Crosstalk between keratinocytes and neutrophils shapes skin immunity against S. aureus infection

IntroductionStaphylococcus aureus (S. aureus) infection of the skin leads to a rapid initial innate immune response with keratinocytes in the epidermis as the initial sensors. Polymorphonuclear neutrophils (PMNs) are the first innate immune cells to infiltrate infection sites where they provide an effective first-line of defense. Previous work of our group showed that in inflamed skin a crosstalk between PMNs and keratinocytes results in enhanced S. aureus skin colonization.MethodsIn this work, we used an in vitro co-culture model to studied the crosstalk between primary human keratinocytes (PHKs) and PMNs in a sterile environment and upon S. aureus infection. We investigated the influence of PHKs on PMN activation by analyzing PMN lifespan, expression of degranulation markers and induction of proinflammatory cytokines. Furthermore, we analyzed the influence of PMNs on the inflammatory response of PHKs. Finally, we investigated the influence of the skin microbiome on PMN-mediated skin inflammation.ResultsWe show that co-culture of PMNs with PHKs induces activation and degranulation of PMNs and significantly enhances their lifespan compared to PMN cultivation alone by an IL-8 mediated mechanism and, furthermore, primes PMNs for enhanced activity after S. aureus infection. The prolonged incubation with PMNs also induces inflammatory responses in PHKs which are further exacerbated in the presence of S. aureus and induces further PMN recruitment thus fueling skin inflammation. Interestingly, infection of PHKs with the skin commensal S. epidermidis reduces the inflammatory effects of PMNs in the skin and exhibits an anti-inflammatory effect.DiscussionOur data indicate that skin infiltrating PMNs and PHKs influence each other in such a way to enhance skin inflammation and that commensal bacteria are able to reduce the inflammatory effect

Biological functions of casein kinase 1 isoforms and putative roles in tumorigenesis

Isoforms of the casein kinase 1 (CK1) family have been shown to phosphorylate key regulatory molecules involved in cell cycle, transcription and translation, the structure of the cytoskeleton, cell-cell adhesion and receptor-coupled signal transduction. They regulate key signaling pathways known to be critically involved in tumor progression. Recent results point to an altered expression or activity of different CK1 isoforms in tumor cells. This review summarizes the expression and biological function of CK1 family members in normal and malignant cells and the evidence obtained so far about their role in tumorigenesis. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/1476-4598-13-231) contains supplementary material, which is available to authorized users

Casein kinase 1α has a non-redundant and dominant role within the CK1 family in melanoma progression

Influence of the modulation of CK1 isoform expression on cell viability and cell cycle. (A) Inhibition of isoform specific CK1-activity via combined siRNA mediated knockdown of CK1α, CK1δ and CK1ε. SbCl2 (left diagram) and SKMEL19 (right diagram) cells were transduced with isoform specific siRNA or a non-silencing control and cell growth was monitored for 4 days using the MUH viability assay. Fluorescence intensities were normalized (100 %) to the start point at 24 h post transfection of the siRNA. Shown is the mean with SD of hexatuplicates. (B) Cell cycle analysis after knockdown of CK1 isoforms in SbCl2 and SKMel19 melanoma cells. After ice-cold ethanol fixation melanoma tumor cells were stained with 50 μg/ml propidium iodide containing RNase in PBS for 30 min and analyzed in a LSRII flow cytometer (BD). (C) Cell cycle analysis at 48 h after induction of CK1 isoforms revealed a significant subG1 apoptotic population only after overexpression of CK1α. (TIF 701 kb

Inhibition of PI3K-AKT-mTOR Signaling Sensitizes Melanoma Cells to Cisplatin and Temozolomide

