Generation of dendritic cells: A critical comparison of different methodological approaches

Dendritic cells (DCs) represent the most important antigen presenting cells (APCs). In vivo they originate from bone marrow myeloid and lymphoid precursors. In a first stage, the so-called immature DCs stay into nonlymphoid tissues and show high capacity of antigen capture and processing, but low T cell stimulatory capacity. After the internalization of the antigen, in the presence of inflammatory mediators, DCs mature and migrate out of nonlymphoid tissues into the blood or lymph, reaching secondary lymphoid organs. In this second stage the mature DCs lose the ability to capture antigens and increase the capacity to stimulate T cells, controlling the immune responses. Indeed, DCs are a family of heterogeneous cells and each subset exerts control over a different area of immunity, activating innate and acquired systems or maintaining the tolerance to antigens. For in vitro studies, dendritic cells can be isolated directly from peripheral blood through blood dendritic cell antibodies (BDCA), but they have been difficult to obtain due to their low concentration. So a wide variety of methods to have DCs has been performed, including the generation from circling CD14+ monocytes or from CD34+ progenitors. A multitude of systems have been carried out also to obtain these precursor cells. The method employed could affect morphological and functional features of the resultant DCs. For example, Ratta M and co-workers affirm that peripheral blood CD34+ cells generate a significantly higher number of fully functional DCs, capable of processing and presenting soluble antigens to autologous T cells. Elkord E and coworkers state that DCs generated from a positive immunoselected monocytes drastically reduce their ability to secrete cytokines as interleukin (IL)-12, IL-10, TNF-α, if compared to those generated from plastic adherence-isolated monocytes. Our own results indicate that monocytes selected by a positive selection method originate partially-mature DCs also without a maturational stimulus, characterized by low capacity of internalizing antigens and of inducing T cell proliferation also after lipopolysaccharide addition. Accordingly, in this commentary we review the recent progress in understanding the correlation between the approach used to obtain dendritic cells and their related properties, in order to indicate a correct application of DCs for research or clinical studies

The Centenary of Immune Thrombocytopenia-Part 2: Revising Diagnostic and Therapeutic Approach

Primary immune thrombocytopenia (ITP) is the most common cause of thrombocytopenia in children and adolescents and can be considered as a paradigmatic model of autoimmune disease. This second part of our review describes the clinical presentation of ITP, the diagnostic approach and overviews the current therapeutic strategies. Interestingly, it suggests an algorithm useful for differential diagnosis, a crucial process to exclude secondary forms of immune thrombocytopenia (IT) and non-immune thrombocytopenia (non-IT), which require a different therapeutic management. Advances in understanding the pathogenesis led to new therapeutic targets, as thrombopoietin receptor agonists, whose role in treatment of ITP will be discussed in this work

The Centenary of Immune Thrombocytopenia - Part 1: Revising Nomenclature and Pathogenesis

The natural history of the immune thrombocytopenia (ITP) is interesting and intriguing because it traces different steps underlying autoimmune diseases. The review points out the main steps that have accompanied the stages of its history and the consequential changes related to its terminology. ITP is an autoimmune disease resulting from platelet antibody-mediated destruction and impaired megakaryocyte and platelet production. However, research advances highlight that a complex dysregulation of the immune system is involved in the pathogenesis of this condition. The review examines the role of the multiple immune components involved in the autoimmunity process, focusing on the more recent mechanisms, which could be new promising therapeutic targets for ITP patients

Effects of Anakinra on Health-Related Quality of Life in a Patient with 1129G>A/928G>A Mutations in MVK Gene and Heterozygosity for the Mutation 2107C>A in CIAS1 Gene

Mevalonate kinase deficiency impairs several aspects of the patient's quality of life, thus early diagnosis and treatment are required to improve health-related quality of life (HRQOL). A 15-year-old patient with double heterozygosity for the mutations 1129G>A and 928G>A in MVK gene, heterozygosity for the mutation 2107C>A in CIAS1 gene and hyper-IgD syndrome phenotype, has been treated with anakinra with a reduction of 50% in the number of fever episodes per month, a reduction of 33% in the days of fever for each attack and normal blood tests in the intercritical phase. The RAND 36-Item Health Survey has been used for the assessment of HRQOL before and after the treatment with anakinra. The patient's quality of life showed an overall improvement of 27%; results showed a better improvement in role limitations due to physical health (50%)

The effects of zoledronate on monocyte-derived dendritic cells from melanoma patients differ depending on the clinical stage of the disease.

