Yuling Shi

Systemic analyses of immunophenotypes of peripheral T cells in non-segmental vitiligo: implication of defective natural killer T cells.

Although it is widely believed that non‐segmental vitiligo (NSV) results from the autoimmune destruction of melanocytes, a clear understanding of defects in immune tolerance, which mediate this uncontrolled self‐reactivity, is still lacking. In the present study, we systemically evaluated circulating regulatory T (Treg) cells, including CD4+CD25+FoxP3+ Treg cells and invariant natural killer T (iNKT) cells, as well as naïve and memory CD4+ and CD8+ T cells and their cytokine production, in a cohort of 43 progressive NSV patients with race‐, gender‐, and age‐matched healthy controls. We found that the general immunophenotypes of CD4+ and CD8+ T cells and the percentage of CD4+CD25+FoxP3+ Tregs were comparable between NSV and healthy controls. However, percentages of peripheral iNKT cells were significantly decreased in NSV patients compared to that in healthy controls. Our data confirm the previous notion that the percentage of peripheral CD4+CD25+FoxP3+ Tregs remains unaltered in NSV and suggests the involvement of defective iNKT cells in the pathogenesis of NSV.

Lack of microRNA miR-150 reduces the capacity of epidermal Langerhans cell cross-presentation.

MicroRNAs (miRNAs) are evolutionarily conserved small non‐coding RNAs that repress target genes at post‐transcriptional level. Langerhans cells (LCs) are skin‐residential dendritic cells (DCs) with a life cycle distinct from other types of DCs. miRNA deficiency interrupts the homoeostasis and function of epidermal LCs, suggesting the critical roles of miRNAs in LC development and function. However, the roles of individual miRNAs in regulating LC development and function remain completely unknown. MiRNA miR‐150 is highly expressed in mature lymphocytes and regulates T‐ and B‐cell development and function. Here, we reported that miR‐150 is also expressed in epidermal LCs, and its expression is significantly down‐regulated during in vitro LC maturation. Using a miR‐150 knockout mouse model, we found that lack of miR‐150 reduces the capacity of LCs to cross‐present a soluble antigen to antigen‐specific CD8+ T cells, but does not disturb the development, maturation, migration and phagocytic capacity of LCs. Thus, our data indicate that miR‐150 is required for LC cross‐presentation.

MicroRNA expression profiling identifies potential serum biomarkers for non-segmental vitiligo

To take out a personal subscription, please click here More information about Pigment Cell & Melanoma Research at www.pigment.org MicroRNA expression profiling identifies potential serum biomarkers for non-segmental vitiligo Yu-Ling Shi, Matthew Weiland, Jia Li, Iltefat Hamzavi, Marsha Henderson, Richard H. Huggins, Bassel H. Mahmoud, Oma Agbai, Xiaofan Mi, Zheng Dong, Henry W. Lim1,2, Qing-Sheng Mi and Li Zhou

Increased circulating Th17 cells and elevated serum levels of TGF-beta and IL-21 are correlated with human non-segmental vitiligo development

Although non‐segmental vitiligo (NSV) results from the autoimmune destruction of melanocytes, the detailed immune mechanisms have not yet been fully elucidated. Th17 cells have been identified to be implicated in human autoimmune diseases. In this study, the frequencies of peripheral blood Th17 cells and serum levels of IL‐17A and Th17 cell‐related cytokines were examined in 45 patients with active NSV compared to 45 race‐, gender‐, and age‐matched healthy controls. Our results showed increased circulating Th17 cell frequencies and elevated serum IL‐17A, TGF‐β1, and IL‐21 levels in patients with NSV. Meanwhile, the increased Th17 cell frequencies are positively correlated with serum TGF‐β1 level, and the body surface area of lesions is positively correlated with elevated TGF‐β1 and IL‐21 levels and Th17 cell frequencies. Furthermore, positive correlation was identified between Th17 and Th1 cell frequencies in patients with NSV. These results further indicate the potential involvement of Th17 cells and the collaborative contribution of Th17 and Th1 in NSV development, and suggest that the elevated serum TGF‐β1 and IL‐21 levels could contribute to enhanced Th17 cell differentiation in NSV.

