The investigation of the amounts and expressions of epidermal growth factor, epidermal growth factor receptor, and epidermal growth factor receptor gene polymorphisms in acne vulgaris.

BACKGROUND: Epidermal growth factor receptor inhibitors (EGFRI) used in cancer chemotherapy cause acneiform folliculitis in 70%-100% of patients in a dose-dependent manner. Acneiform folliculitis is considered to be caused by an inflammatory process due to follicular hyperkeratosis and subsequently a set of changes both in epidermis and hair follicles as a result of epidermal growth factor receptor (EGFR) blockade. Both acne vulgaris and acneiform folliculitis due to EGFRIs show similar changes in the pilosebaceous unit. Furthermore, in both groups of patients, topical application of recombinant human epidermal growth factor (EGF) has been reported to improve the disease. AIMS: In this study, it was aimed to investigate the role of EGF and EGFR amount, expression, and EGFR gene polymorphisms in the etiopathogenesis of acne vulgaris. PATIENTS/METHODS: 156 acne vulgaris patients, within 18-25 years of age, who had 15 or more inflammatory acne lesions on dermatologic evaluation were included in this study. The absence of any known systemic or genetic disease or cancer and any systemic or topical treatment for the last 1 month were prerequisites. In the control group, 154 volunteers in the same age range who were examined at the outpatient clinic with diagnoses of melanocytic nevus, ephelid, cherry angioma, and callus and who had no more than 3 inflammatory acne lesions were recruited. The amounts of EGF and EGFR were determined by sandwich ELISA, expressions of EGF and EGFR by reverse transcriptase polymerase chain reaction; EGFR polymorphisms were examined by restriction enzyme digestion, Sanger, and high-resolution melting methods. RESULTS: The patient and control groups were compared in terms of EGFR gene polymorphisms in addition to the amount and expressions of EGF and EGFR. The amount of EGF in the serum was found to be significantly higher in the acne group. (P = .0012). There was no significant difference in other parameters studied. CONCLUSION: The results of our study showed a significant increase in the amount of EGF in the acne group. Though EGF may be incriminated in the etiopathogenesis of AV, the most likely explanation about its role may be controlling inflammation from the very first stage.

Comparison of Dendritic Cell Activation by Virus-Based Vaccine Delivery Vectors Emphasizes the Transcriptional Downregulation of the Oxidative Phosphorylation Pathway.

Antigen delivery platforms based on engineered viruses or virus-like particles are currently developed as vaccines against infectious diseases. As the interaction of vaccines with dendritic cells (DCs) shapes the immunological response, we compared the interaction of a range of virus-based vectors and virus-like particles with DCs in a murine model of systemic administration and transcriptome analyses of splenic DCs. The transcriptome profiles of DCs separated the vaccine vectors into two distinct groups characterized by high- and low-magnitude differential gene expression, which strongly correlated with (1) the surface expression of costimulatory molecules CD40, CD83, and CD86 on DCs, and (2) antigen-specific T-cell responses. Pathway analysis using PANOGA (Pathway and Network-Oriented GWAS Analysis) revealed that the JAK/STAT pathway was significantly activated by both groups of vaccines. In contrast, the oxidative phosphorylation pathway was significantly downregulated only by the high-magnitude DC-stimulating vectors. A gene signature including exclusively chemokine-, cytokine-, and receptor-related genes revealed a vector-specific pattern. Overall, this in vivo DC stimulation model demonstrated a strong relationship between the levels of induced DC maturation and the intensity of T-cell-specific immune responses with a distinct cytokine/chemokine profile, metabolic shifting, and cell surface expression of maturation markers. It could represent an important tool for vaccine design.

Murine Bone Marrow-Derived Dendritic Cells Transduced by Light-Helper-Dependent Herpes Simplex Virus-1 Amplicon Vector Acquire a Mature Dendritic Cell Phenotype.

Dendritic cells (DCs) turn into the most potent antigen-presenting cells following a complex transforming process, which leads to their maturation. Herpes simplex virus-1 (HSV-1) amplicon vectors represent highly versatile viral vector platforms with the ability to transduce immature DCs at exceedingly high efficiencies, while the efficiency of infection of mature DCs is significantly low. However, the bacterial artificial chromosome (BAC)-dependent (BD) amplicon vectors tested so far do not result in the maturation of mouse bone marrow-derived DCs (BMDCs) in vitro. In this study we investigated the effects of light-helper-dependent (LHD) amplicon vectors produced with the replication-defective HSV-1 LaLΔJ helper virus system. First, we observed that transgene expression in BMDC cultures was equally potent between the LHD and the BD amplicon vectors. We determined that the percentage of transduced cells and the duration of transgene expression were negatively influenced by the presence of increasing levels of helper virus. Second, infection by the LHD amplicon vector as well as the helper HSV-1 LaLΔJ virus alone resulted in the phenotypic maturation of BMDCs and the expression of both interferon-stimulated genes and proinflammatory cytokines. Further comparisons of the gene expression of infected DCs showed that while interferon-stimulated genes such as Ifit1, Ifit3, Mx2, Isg15, and Cxcl10 were induced by both BD and LHD amplicon vectors, early proinflammatory cytokine gene expression (Tnfa, Il1a, Il1b, Il6, Il10, Il12b, Cxcl1, and Cxcl16) and DC maturation were mediated only by the LHD amplicons.