Zoledronic acid has shown indirect anticancer effects on angiogenesis, the tumor microenvironment and immune responses. Its immunological action is exerted, at least in part, via its modulating properties. The aim of this study was to investigate the in vitro effects of zoledronic acid on the dendritic cells of melanoma patients. Peripheral blood samples were collected from 26 patients with melanoma and 11 healthy donors. Dendritic cells were derived from purified monocytes, and zoledronic acid (ZA) was added on the first day of culture. The phenotype and function of the generated cells were evaluated by flow cytometry. The ZA-treated monocytes from patients with early-stage disease generated DCs characterized by reduced endocytic activity and increased allostimulatory capacity compared with the untreated samples, allowing restoration of the DC function observed in normal subjects. In contrast, the ZA-treated monocytes from patients at stage III generated cells with higher CD14 antigen expression and endocytosis than the untreated samples. Therefore, in melanoma patients, the in vitro ZA effects differ according to the progression of the disease. In addition, our preliminary results appear to suggest that ZA effects are also influenced by the expression of CD14 antigen, indicating that the DC phenotype together with clinical characteristics must be considered in the choice of patients to be treated with ZA. Our work focus on the effect of ZA on monocyte-derived DCs from melanoma patients, showing that the effects of therapeutic doses of this drug might be mediated at least in part by modulation of myeloid cell function

UN OSPITE INATTESO: UNA RICKETTSIA

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Efficacy and Feasibility of the Epithelial Cell Adhesion Molecule (EpCAM) Immunomagnetic Cell Sorter for Studies of DNA Methylation in Colorectal Cancer

The aim of this work was to assess the impact on measurements of methylation of a panel of four cancer gene promoters of purifying tumor cells from colorectal tissue samples using the epithelial cell adhesion molecule (EpCAM)-immunomagnetic cell enrichment approach. We observed that, on average, methylation levels were higher in enriched cell fractions than in the whole tissue, but the difference was significant only for one out of four studied genes. In addition, there were strong correlations between methylation values for individual samples of whole tissue and the corresponding enriched cell fractions. Therefore, assays on whole tissue are likely to provide reliable estimates of tumor-specific methylation of cancer genes. However, tumor cell tissue separation using immunomagnetic beads could, in some cases, give a more accurate value of gene promoter methylation than the analysis of the whole cancer tissue, although relatively expensive and time-consuming. The efficacy and feasibility of the immunomagnetic cell sorting for methylation studies are discussed

Therapeutic aspects of Sydenham's Chorea: an update

Sydenham's Chorea (SC) is a hyperkinetic movement disorder associated with neuropsychiatric manifestations. It is believed to be caused by the autoimmune response following a group A beta-hemolytic streptococcal (GABHS) pharyngitis, and it is one of the major diagnostic criteria for Acute Rheumatic Fever (ARF) diagnosis. Despite having been known and studied for centuries, there are still no standardized therapies or official guidelines for SC treatment, so that it is necessarily left to physicians' clinical experience. Antibiotic treatment, symptomatic therapies, and immunomodulatory treatment are the three pillars upon which SC patients' management is currently based, but they still lack a solid scientific basis. The aim of this writing is precisely to review the state of the art of SC's treatment, with an overview of the advances made in the last 5 years. However, since the therapeutic uncertainties are a mere reflection of the severe gap of knowledge that concerns SC's pathogenesis and manifestations, the importance of high-quality research studies based on homogenized methodologies, instruments, and measured outcomes will also be stressed

Paediatric recurrent pericarditis: Appropriateness of the standard of care and response to IL1-blockade

Objective: To analyse, in a cohort of paediatric patients with recurrent pericarditis (RP) undergoing anti-IL-1 treatment: the agent and dosing used as first line treatment, the long-term efficacy of IL1-blockers, the percentage of patients achieving a drug-free remission, the presence of variables associated with drug-free remission. Study design: Data were collected from patients' charts. Annualized relapse rate (ARR) was used for evaluation of treatment efficacy, bivariate logistic regression analysis for variables associated with drug-free remisison. Results: 58 patients, treated between 2008 and 2018, were included in the study (mean follow-up 2.6 years). 14/56 patients non-responsive to first line drugs were under-dosed. 57 patients were treated with anakinra: the ARR before and during daily treatment was 3.05 and 0.28, respectively (p<0.0001); an increase to 0.83 was observed after the reduction/withdrawal of treatment (p<.0001). The switch from anakinra to canakinumab (5 patients) was associated to an increase of the ARR (0.49 vs 1.46), but without statistical significance (p=0.215). At last follow-up only 9/58 patients had withdrawn all treatments. With the limits of a retrospective study and the heterogeneity between the patients enrolled in the study, a shorter duration of treatment with anakinra was the only variable associated with drug-free remission. Conclusion: This study shows that most of the pediatric patients with RP needing IL-1 blockade received an inadequate treatment with first line agents. The effectiveness of anakinra is supported by this study, but few patients achieved drug free-remission. The different rate of response to anakinra and canakinumab may suggest a possible role of IL1α in the pathogenesis of RP