Interaction between IL-33 Gene Polymorphisms and Current Smoking with Susceptibility to Systemic Lupus Erythematosus.

AIMS: This study is aimed at exploring the relation between IL-33 single-nucleotide polymorphisms (SNPs) and the risk of systemic lupus erythematosus (SLE). METHODS: SNPStats (online software) was used to test the Hardy-Weinberg equilibrium in controls. Generalized multifactor dimensionality reduction (GMDR) was adopted to screen the preferable interaction between IL-33 SNPs and current smoking. RESULTS: Logistic regression analysis based on the fundamental data of age, gender, BMI, current smoking, and alcohol drinking showed that both rs1929992-G and rs1891385-C alleles were correlated with an increasing risk of SLE, the ORs (95% CI) of which were 1.62 (1.21-2.05) and 1.64 (1.22-2.10), respectively. One two-locus model (rs1929992×current smoking) had a testing accuracy of 60.11% (P = 0.0010). Through an overall multidimensional model, optimum cross-validation consistency was obtained. The analysis indicated that current smoking status influenced the SLE risk depending on the genotypes at rs1929992. Pairwise LD analysis indicated that haplotype rs1929992G-rs7044343T was statistically related to the elevating risk of SLE (P < 0.05). Those subjects with the G-T haplotype had a higher SLE risk than those with other haplotypes, after correction with factors, including gender, alcohol drinking, age, BMI, and current smoking. CONCLUSIONS: The rs1929992-G and rs1891385-C allele, interaction between the rs1929992 gene and current smoking, and haplotype rs1929992G-rs7044343T were all risk factors of SLE.

Evaluation of the efficacy and tolerance of artemether emulsion for the treatment of papulopustular rosacea: a randomized pilot study.

Objective: To assess the efficacy and safety of artemether emulsion in patients with papulopustular rosacea. Methods: A total of 130 (randomized 1:1) were externally administered either artemether emulsion (1%) or metronidazole emulsion (3%) twice daily for 4 weeks with an open-label 8-week follow-up. The primary endpoints included the proportion of patients who achieved clinical effective responses, as well as erythema and papule and pustule score at week 4. Results: Numerically more patients achieved an effective response at week 4 with artemether emulsion (87.1%) than metronidazole emulsion (80.0%) (p > .05). Patients with artemether emulsion had comparable baseline erythema score (2.45 ± 0.67 versus 2.42 ± 0.70, p = .809) and papule and pustule score (2.11 ± 0.96 versus 2.32 ± 0.83, p = .264), but significantly lower papule and pustule score (0.21 ± 0.52 versus 0.42 ± 0.83, p = .001) and comparable erythema score (0.53 ± 0.88 versus 0.62 ± 0.88, p = .999) compared to patients with metronidazole emulsion at week 4. There was a significantly higher proportion of patients with metronidazole emulsion relapse compared to metronidazole emulsion during the open-label 8-week follow-up period (21.6% versus 2.4%, p < .01). Conclusions: Artemether emulsion improved papulopustular rosacea in the metronidazole emulsion group as early as 4 weeks, but its beneficial effect was maintained through the 8-week follow-up period compared to metronidazole emulsion.

Association of TNFAIP3 and TNIP1 polymorphisms with systemic lupus erythematosus risk: A meta-analysis.