In melanoma, the PI3K-AKT-mTOR (AKT) and RAF-MEK-ERK (MAPK) signaling pathways are constitutively activated and appear to play a role in chemoresistance. Herein, we investigated the effects of pharmacological AKT and MAPK pathway inhibitors on chemosensitivity of melanoma cells to cisplatin and temozolomide. Chemosensitivity was tested by examining effects on growth, cell cycle, survival, expression of antiapoptotic proteins, and invasive tumor growth of melanoma cells in monolayer and organotypic culture, respectively. MAPK pathway inhibitors did not significantly increase chemosensitivity. AKT pathway inhibitors consistently enhanced chemosensitivity yielding an absolute increase of cell growth inhibition up to 60% (P<0.05, combination therapy vs monotherapy with inhibitors or chemotherapeutics). Cotreatment of melanoma cells with AKT pathway inhibitors and chemotherapeutics led to a 2- to 3-fold increase of apoptosis (P<0.05, combination therapy vs monotherapy) and completely suppressed invasive tumor growth in organotypic culture. These effects were associated with suppression of the antiapoptotic Bcl-2 family protein Mcl-1. These data suggest that inhibition of the PI3K-AKT-mTOR pathway potently increases sensitivity of melanoma cells to chemotherapy

β-Catenin Signaling Increases during Melanoma Progression and Promotes Tumor Cell Survival and Chemoresistance

Beta-catenin plays an important role in embryogenesis and carcinogenesis by controlling either cadherin-mediated cell adhesion or transcriptional activation of target gene expression. In many types of cancers nuclear translocation of beta-catenin has been observed. Our data indicate that during melanoma progression an increased dependency on the transcriptional function of beta-catenin takes place. Blockade of beta-catenin in metastatic melanoma cell lines efficiently induces apoptosis, inhibits proliferation, migration and invasion in monolayer and 3-dimensional skin reconstructs and decreases chemoresistance. In addition, subcutaneous melanoma growth in SCID mice was almost completely inhibited by an inducible beta-catenin knockdown. In contrast, the survival of benign melanocytes and primary melanoma cell lines was less affected by beta-catenin depletion. However, enhanced expression of beta-catenin in primary melanoma cell lines increased invasive capacity in vitro and tumor growth in the SCID mouse model. These data suggest that beta-catenin is an essential survival factor for metastatic melanoma cells, whereas it is dispensable for the survival of benign melanocytes and primary, non-invasive melanoma cells. Furthermore, beta-catenin increases tumorigenicity of primary melanoma cell lines. The differential requirements for beta-catenin signaling in aggressive melanoma versus benign melanocytic cells make beta-catenin a possible new target in melanoma therapy

A rare loss-of-function genetic mutation suggest a role of dermcidin deficiency in hidradenitis suppurativa pathogenesis

Hidradenitis suppurativa (HS) is a chronic inflammatory skin disease with a multifactorial aetiology that involves a strict interplay between genetic factors, immune dysregulation and lifestyle. Familial forms represent around 40% of total HS cases and show an autosomal dominant mode of inheritance of the disease. In this study, we conducted a whole-exome sequence analysis on an Italian family of 4 members encompassing a vertical transmission of HS. Focusing on rare damaging variants, we identified a rare insertion of one nucleotide (c.225dupA:p.A76Sfs*21) in the DCD gene encoding for the antimicrobial peptide dermcidin (DCD) that was shared by the proband, his affected father and his 11-years old daughter. Since several transcriptome studies have shown a significantly decreased expression of DCD in HS skin, we hypothesised that the identified frameshift insertion was a loss-of-function mutation that might be associated with HS susceptibility in this family. We thus confirmed by mass spectrometry that DCD levels were diminished in the affected members and showed that the antimicrobial activity of a synthetic DCD peptide resulting from the frameshift mutation was impaired. In order to define the consequences related to a decrease in DCD activity, skin microbiome analyses of different body sites were performed by comparing DCD mutant and wild type samples, and results highlighted significant differences between the groins of mutated and wild type groups. Starting from genetic analysis conducted on an HS family, our findings showed, confirming previous transcriptome results, the potential role of the antimicrobial DCD peptide as an actor playing a crucial part in the etio-pathogenesis of HS and in the maintenance of the skin’s physiological microbiome composition; so, we can hypothesise that DCD could be used as a novel target for personalised therapeutic approach