Cleavage of Atg3 protein by caspase-8 regulates autophagy during receptor-activated cell death.

Autophagy is an evolutionarily conserved mechanism contributing to cell survival under stress conditions including nutrient and growth factor deprivation. Connections and cross-talk between cell death mechanisms and autophagy is under investigation. Here, we describe Atg3, an essential regulatory component of autophagosome biogenesis, as a new substrate of caspase-8 during receptor-mediated cell death. Both, tumor necrosis factor α and tumor necrosis factor-related apoptosis inducing ligand induced cell death was accompanied by Atg3 cleavage and this event was inhibited by a pan-caspase inhibitor (zVAD) or a caspase-8-specific inhibitor (zIETD). Indeed, caspase-8 overexpression led to Atg3 degradation and this event depended on caspase-8 enzymatic activity. Mutation of the caspase-8 cleavage site on Atg3 abolished its cleavage both in vitro and in vivo, demonstrating that Atg3 was a direct target of caspase-8. Autophagy was inactive during apoptosis and blockage of caspases or overexpression of a non-cleavable Atg3 protein reestablished autophagic activity upon death receptor stimulation. In this system, autophagy was important for cell survival since inhibition of autophagy increased cell death. Therefore, Atg3 provides a novel link between apoptosis and autophagy during receptor-activated cell death.

Green fluorescent protein - Tagged HCV non-enveloped capsid like particles: development of a new tool for tracking HCV core uptake.

Circulating 'free' non-enveloped Hepatitis C virus (HCV) core protein has been demonstrated in HCV-infected patients, and HCV subgenomes with deletions of the envelope proteins have been previously identified. Initial studies from our laboratory, previously published, indicated that expression of HCV core in insect cells can direct the formation of capsid-like particles lacking the envelope glycoproteins. These protein nanospheres, morphologically similar to natural capsids, were shown to be taken up by human hepatic cells and to produce cell-signalling events. To follow the intracellular fate of these particles we fused the core protein to eGFP. We demonstrate that the chimeric proteins core(173)-eGFP, eGFP-core(191) and eGFP-core(173) can be efficiently expressed, self-assembled, and form fluorescent non-enveloped capsid-like particles. By using confocal microscopy and FACS analysis, we provide evidence that the fluorescent nanospheres can not only enter human hepatic cells - the main target of HCV - but also human immune cells such as T and B lymphocytes, as well as human myeloid leukaemia cells differentiated along the monocyte/macrophage-like pathway. The fluorescent particles might thus be used to trace the intracellular trafficking of naked HCV capsids as showed by live microscopy and to further understand their biological significance.

IL-6 and IL-8 release is mediated via multiple signaling pathways after stimulating dendritic cells with lysophospholipids.

Lysophosphatidic acid (LPA) and sphingosine 1-phosphate (S1P) are bioactive lipid mediators, which are known to play major roles in allergic reactions as well as in tumor pathogenesis. Here, the biological activities and signal pathways of these lysophospholipids (LPLs) in dendritic cells (DCs) were characterized further. Flow cytometric and immunoblot analyses indicate that immature as well as mature DCs express the LPL receptors S1P1, S1P3, S1P5, and LPA2, but not S1P2, S1P4, LPA1, or LPA3. Moreover, enzyme-linked immunosorbent assay experiments demonstrate that simultaneous addition of these LPLs to immature DCs in the presence of lipopolysaccharide enhanced the secretion of the inflammatory cytokines interleukin (IL)-6 and IL-8 in maturing DCs. In contrast, no modification of IL-6 or IL-8 release was observed after exposure of mature DCs to LPLs alone. In addition, studies with pertussis toxin and mitogen-activated protein kinase (MAPK) kinase inhibitor PD98059 suggested that Gi proteins and MAPK pathway are involved in these LPL-induced cell responses. Corroborating these findings, we observed that LPLs induce the phosphorylation of extracellular signal-regulated kinase 1/2 in immature DCs but not in mature DCs. Further analyses show that inhibitors of phosholipase D, Rho, and protein kinase C also inhibited the LPL-induced release of IL-6 and IL-8. Therefore, our findings suggest that lipopolysaccharide in DCs uncouples LPL receptors from the signal-transducing machinery during maturation and that exposure of LPLs at early time-points to maturing DCs modifies the proinflammatory capacity of mature DCs.