OBJECT: With the development of GWAS, both TNFAIP3 and TNIP1 were revealed to be susceptibility genes of SLE. However, some other studies revealed no association between TNFAIP3, TNIP1 and SLE susceptibility. In order to estimate such association more precisely and systemically, a meta-analysis was conducted. METHOD: Studies on the association between TNFAIP3 rs2230926, TNIP1 rs7708392 and SLE risk were carefully selected via searching 3 databases (Pubmed, Embase, and Web of Science). A fixed- or random-effect model was used according to the heterogeneity, and a subgroup analysis by ethnicity was also performed. RESULTS: 26 studies from 18 articles involving a total of 21,372 patients and 30,165 controls were analyzed for TNFAIP3 rs2230926. A significant association between the minor G allele of TNFAIP3 rs2230926 and SLE risk was found via a random-effect model (OR = 1.643, 95% CI = (1.462, 1.847), p < 0.01). In the subgroup analysis by ethnicity, significant correlations were also found in all Caucasians, Asians, and Africans (OR = 1.675, 95% CI = (1.353, 2.074), p < 0.01; OR = 1.738, 95% CI = (1.557, 1.940), p < 0.01; OR = 1.324, 95% CI = (1.029, 1.704), p < 0.05). As for TNIP1 rs7708392, 21 studies from 12 articles involving 24,716 cases and 32,200 controls were analyzed. A significant association of the minor C allele of TNIP1 rs7708392 and SLE risk was found via a random-effect model (OR = 1.247, 95% CI = (1.175, 1.323), p < 0.01). In the subgroup analysis by ethnicity, significant correlations were found in Caucasians, and Africans (OR = 1.317, 95% CI = (1.239, 1.401), p < 0.01; OR = 1.210, 95% CI = (1.108, 1.322), p < 0.01). However, there was no significant association in Asians (OR = 1.122, 95% CI = (0.953, 1.321), p > 0.05). CONCLUSION: The minor G allele of TNFAIP3 rs2230926 was associated with increased risk of SLE in all Caucasians, Asians, and Africans. The minor C allele of TNIP1 rs7708392 was associated with the increased risk of SLE in Caucasians and Africans, while it was not associated with SLE susceptibility in Asians.

The natural phenolic peperobtusin A induces apoptosis of lymphoma U937 cells via the Caspase dependent and p38 MAPK signaling pathways.

Our previous research found the ethyl acetate extract of Peperomia tetraphylla (EAEPT) inhibited the growth of U937 cells by blocking the cell cycle and prompted apoptosis via the reactive oxygen species (ROS)-medicated mitochondria pathway. While the compounds in EAEPT which possessed the anti-tumor activity were unclear. Peperobtusin A is a phenolic compound, which was isolated from the whole plant of Peperomia tetraphylla. In this work, we found that peperobtusin A had the anti-proliferative effects against human lymphoma U937 cells and induced apoptosis in a dose dependent manner. Peperobtusin A significantly enhanced the formation of intracellular ROS and induced the loss of mitochondrial membrane potential (Δψm). And peperobtusin A could increase the ratio of Bax/Bcl-2, induce the cleavage of Bid, Caspase-3, Caspase-8 and Caspase-9 and enhance the level of P-P38. Moreover, peperobtusin A induced the accumulation of cells at S phase. Through using of inhibitors such as antioxidant NAC, pan-caspase inhibitor Z-VAD-FMK, p38 MAPK specific inhibitor SB203580, we found that intracellular ROS generation, activation of Caspases and p38 MAPK played very important roles in the apoptosis induced by peperobtusin A in U937 cells. Our results indicated that intracellular ROS generation, the Caspase-dependent and p38 MAPK signaling pathways involved in apoptosis induced by peperobtusin A in U937 cells.

The Role of Autophagy in the Resistance to BRAF Inhibition in BRAF-Mutated Melanoma.

Malignant melanoma is the most aggressive and notorious skin cancer, and metastatic disease is associated with very poor long-term survival outcomes. Although metastatic melanoma patients with oncogenic mutations in the BRAF gene initially respond well to the treatment with specific BRAF inhibitors, most of them will eventually develop resistance to this targeted therapy. As a highly conserved catabolic process, autophagy is responsible for the maintenance of cellular homeostasis and cell survival, and is involved in multiple diseases, including cancer. Recent study results have indicated that autophagy might play a decisive role in the resistance to BRAF inhibitors in BRAF-mutated melanomas. In this review, we will discuss how autophagy is up-regulated by BRAF inhibitors, and how autophagy induces the resistance to these agents.

The caspase-1 inhibitor CARD18 is specifically expressed during late differentiation of keratinocytes and its expression is lost in lichen planus.