The tumor necrosis factor receptor superfamily member 1B polymorphisms predict response to anti-TNF therapy in patients with autoimmune disease: A meta-analysis

Numerous published data on the tumor necrosis factor receptor superfamily member 1B (TNFRSF1B) gene polymorphisms are shown to be associated with response or non-response to anti-TNF therapy in autoimmune diseases such as rheumatoid arthritis (RA), psoriasis and Crohns Disease (CD). The aim of this study is to investigate whether the TNFRSF1B rs1061622 T/G or TNFRSF1A A/G rs767455 polymorphisms can predict the response to anti-TNF-based therapy in patients with autoimmune diseases. We conducted a meta-analysis of studies on the association between TNFRSF1B rs1061622 T/G polymorphism or TNFRSF1A A/G rs767455 polymorphism and non-responsiveness to anti-TNF therapy in autoimmune diseases. A total of 8 studies involving 929 subjects for TNFRSF1B rs1061622 and 564 subjects for TNFRSF1A rs767455 were finally considered. These studies consisted of seven studies on the TNFRSF1B polymorphism and four studies on the TNFRSF1A polymorphism. Meta-analysis showed significant association between the TNFRSF1B rs1061622 allele and non-responders to anti-TNF therapy [T/G odds ratio (OR) 0.72, 95% confidence interval (CI) 0.57-0.93, p=0.01]. Stratification by disease type indicated an association between the TNFRSF1B rs1061622 allele and non-responders to TNF antagonist in RA (T/G OR 0.69, 95% CI 0.48-0.99, p<0.05) and psoriasis (T/G OR 0.39, 95% CI 0.23-0.67, p<0.001), but not in CD (T/G OR 1.14, 95% CI 0.57-0.93, p=0.57). And there was no association between TNFRSF1A rs767455 genotype and non-responders to the anti-TNF therapy (A/G OR 0.93, 95% CI 0.70-1.23, p=0.59). This meta-analysis demonstrates that TNFRSF1B T allele carriers show a better response to anti-TNF therapy, and individuals carrying TNFRSF1A A allele have no relationship with the response to anti-TNF therapy for autoimmune diseases. The genotyping of this polymorphism could help to optimize the treatment by identifying patients with a likely poor response to biological drugs.

Naringin protects ultraviolet B-induced skin damage by regulating p38 MAPK signal pathway

BACKGROUND Naringin is a bioflavonoid and has free radical scavenging and anti-inflammatory properties. METHODS We examined the effects of naringin on skin after ultraviolet radiation B (UVB) irradiation and the signal pathways by in vitro and in vivo assay. RESULTS HaCaT cells pretreated with naringin significantly inhibited UVB induced-cell apoptosis and production of intracellular reactive oxygen species (ROS). The expressions of interleukin-1β (IL-1β), interleukin-6 (IL-6), interleukin-8 (IL-8) and cyclooxygenase-2 (COX-2) in HaCaT cells pretreated with naringin were decreased compared with the only UVB group. Also, the activation of p38 induced by UVB in HaCaT cells was reversed by naringin treatments. The inhibition function of naringin on p38 activity was more obvious than JNK. In vivo, topical treatments with naringin prevented the increase of epidermal thickness, IL-6 production, cell apoptosis and the overexpression of COX-2 in BALB/c mice skin irradiated with UVB. Naringin treatment also markedly blocked the activation of p38 in response to UVB stimulation in the mouse skin. CONCLUSION Naringin can effectively protect against UVB-induced keratinocyte apoptosis and skin damage by inhibiting ROS production, COX-2 overexpression and strong inflammation reactions. It seemed that naringin played its role against UVB-induced skin damage through inhibition of mitogen-activated protein kinase (MAPK)/p38 activation.

Histone deacetylases inhibitor Trichostatin A ameliorates DNFB-induced allergic contact dermatitis and reduces epidermal Langerhans cells in mice

BACKGROUND Histone deacetylases (HDACs) influence chromatin organization, representing a key epigenetic regulatory mechanism in cells. Trichostatin A (TSA), a potent HDAC inhibitor, has anti-tumor and anti-inflammatory effects. Allergic contact dermatitis (ACD) is a T-cell-mediated inflammatory reaction in skin and is regulated by epidermal Langerhans cells (LCs). OBJECTIVE The aim of this study was to investigate if TSA treatment prevents 2,4-dinitrofluorobenzene (DNFB)-induced ACD in mice and regulates epidermal LCs and other immune cells during ACD development. METHODS ACD was induced by sensitizing and challenging with DNFB topically. Mice were treated intraperitoneally with TSA or vehicle DMSO as a control every other day before and during induction of ACD. The ear swelling response was measured and skin biopsies from sensitized skin areas were obtained for histology. Epidermal cells, thymus, spleen and skin draining lymph nodes were collected for immune staining. RESULTS TSA treatment ameliorated skin lesion severity of DNFB-induced ACD. The percentages of epidermal LCs and splenic DCs as well as LC maturation were significantly reduced in TSA-treated mice. However, TSA treatment did not significantly affect the homeostasis of conventional CD4(+) and CD8(+) T cells, Foxp3(+)CD4(+) regulatory T cells, iNKT cells, and γδ T cells in thymus, spleen and draining lymph nodes (dLNs). Furthermore, there were no significant differences in IL-4 and IFN-γ-producing T cells and iNKT cells between TSA- and DMSO-treated mice. CONCLUSION Our findings suggest that TSA may ameliorate ACD through the regulation of epidermal LCs and HDACs could serve as potential therapeutic targets for ACD and other LCs-related skin diseases.

