Sex hormone modulation of cell growth and apoptosis of the human monocytic/macrophage cell line

Sex hormones seem to modulate the immune/inflammatory responses by different mechanisms in female and male rheumatoid arthritis patients. The effects of 17β-oestradiol and of testosterone were tested on the cultured human monocytic/macrophage cell line (THP-1) activated with IFN-γ in order to investigate their role in cell proliferation and apoptosis. Activated human THP-1 cells were cultured in the presence of 17β-oestradiol and testosterone (final concentration, 10 nM). The evaluation of markers of cell proliferation included the NF-κB DNA-binding assay, the NF-κB inhibition complex, the proliferating cell nuclear antigen expression and the methyl-tetrazolium salt test. Apoptosis was detected by the annexin V-propidium assay and by the cleaved poly-ADP ribose polymerase expression. Specific methods included flow analysis cytometry scatter analysis, immunocytochemistry and western blot analysis. Cell growth inhibition and increased apoptosis were observed in testosterone-treated THP-1 cells. Increased poly-ADP ribose polymerase-cleaved expression and decreased proliferating cell nuclear antigen expression, as well as an increase of IκB-α and a decrease of the IκB-α phosphorylated form (ser 32), were found in testosterone-treated THP-1 cells. However, the NF-κB DNA binding was found increased in 17β-oestradiol-treated THP-1 cells. The treatment with staurosporine (enhancer of apoptosis) induced decreased NF-κB DNA binding in all conditions, but particularly in testosterone-treated THP-1 cells. Treatment of THP-1 by sex hormones was found to influence cell proliferation and apoptosis. Androgens were found to increase the apoptosis, and oestrogens showed a protective trend on cell death – both acting as modulators of the NF-κB complex

APP1 Transcription Is Regulated by Inositol-phosphorylceramide Synthase 1-Diacylglycerol Pathway and Is Controlled by ATF2 Transcription Factor in Cryptococcus neoformans

Inositol-phosphorylceramide synthase 1 (Ipc1) is a fungal-specific enzyme that regulates the level of two bioactive molecules, phytoceramide and diacylglycerol (DAG). In previous studies, we demonstrated that Ipc1 regulates the expression of the antiphagocytic protein 1 (App1), a novel fungal factor involved in pathogenicity of Cryptococcus neoformans. Here, we investigated the molecular mechanism by which Ipc1 regulates App1. To this end, the APP1 promoter was fused to the firefly luciferase gene in the C. neofor-mans GAL7:IPC1 strain, in which the Ipc1 expression can be modulated, and found that the luciferase activity was indeed regulated when Ipc1 was modulated. Next, using the luciferase reporter assay in both C. neoformans wild-type and GAL7:IPC1 strains, we investigated the role of DAG and sphingolipids in the activation of the APP1 promoter and found that treatment with 1,2-dioctanoylglycerol does increase APP1 transcription, whereas treatment with phytosphingosine or ceramides does not. Two putative consensus sequences were found in the APP1 promoter for ATF and AP-2 transcription factors. Mutagenesis analysis of these sequences revealed that they play a key role in the regulation of APP1 transcription: ATF is an activator, whereas AP-2 in a negative regulator. Finally, we identified a putative Atf2 transcription factor, which is required for APP1 transcription and under the control of Ipc1-DAG pathway. These studies provide novel regulatory mechanisms of the sphingolipid pathway involved in the regulation of gene transcription of C. neoformans

Polymorphisms of the SCN1A gene in children and adolescents with primary headache and idiopathic or cryptogenic epilepsy: is there a linkage?

The purpose of this study was to evaluate the distribution of the polymorphisms of the SCN1A gene in a series of children and adolescents with primary headache and idiopathic or cryptogenic epilepsy compared to controls. Five non-synonymous exonic polymorphisms (1748A > T, 2656T > C, 3199A > G, 5771G > A, 5864T > C) of the SCN1A gene were selected and their genotyping was performed, by high resolution melting (HRM), in 49 cases and 100 controls. We found that among the five polymorphisms, only 3199A > G was a true polymorphism. We did not find a statistically significant difference between distribution of 3199A > G genotypes between cases and controls. We excluded the role of the SCN1A gene in the pathogenesis of comorbidity between headache (especially migraine) and epilepsy. The SCN1A gene is a major gene in different epilepsies and epilepsy syndromes; the HRM could be the new methodology, more rapid and efficacious, for molecular analysis of the SCN1A gene

Correction: Alternatively Activated (M2) Macrophage Phenotype Is Inducible by Endothelin-1 in Cultured Human Macrophages

[This corrects the article DOI: 10.1371/journal.pone.0166433.]

CTLA4-Ig interacts with cultured synovial macrophages from rheumatoid arthritis patients and downregulates cytokine production