BACKGROUND: CARD18 contains a caspase recruitment domain (CARD) via which it binds to caspase-1 and thereby inhibits caspase-1-mediated activation of the pro-inflammatory cytokine interleukin (IL)-1ß. OBJECTIVES: To determine the expression profile and the role of CARD18 during differentiation of keratinocytes and to compare the expression of CARD18 in normal skin and in inflammatory skin diseases. METHODS: Human keratinocytes were induced to differentiate in monolayer and in 3D skin equivalent cultures. In some experiments, CARD18-specific siRNAs were used to knock down expression of CARD18. CARD18 mRNA levels were determined by quantitative real-time PCR, and CARD18 protein was detected by Western blot and immunofluorescence analyses. In situ expression was analyzed in skin biopsies obtained from healthy donors and patients with psoriasis and lichen planus. RESULTS: CARD18 mRNA was expressed in the epidermis at more than 100-fold higher levels than in any other human tissue. Within the epidermis, CARD18 was specifically expressed in the granular layer. In vitro CARD18 was strongly upregulated at both mRNA and protein levels in keratinocytes undergoing terminal differentiation. In skin equivalent cultures the expression of CARD18 was efficiently suppressed by siRNAs without impairing stratum corneum formation. Epidermal expression of CARD18 was increased after ultraviolet (UV)B irradiation of skin explants. In skin biopsies of patients with psoriasis no consistent regulation of CARD18 expression was observed, however, in lesional epidermis of patients with lichen planus, CARD18 expression was either greatly diminished or entirely absent whereas in non-lesional areas expression was comparable to normal skin. CONCLUSIONS: Our results identify CARD18 as a differentiation-associated keratinocyte protein that is altered in abundance by UV stress. Its downregulation in lichen planus indicates a potential role in inflammatory reactions of the epidermis in this disease.

The role of autophagy in the pathogenesis of systemic lupus erythematosus.

Autophagy is a highly conserved catabolic process, whereby unwanted cytoplasmic contents are enclosed by the double-membrane autophagosomes and delivered to the lysosomes for degradation. It is responsible for the recycling of nutrients and cellular components, thus playing a pivotal role in maintaining cellular homeostasis as well as cell survival during stress conditions. Perturbations in autophagy are implicated in multiple diseases, such as cancers and neuro-degeneration diseases. Recent studies demonstrate that autophagy may participate in almost every step of immune responses, including pathogen recognition, antigen processing and presentation, immune cell development and function, and immunoregulation. The pathogenesis of some autoimmune diseases, such as multiple sclerosis and Crohn's disease, has been reported to be associated with dysregulated autophagy. Systemic lupus erythematosus (SLE) is a chronic, potentially fatal autoimmune disease, characterized by dysregulation of immune cells and production of autoantibodies that cause widespread tissue and organ damage. The pathogenesis of SLE remains unclear. With several single nucleotide polymorphisms (SNPs) in autophagy-related gene5 (ATG5) being linked to SLE susceptibility, more and more lines of evidence from animal model, cell biology, immunology, and genetics studies show that autophagy contributes to the occurrence, development, and severity of SLE.

Paeoniflorin attenuates ultraviolet B-induced apoptosis in human keratinocytes by inhibiting the ROS-p38-p53 pathway.

Ultraviolet (UV) light is one of the most harmful environmental factors that contribute to skin damage. Exposure to UV induces extensive generation of reactive oxygen species (ROS), and results in photoaging and skin cancer development. One approach to protecting human skin against UV radiation is the use of antioxidants. In recent years, naturally occurring herbal compounds have gained considerable attention as protective agents for UV exposure. Paeoniflorin (PF) is a novel natural antioxidant, which is isolated from peony root (Radix Paeoniae Alba). The present study evaluated the protective effects of PF on UV­induced skin damage in vitro, and demonstrated that the effects were mediated via the ROS­p38­p53 pathway. The results of the present study demonstrated that treatment with PF (25, 50, and 100 µM) significantly increased the percentage of viable keratinocytes after UV­B exposure. In addition, cell death analysis indicated that PF treatment markedly reduced UV­B­radiation­induced apoptosis in keratinocytes, which was accompanied by increased procaspase 3 expression and decreased cleaved caspase 3 expression. Treatment with PF markedly reduced the production of ROS, and inhibited the activation of p38 and p53 in human keratinocytes, thus suggesting that the ROS­p38­p53 pathway has a role in UV­B­induced skin damage. In conclusion, the present study reported that PF was able to attenuate UV­B­induced cell damage in human keratinocytes. Notably, these effects were shown to be mediated, at least in part, via inhibition of the ROS-p38-p53 pathway.