TGFβ/Smad3 signal pathway is not required for epidermal langerhans cell development

TO THE EDITOR Epidermal Langerhans cells (LCs) are professional antigen-presenting dendritic cells (DCs) that reside in the epidermis and form the first immunological barrier to the external environment (Romani et al., 2010). Although considerable progress has been made in identifying the developmental requirement of LCs, the mechanisms that regulate LC development and homeostasis are still not fully elucidated. TGF-β1 is one of key regulators to control LC development and homeostasis. LCs are absent in mice that lack TGF-β1, owing to a failure in LC differentiation, survival or both (Borkowski et al., 1996; Kaplan et al., 2007; Zahner et al., 2011). This finding is further supported by the dependence of LCs development on other TGFβ1-related molecules, including inhibitor of DNA binding 2 (ID2), a TGFβ1-induced inhibitor of helix–loop–helix transcription factors (Hacker et al., 2003), and Runx3, specifically expressed by mature DCs and mediates their response to TGFβ1 (Fainaru et al., 2004). Furthermore, TGF-β1 is also required to maintain the pool of immature LCs in the epidermis (Kel et al., 2010). Although TGF-β1 is expressed by both keratinocytes and LCs, an autocrine source of TGF-β1 from LCs is required for LC development (Kaplan et al., 2007). Taken together, TGF-β1 is essential for the ontogeny, homeostasis, and function of epidermal LCs. However, the intracellular signaling pathway of TGF-β1 in epidermal LCs remains elusive. In a well-defined classical TGF-β linear signaling pathway, once activated, TGF-β1 signals through its two cell surface receptors, TGF-β receptor 1 (TβR1) and TGF-β receptor 2 (TβR2), leading to Smad-mediated transcriptional regulation. There are eight Smads: Smad1 to Smad8. Smad2 and Smad3 are activated through carboxy-terminal phosphorylation by TGFβ1. These receptor-activated Smads (R-Smads) are released from the receptor complex to form a heterotrimeric complex with a common Smad4, and translocate into the nucleus to regulate the transcription of target genes (Derynck and Zhang, 2003; Tsukazaki et al., 1998). In addition, TGF-β1 also can activate other signaling cascades, including MAPK pathways (Itoh et al., 2000; Massague, 2000; Moustakas et al., 2001). Unlike mice with the conventional disruptions of Smad2 and Smad4 that are lethal, mice with Smad3 deficiency are viable and can survive. However, Smad3 knockout (KO) mice develop progressive diseases, including leucocytosis and massive inflammation. The loss of Smad3 results in multiple cell defects, including T cells, neutriphils and macrophage (Werner et al., 2000; Yang et al., 1999). Here we used Smad3 KO mice (Datto et al., 1999) to directly test if TGFβ/Smad3 signaling pathway is involved in the ontogeny and homeostasis of epidermal LCs. During LC early development, a single wave of LC precursors recruited in the epidermis around embryonic day 18, which subsequently differentiate into LCs after birth, and LCs then undergo a massive burst proliferation during the first week of life to form a typical LC network (Chorro et al., 2009). Given that TGF-β signals are required for ontogeny and maintaining of LCs in the epidermis, we first assessed if Smad3 is required for LC homeostasis in vivo. Epidermal cell suspensions prepared from ears and dorsal skin of Smad3KO mice and WT littermates, on day 0, 1, 3, 8 after birth, as well as at 3 and 5-week old, were stained with anti-Langerin and anti-MHC-II antibodies. As shown in Figure 1 a & b, we did observe a 3–4-fold expansion of LCs from 0.78 ± 0.11% on day 0 to 2.72 ± 0.14% on day 3 in WT mice. Langerin expression was undetectable in MHCII+ LCs in the epidermis on day 0 and 1, but upregulated Langerin expression was detected in about 70%–80% LCs by day 3, which is consistent with recent report (Chorro et al., 2009; Kel et al., 2010). However, there were no significant differences on the rations of epidermal LCs between Smad3 KO and WT mice at any time points by flow cytometry (Figure 1a & b) and immunohistological staining at day 0 after born and 5 weeks old (Figure 1c & d). Thus, Smad3 is not required in the TGF-β signal pathway for oncogeny and homeostasis of epidermis LCs. Figure 1 Smad3 is not required for ontogeny and homeostasis of epidermis LCs. (a) and (b) Epidermal suspensions prepared from ears and trunk skin of Smad3 KO and WT littermates on day 0, 1, 3, 8 after birth, as well as at 3 and 5 weeks old, were stained with anti-Langerin ... Upon activation by various stimuli, immature LCs residing in epidermis collect antigen and increase their MHC II and costimulatory molecules, including CD80 and CD86, and then migrate to T cells areas of draining lymph nodes, leading to immune responses (Romani et al., 2010). However, loss of TβR1 in LCs makes LC spontaneous maturation, suggesting the TGF-β signal as an essential pathway to control the immature state of LCs (Kel et al., 2010). We next tested if TGF-β/Smad3 signaling pathway is required for maintaining the immature state of epidermal LCs. As shown in Figure 2a, the frequencies of CD80+ LCs and CD86+LCs were comparable between Smad3 KO and WT mice. Furthermore, the expression levels of MHCII, CD80 and CD86 on LCs, represented by mean fluorescence intensity (MFI), were not changed (Figure 2b). LC maturation in Smad3 KO mice were further evaluated after in vitro epidermal culture. As shown in Figure 2c, after 60h culture, both the percentages of CD86- and CD80-positive LCs and MHCIIhigh LCs as well as the relative CD80, CD86, and MHCII expression levels on LCs (Figure 2d) were significantly increased in Smad3KO and WT mice compared to unstimulation condition (Figure 2a & b), but there was no significant difference between Smad3 KO and WT mice (P > 0.05). Thus, the lack of Smad3 signaling does not affect the immature state of LCs as well as LC maturation. Figure 2 Smad3 is not an essential factor to control epidermis LC maturation and LC uptaking-antigen ability. (a) and (b) Epidermal suspensions freshly isolated from ears and trunk skin of Smad3 KO and WT littermates at 5 weeks old, were stained with anti-Langerin, ... Due to their physical location, LCs acquire and process antigens. To evaluate the role of Smad3 in antigen phagocytic function of LCs, freshly isolated epidermal cells from KO and WT mice were incubated at 37°C or 4°C (as control) with FITC-Dextran for 45 minutes and then stained with anti-mouse MHC-II and CD45.2 antibodies. As shown in Figure 2e, the phagocytic capacity of LCs in Smad3 KO mice had no significant difference (P > 0.05) compared to the WT LCs, based on the ratio of FITC-positive LCs (Figure 2e) or MFI expression levels (Figure 2f). Thus, lack of Smad3 signal pathway does not affect LC phagocytosis. In summary, lack of Smad3 surprisingly does not significantly interrupt the development and immature state of epidermal LCs, and Smad3-deficient LCs have normal maturation and phagocytosis. Our data suggest that Smad3 is not required in the TGF-β signal pathway for ontogeny, homeostasis, and function of epidermal LCs. Recent report indicated that Smad2 and Smad3 were redundantly essential for TGF-β–mediated induction of Foxp3-expressing regulatory T cells and suppression of IFN-γ production in CD4+ T cells (Takimoto et al., 2010). This raises the possibility that Smad2/Smad3 redundancy may also exist in TGF-β/Smads pathways in LCs. In addition, non-TGF-β/Smads pathways may also regulate LC ontogeny and homeostasis (Figure 2g). Further investigations are warranted to clarify the TGF-β signaling pathways through which TGF-β controls LC ontogeny and homeostasis.