Introduction: Co-stimulatory signal B7(CD80/CD86):CD28 is needed in order to activate T cells in immune response. Cytotoxic T lymphocyte-associated antigen-4-immunoglobulin (CTLA4-Ig) binding to the B7 molecules on antigen-presenting cells downregulates this activation and represents a recent biological treatment in rheumatoid arthritis (RA). Objectives of the study were to investigate the presence of the B7.2 (CD86) molecule and its masking by CTLA4-Ig on cultures of both RA synovial macrophages (RA SM), and of macrophages differentiated from THP-1 cells (M). In addition, the anti-inflammatory effects of CTLA4-Ig on co-cultures of RA SM and M with activated T cells were tested.Methods: All macrophages were co-cultured for 24 hours with activated T cells, without or with CTLA4-Ig (10, 100, 500 \u3bcg/ml for 1 hour, 3 hours and overnight, respectively). Immunofluorescence (IF) staining for B7.2, and an analysis of inflammatory cytokine expression (interleukin (IL) -6, tumor necrosis factor (TNF) \u3b1, IL-1\u3b2, transforming growth factor (TGF) \u3b2) by immunocytochemistry (ICC), western blot (WB) and reverse transcriptase-polymerase chain reaction (RT-PCR) were performed.Results: Macrophages showed intense B7.2 expression. CTLA4-Ig/B7.2 masking was evident for all macrophages, even after only 1 hour of cell culture (range from 10 to 100 \u3bcg/ml). ICC of co-cultures showed a dose-dependent decrease in inflammatory cytokines (P < 0.001 for IL-6, TNF\u3b1, IL-1\u3b2 and TGF\u3b2). Data were confirmed by WB and RT-PCR analysis.Conclusions: Optimal concentrations of CTLA4-Ig for the CTLA4-Ig/B7.2 masking on activated macrophages were identified and were found to induce significant downregulation in the cell production of IL-6, TNF\u3b1, IL1-\u3b2 and TGF\u3b2. In conclusion, macrophages would appear to be a sensitive target for CTLA4-Ig treatment in RA

Role of Sterylglucosidase 1 (Sgl1) on the pathogenicity of Cryptococcus neoformans: Potential applications for vaccine development

Cryptococcosis caused by C. neoformans and C. gattii affects a large population and is a cause of significant morbidity and mortality. Despite its public health burden, there are currently no vaccines against cryptococcosis and new strategies against such infections are needed. In this study, we demonstrate that C. neoformans has the biochemical ability to metabolize sterylglucosides (SGs), a class of immunomodulatory glycolipids. Genetic manipulations that eliminate cryptococccal sterylglucosidase lead to the accumulation of SGs and generate a mutant strain (Îsgl1) that is non-pathogenic in the mouse models of cryptococcosis. Interestingly, this mutant strain acts as a vaccine strain and protects mice against cryptococcosis following infection with C. neoformans or C. gattii. The immunity induced by the Îsgl1 strain is not CD4+ T-cells dependent. Immunocompromised mice, which lack CD4+ T-cells, are able to control the infection by Îsgl1 and acquire immunity against the challenge by wild-type C. neoformans following vaccination with the Îsgl1 strain. These findings are particularly important in the context of HIV/AIDS immune deficiency and suggest that the Îsgl1 strain might provide a potential vaccination strategy against cryptococcosis

A Mediterranean Diet Mix Has Chemopreventive Effects in a Murine Model of Colorectal Cancer Modulating Apoptosis and the Gut Microbiota

Objectives: Unhealthy dietary patterns have been associated with colorectal cancer (CRC) onset while Mediterranean Diet (MD) has been proposed for CRC prevention. This study evaluated the effect of a Mediterranean Diet Mix (MD-MIX) on colonic tumors development in A/J mice fed a low-fat (LFD) or a high-fat western diet (HFWD), and injected with the procarcinogen azoxymethane (AOM).Materials and Methods: Forty A/J male mice were randomly assigned into four feeding arms (10 mice/arm; LFD, LFD-MD-MIX, HFWD, HFWD-MD-MIX) to be treated with AOM. Ten mice were exposed to the diets alone (Healthy LFD and Healthy HFWD) to be used as control. Tumor incidence and multiplicity were evaluated at sacrifice. Mucosal fatty acid content and urinary phenolic compounds were assayed by mass spectrometry. Apoptosis was evaluated by TUNEL assay and gene expression markers. Cell proliferation was evaluated by Ki67 immunohistochemistry. Microbiota composition was assessed at different time points by 16S RNA sequencing.Results: A tumor incidence of 100% was obtained in AOM-treated mice. The MD-MIX supplementation was able to reduce the number of colonic lesions in both LFD and HFWD-fed mice and to induce apoptosis, in particular in the LFD-MD-MIX arm. Moreover, a preventive effect on low-grade dysplasia and macroscopical lesions (>1 mm) development was found in HFWD-fed mice together with a regulation of the AOM-driven intestinal dysbiosis.Conclusions: MD-MIX was able to counteract CRC development in mice under different dietary backgrounds through the regulation of apoptosis and gut microbiota

The impact of the COVID-19 pandemic on surgical management of breast cancer:global trends and future perspectives

Introduction: The rapid spread of COVID-19 across the globe is forcing surgical oncologists to change their daily practice. We sought to evaluate how breast surgeons are adapting their surgical activity to limit viral spread and spare hospital resources. Methods: A panel of 12 breast surgeons from the most affected regions of the world convened a virtual meeting on April 7, 2020, to discuss the changes in their local surgical practice during the COVID-19 pandemic. Similarly, a Web-based poll based was created to evaluate changes in surgical practice among breast surgeons from several countries. Results: The virtual meeting showed that distinct countries and regions were experiencing different phases of the pandemic. Surgical priority was given to patients with aggressive disease not candidate for primary systemic therapy, those with progressive disease under neoadjuvant systemic therapy, and patients who have finished neoadjuvant therapy. One hundred breast surgeons filled out the poll. The trend showed reductions in operating room schedules, indications for surgery, and consultations, with an increasingly restrictive approach to elective surgery with worsening of the pandemic. Conclusion: The COVID-19 emergency should not compromise treatment of a potentially lethal disease such as breast cancer. Our results reveal that physicians are instinctively reluctant to abandon conventional standards of care when possible. However, as the situation deteriorates, alternative strategies of de-escalation are being adopted. Implications for Practice: This study aimed to characterize how the COVID-19 pandemic is affecting breast cancer surgery and which strategies are being adopted to cope with the situation. © 2020 AlphaMed Pres