Microarray-based identification of differentially expressed genes in extramammary Paget's disease.

Extramammary Paget's disease (EMPD) is a rare cutaneous malignancy accounting for approximately 1-2% of vulvar cancers. The rarity of this disease has caused difficulties in characterization and the molecular mechanism underlying EMPD development remains largely unclear. Here we used microarray analysis to identify differentially expressed genes in EMPD of the scrotum comparing with normal epithelium from healthy donors. Agilent single-channel microarray was used to compare the gene expression between 6 EMPD specimens and 6 normal scrotum epithelium samples. A total of 799 up-regulated genes and 723 down-regulated genes were identified in EMPD tissues. Real-time PCR was conducted to verify the differential expression of some representative genes, including ERBB4, TCF3, PAPSS2, PIK3R3, PRLR, SULT1A1, TCF7L1, and CREB3L4. Generally, the real-time PCR results were consistent with microarray data, and the expression of ERBB4, PRLR, TCF3, PIK3R3, SULT1A1, and TCF7L1 was significantly overexpressed in EMPD (P<0.05). Moreover, the overexpression of PRLR in EMPD, a receptor for the anterior pituitary hormone prolactin (PRL), was confirmed by immunohistochemistry. These data demonstrate that the differentially expressed genes from the microarray-based identification are tightly associated with EMPD occurrence.

Vitamin C protects against UV irradiation-induced apoptosis through reactivating silenced tumor suppressor genes p21 and p16 in a Tet-dependent DNA demethylation manner in human skin cancer cells.

AIM: DNA methylation plays important roles in various kinds of carcinogenesis. Vitamin C could induce Tet-dependent DNA demethylation in embryonic stem cells. Therefore, the antagonizing activity of vitamin C on ultraviolet (UV)-induced apoptosis was investigated in this study. METHODS: Apoptosis of human epidermoid carcinoma A431 cells and p16-knockout (KO) or p21-KO fibroblasts was assessed by a fluorescence-activated cell sorter. Real-time PCR and western blot were used to determine the relative expression levels of p12, p21, and Tet1/2/3 genes. The global DNA methylation levels were determined using MethylFlash Methylated DNA Quantification Kit in A431 cells with or without vitamin C treatment. To examine the DNA demethylation activity of vitamin C, DNA immunoprecipitation (DIP)-qPCR was performed to determine the relative levels of 5-methylcytosine (5mC) or 5-hydroxymethylcytosine (5hmC) in p16 and p21 promoter regions containing cytosine-phosphorothiolated guanine (CpG) islands. RESULTS: The increasing apoptosis of A431 cells under prolonged UV irradiation was remarkably decreased by the combination of vitamin C treatment, suggesting that vitamin C protects against UV-induced apoptosis. Concurrently, vitamin C induced a significant reduction of global DNA methylation in a time- and dose-dependent manner in A431 cells. Vitamin C also reactivated the expression of p16 and p21 at mRNA and protein levels. Mechanistically, about 27% 5hmC-positive cells were observed in vitamin C-treated A431 cells, and the 5hmC enrichment at p16 and p21 promoter regions was also largely increased by vitamin C. Moreover, the expression of p16 and p21 was decreased in Tet1/2 double-knockdown cells, in which the inhibitory effect of vitamin C on UV-induced apoptosis was dismissed. Furthermore, the inhibition of UV-induced apoptosis on vitamin C treatment nearly disappeared in p16- or p21-knockout primary cultured fibroblasts. CONCLUSION: These results demonstrate that vitamin C effectively antagonizes UV-induced apoptosis through regulation of Tet activity, DNA demethylation, and subsequent tumor suppressor gene activation in skin cancer cells.

Icariside II induces apoptosis via inhibition of the EGFR pathways in A431 human epidermoid carcinoma cells.