Daidzein stimulates collagen synthesis by activating the TGF-β/smad signal pathway

The objective of this study was to investigate the effects of daidzein on collagen metabolism and its underlying mechanism in cultured skin fibroblast and nude mouse skin.

Serum miRNA expression profiles change in autoimmune vitiligo in mice

It is widely believed that non‐segmental vitiligo results from the autoimmune destruction of melanocytes. MicroRNAs (miRNAs), a class of small non‐coding RNAs that negatively regulate gene expression, are involved in the immune cell development and function and regulate the development of autoimmune diseases. Recent studies demonstrate that functional miRNAs can be detected in the serum and serve as biomarkers of various diseases. In the present study, we used a mouse autoimmune vitiligo model, in which melanocyte autoreactive CD4+ T cells were adoptively transferred into Rag1−/− host mice. Serum miRNA expression was profiled in vitiligo developed mice and control mice using TaqMan RT‐PCR arrays. We have found that the expressions of 20 serum miRNAs were changed in vitiligo mice compared to control mice. Three increased miRNAs, miR‐146a, miR‐191, and miR‐342‐3p, were further confirmed by a single TaqMan RT‐PCR. Our findings suggest that miRNAs may be involved in vitiligo development and serum miRNAs could serve as serum biomarkers for vitiligo in mice.