactivation cultures. Mechanistically, curdlan co-administration decreased IL-4 and IL-1ß expressions in draining lymph nodes. Curdlan co-administration also lower the migration of langerin+ CD103- epidermal Langerhans cells into draining lymph nodes at 96 hours post-sensitization which might be attributed to decreased expressions of IL-18 and IL-1ß in patched skin. Moreover, adoptive transfer of CFSE-labeled transgenic CD4 T cells confirmed that curdlan co-administration decreased the proliferation and IL-4-production of ovalbumin -specific T cells primed by epidermal Langerhans cells. CONCLUSIONS: These results indicated that concurrent exposure to a dectin-1 agonist suppresses the epicutaneously induced Th2 response by modulating the cytokine expression profiles in draining LNs and the migration of epidermal Langerhans cells. These results highlight the effects of fungal components on epicutaneous allergen sensitization in atopic diseases.

Antigen-driven bystander effect accelerates epicutaneous sensitization with a new protein allergen.

Exposure to protein allergen epicutaneously, inducing a Th2-dominant immune response, sensitizes the host to the development of atopic disease. Antigen-driven bystander effect demonstrates that polarized T cells could instruct naïve T cells to differentiate into T cells with similar phenotype. In this study, we aimed to determine the contribution of antigen-driven bystander effect on epicutaneous sensitization with a newly introduced protein allergen. BALB/c mice were immunized intraperitoneally with BSA emulsified in alum, known to induce a Th2 response, three weeks before given BSA and OVA epicutaneously. Lymph node cells from these mice restimulated with OVA secreted higher levels IL-4, IL-5 and IL-13 as compared with cells from mice without BSA immunization. In addition, BALB/c mice immunized subcutaneously with BSA emulsified in complete Freund's adjuvant, known to induce a Th1-predominant response, also induced higher Th1 as well as Th2 cytokine response when restimulated with OVA as compared with mice without immunization. We demonstrated that subcutaneous immunization with BSA in CFA induced Th2 as well as Th1 response. The threshold of epicutaneous sensitization to OVA was also reduced, possibly due to increased expressions of IL-4 and IL-10 in the draining lymph nodes during the early phase of sensitization. In conclusion, antigen-driven bystander effect, whether it is of Th1- or Th2-predominant nature, can accelerate epicutaneous sensitization by a newly introduced protein allergen. These results provide a possible explanation for mono- to poly-sensitization spread commonly observed in atopic children.

Low-energy visible light irradiation modulates immune responses induced by epicutaneous sensitization with protein antigen.

Epicutaneous sensitization has been an important route for protein allergen sensitization in atopic disease. Although the skin is irradiated by sunlight daily, the influence of visible light on epicutaneous sensitization has not been explored. In this study, by using a well-established murine protein-patch model, we show that low-energy visible light (LEVL) irradiation could differentially modulate the predominant Th2 immune response induced by epicutaneous sensitization with protein antigen. When the induced Th2 response was strong, as usually observed in BALB/c mice, LEVL irradiation suppressed the response. In contrast, LEVL irradiation enhanced the weaker Th2 response in C57BL/6 mice. Increased IL-18 and decreased TGF-beta expression in draining lymph nodes after LEVL irradiation was observed in BALB/c mice, but not in C57BL/6 mice. LEVL irradiation also enhanced IL-18 expression in skin and reduced the downregulation of CD24 expression on epidermal Langerhans cells in draining lymph nodes of BALB/c mice. Collectively, these results provide evidence for immunomodulatory effects of LEVL irradiation and will help us develop a useful strategy for prevention of allergen sensitization.