Improvements in skin cancer treatment are likely to derive from novel agents targeting the molecular pathways that promote tumor cell growth and survival. Icariside II (IS) is a metabolite of icariin, which is derived from Herba Epimedii. The aim of the present study was to evaluate the antitumor effects of IS and to determine the mechanism of apoptosis in A431 human epidermoid carcinoma cells. A431 cells were treated with IS (0­100 µM) for 24 or 48 h and cell viability was detected using the WST­8 assay. Apoptosis was measured by the Annexin­V/propidium iodide (PI) flow cytometric assay. Western blot analysis was used to measure the expression of cleaved caspase­9, cleaved poly ADP ribose polymerase (PARP), phosphorylated signal transducer and activator of transcription 3 (P­STAT3), phosphorylated extracellular signal-regulated kinase (P­ERK), and P­AKT. A431 cells were also pretreated with IS (0­100 µM) 2 h prior to treatment with epidermal growth factor (EGF; 100 ng/ml) for 10 min. Phosphorylated EGF receptor (P­EGFR), P­STAT3, P­ERK and P­AKT were detected by western blot analysis. The results demonstrated that IS inhibited the cell viability of the A431 cells in a dose­dependent manner. Pretreatment with LY294002 [a phosphatidylinositol 3-kinase (PI3K) inhibitor], EGF (an EGFR agonist) and AG1478 (an EGFR inhibitor) partially reversed IS­induced decreases in cell viability. Treatment with 50 µm IS resulted in an increased number of apoptotic cells mirrored by increases in cleaved caspase­9 and cleaved PARP. In addition, treatment with 50 µM IS significantly inhibited the activation of the Janus kinase (JAK)­STAT3 and mitogen­activated protein kinase (MAPK)­ERK pathways, but promoted the activation of the PI3K­AKT pathway. Furthermore, IS effectively inhibited the EGF-induced activation of the EGFR pathways. In conclusion, IS inhibited the cell viability of the A431 cells through the regulation of apoptosis. These effects were mediated, at least in part, by inhibiting the activation of the EGFR pathways.

MicroRNA-29b contributes to DNA hypomethylation of CD4+ T cells in systemic lupus erythematosus by indirectly targeting DNA methyltransferase 1.

BACKGROUND: The mechanism of DNA hypomethylation in systemic lupus erythematosus (SLE) has not been fully elucidated. Recent studies showed that miR-29b could regulate DNA methylation by targeting the DNA methylation machinery. However, the role of miR-29b in T cell aberrant DNA hypomethylation of SLE still remains unclear. OBJECTIVE: In this study, we asked whether miR-29b regulate DNA methylation in lupus CD4+ T cells. METHODS: The miR-29b expression was analyzed by quantitative polymerase chain reaction (qPCR). Sp1, DNMT1, CD11a and CD70 mRNA and protein levels were determined by qPCR, Western-blotting and flow cytometry, respectively. The global DNA methylation levels were evaluated by the Methyflash™ DNA Methylation Quantification Kit. CD11a and CD70 promoter methyaltion levels were detected by bisulfate modification and methylation-sensitive high resolution melting analysis. RESULTS: In SLE patients, the miR-29b levels were up-regulated as compared to healthy donors and its degree of overexpression was negatively correlated with sp1 and DNMT1 protein levels, respectively. Overexpression of miR-29b resulted in significant reduction of sp1 and DNMT1 expression. Further analysis demonstrated that overexpression of miR-29b in CD4+ T cells from healthy donors led to the DNA hypomethylation and up-regulation of genes encoding CD11a and CD70, and inhibition of miR-29b expression in CD4+ T cells from patients with lupus caused reverse effects. CONCLUSION: Our study suggests that miR-29b negatively regulates DNMT1 expression by targeting sp1 in T cells. The overexpression of miR-29b contributes to the reduction of DNMT1 levels and thereby DNA hypomethylation in SLE. This finding provides potential novel strategies for therapeutic interventions.

Preventive effects of zinc against psychological stress-induced iron dyshomeostasis, erythropoiesis inhibition, and oxidative stress status in rats.

Psychological stress (PS) could cause decreased iron absorption and iron redistribution in body resulting in low iron concentration in the bone marrow and inhibition of erythropoiesis. In the present study, we investigated the effect of zinc supplementation on the iron metabolism, erythropoiesis, and oxidative stress status in PS-induced rats. Thirty-two rats were divided into two groups randomly: control group and zinc supplementation group. Each group was subdivided into two subgroups: control group and PS group. Rats received zinc supplementation before PS exposure established by a communication box. We investigated the serum corticosterone (CORT) level; iron apparent absorption; iron contents in liver, spleen, cortex, hippocampus, striatum, and serum; hematological parameters; malondialdehyde (MDA); reduced glutathione (GSH); and superoxide dismutase (SOD). Compared to PS-treated rats with normal diet, the PS-treated rats with zinc supplementation showed increased iron apparent absorption, serum iron, hemoglobin, red blood cell, GSH, and SOD activities; while the serum CORT; iron contents in liver, spleen, and regional brain; and MDA decreased. These results indicated that dietary zinc supplementation had preventive effects against PS-induced iron dyshomeostasis, erythropoiesis inhibition, and oxidative stress status in rats.

Concordant correlation of LIV-1 and E-cadherin expression in human breast cancer cell MCF-7.

The recent report highlighted a significant association between signal transducer and activator of transcription 3 (STAT3) and Snail and LIV-1 (SLC39A6 or ZIP6), the breast cancer-associated protein that belongs to a new subfamily of zinc transporters. LIV-1 is a downstream target of STAT3, both in zebrafish and mammalian cells and provides control over epithelial-mesenchymal transition (EMT). Crucially, these observations link LIV-1, previously demonstrated to be associated with lymph node metastasis in breast cancer, to genes with a proven role in development. A putative role of LIV-1 as a regulator of E-cadherin that modulates the cell-cell adhesion is thus inferred. In present study, the correlation of LIV-1 and E-cadherin expression in human breast cancer cell MCF-7 and the effect of LIV-1 expression on the cell growth were assessed to explore the possible mechanisms associated with this observation in breast cancer. It was shown that the silencing of LIV-1 would induce the down-expression of E-cadherin. There was opposite results if the cells were overexpressed with LIV-1. In addition, the results showed that promotion effect after silencing of LIV-1 and inhibition effect after overexpression of LIV-1 in transfected cells. To our knowledge, this is the first evidence that the expression of E-cadherin could be regulated by the zinc transporter LIV-1. The results suggest that there is an association of LIV-1 expression with less aggressive tumors due to high E-cadherin expression because of high LIV-1 expression. LIV-1 may be a regulator of E-cadherin.

Cooperation of metallothionein and zinc transporters for regulating zinc homeostasis in human intestinal Caco-2 cells.

This investigation examined the effects of zinc status on cell proliferation and the synergic roles of the metallothionein (MT) and zinc transporter (ZnT) in the human colon adenocarcinoma cell line Caco-2. Cells were treated with 0 to 300 micromol/L ZnSO(4) or 0 to 10 micromol/L N,N,N',N'-tetrakis(2-phridylmethyl) ethylenediamine (TPEN). Cell proliferation was determined by MTT assay and apoptotic cells detected by flow cytometry (Hoechst 33258 dye). mRNA expression of MT1; ZnT1; zrt, irt-like protein 1, 4 (ZIP1, 4); and divalent metal transporter (DMT1) were determined by the reverse transcription polymerase chain reaction or real-time polymerase chain reaction. The results showed that either high or low zinc could inhibit the cell proliferation. The number of apoptotic cells increased with incremental increases in the concentrations of ZnSO(4) and TPEN. The mRNA expression of ZnT1 and MT1 responded significantly after 6 and 12 hours with 200 micromol/L zinc treatment, respectively, and increased gradually with zinc levels from 0 to 200 micromol/L. Compared with the unchanged ZIP1 mRNA expression, ZIP4 was closely dependent on TPEN treatment duration and concentration. The DMT1 mRNA expression was upregulated time-dependently but not concentration-dependently in the late TPEN treatment duration. The results suggest that ZIP4 and DMT1 mRNA expressions are susceptible to low extracellular zinc concentration and upregulated to enhance zinc absorption, whereas the ZnT1 and MT1 act as the key regulators under high zinc conditions to enhance the intracellular zinc efflux to maintain zinc homeostasis. We propose that in response to variations in zinc concentration, the cooperated regulative roles of ZnT1, MT1, DMT1, and ZIP4 contribute to zinc homeostasis.