Differential microRNA expression between decidual and peripheral blood natural killer cells in early pregnancy.

STUDY QUESTION: Have decidual natural killer (dNK) cells a different microRNA (miRNA or miR) expression pattern compared to NK cells circulating in the peripheral blood (pb) of healthy pregnant women in the first trimester of gestation? SUMMARY ANSWER: dNK cells have a unique miRNA profile, showing exclusive expression of a set of miRNAs and significant up- or down-regulation of most of the miRNAs shared with pbNK cells. WHAT IS KNOWN ALREADY: dNK cells differ from pbNK cells both phenotypically and functionally, and their origin is still debated. Many studies have indicated that miRNAs regulate several important aspects of NK cell biology, such as development, activation and effector functions. STUDY DESIGN, SIZE, DURATION: Decidua basalis and peripheral blood specimens were collected from women (n = 7) undergoing voluntary termination of gestation in the first trimester of pregnancy. dNK and pbNK cells were then highly purified by cell sorting. PARTICIPANTS/MATERIALS, SETTING, METHODS: miRNAs expression was analysed by quantitative RT-PCR (qRT-PCR)-based arrays using RNA purified from freshly isolated and highly purified pbNK and dNK cells. Results from arrays were validated by qRT-PCR assays. The bioinformatics tool ingenuity pathway analysis (IPA) was applied to determine the cellular network targeted by validated miRNAs and the correlated biological functions. MAIN RESULTS AND THE ROLE OF CHANCE: Herein, we identified the most differentially expressed miRNAs in NK cells isolated from peripheral blood and uterine decidua of pregnant women. We found that 36 miRNAs were expressed only in dNK cells and two miRNAs only in pbNK cells. Moreover, 48 miRNAs were commonly expressed by both NK cell preparations although at different levels: 28 were upregulated in dNK cells, while 15 were downregulated compared to pbNK cells. Validation of a selected set (n = 11) of these miRNAs confirmed the differential expression of nine miRNAs: miR-10b and miR-214 expressed only in dNK cells and miR-200a-3p expressed only in pbNK cells; miR-130b-3p, miR-125a-5p, miR-212-3p and miR-454 were upregulated while miR-210-3p and miR-132 were downregulated in dNK cells compared to pbNK cells. IPA network analysis identified a single network connecting all the miRNAs as well as their significant involvement in several classes of functions: 'Organismal injury, Reproductive system disease, Inflammatory disease' and 'Cellular development'. These miRNAs target molecules such as argonaute 2, tumour protein p53, insulin and other genes that belong to the same network and significantly influence cell differentiation and pregnancy. LIMITATIONS, REASONS FOR CAUTION: In the present study, the cellular network and biological functions modulated by miRNAs differentially expressed in dNK and pbNK cells were identified by IPA considering only molecules and relationships that were with confidence 'experimentally observed' in leucocytes. The decidual and pbNK cells that were analysed here are a heterogeneous population and further study will help to disentangle whether there are differences in miRNA production by the different subsets of NK cells. WIDER IMPLICATIONS OF THE FINDINGS: This is the first study describing a different miRNA expression profile in dNK cells compared to matched pbNK cells during the first trimester of pregnancy. Our findings improved the body of knowledge on dNK cell biology and strongly suggest further investigation into the roles of miRNAs that are differentially expressed in human dNK compared to pbNK cells. Our results suggest that specific miRNAs can modulate dNK cell origin and functions, highlighting a potential role of this miRNA signature in human development and diseases. STUDY FUNDING/COMPETING INTEREST(S): This work was supported by grants from the Istituto Pasteur, Fondazione Cenci Bolognetti, the European NoE EMBIC within FP6 (Contract number LSHN-CT-2004-512040), Istituto Italiano di Tecnologia, and Ministero dell'Istruzione, dell'Università e della Ricerca (Ricerche Universitarie), and from Università Politecnica delle Marche. There are no conflicts of interest to declare.

Selective inhibitors of aurora kinases inhibit proliferation, reduce cell viability and impair cell cycle progression in papillary thyroid carcinoma cells.

The three members of the Aurora kinase family, Aurora-A, -B and -C, regulate several aspects of the mitotic process, and their aberrant expression and/or function causes mitotic abnormalities leading either to cell death or aneuploidy. They are found overexpressed in several human malignancies, including the papillary thyroid carcinoma (PTC). In the present study, we sought to establish whether Aurora kinase inhibition could be of any therapeutic value in the treatment of aggressive forms of PTC, enduring to radioactive iodide (RAI) ablation. To this end, the effects of selective inhibitors of Aurora-A (MLN8237) and Aurora-B (AZD1152) were analyzed on 3 human PTC cell lines expressing either wild-type (K1 and TPC1) or mutant p53 (BCPAP). The two inhibitors were capable of reducing cell proliferation in a time- and dose-dependent manner, with IC50 comprised between 65.4 and 114.9 nM for MLN8237, and between 26.6 and 484.6 nM for AZD1152. Immunofluorescence experiments confirmed that AZD1152 inhibited Aurora-B phosphorylation of histone H3 on Ser10, however, it did not affect Aurora-A autophosphorylation. MLN8237 inhibited Aurora-A autophosphorylation as expected, but at concentrations required to achieve the maximum antiproliferative effects it also abolished H3 (Ser10) phosphorylation. Time-lapse videomicroscopy evidenced that both inhibitors prevented the completion of cytokinesis, and cytofluorimetric analysis showed accumulation of cells in G2/M phase and/or polyploidy. Apoptosis was induced in all the cells by both inhibitors independently from the p53 status. In conclusion, in the present preclinical study MLN8237 and AZD1152 have emerged as promising drug candidates for RAI-insensitive PTC.

Nutritional management of hyperlipaemia in a jenny: a brief report.

An adult jenny (5-years-old, non-pregnant) was presented to the Veterinary Teaching Hospital (VTH) of the University of Sassari, with a recent history of appetite loss, extreme underweight condition and reluctance to move. On physical inspection, emaciation [body condition score, BCS: 3/9], muscular waste [muscular condition score, MCS: 1/5], loose/running faeces [faecal score, FS: 2/8], and a general state of mild dehydration were found. Blood analyses outlined a general undernourishment condition [circulating albumins, ALB: 17.6 g/L (21.6-31.6 g/L)] with underlying systemic inflammatory profile and moderate increase in circulating enzymes to explore liver function [aspartate amino-transferase, AST: 657 u/L (279-430 u/L); alanine amino-transferase ALT: 60 u/L (5-14 u/L); gamma-glutamyl-transferase, γ-GT: 87 IU/L (14-69 IU/L); total bilirubin close to the upper limit, TB: 0.20 mg/dL(0.07-0.21 mg/dL)]and hyperlipaemia [TG: 8.70 mmol/L (0.60-2.87 mmol/L)], following fat depots mobilisation, with total cholesterol closed to the lower limit of the physiological range. Hyper-phosphataemia was linked to haemolytic anaemia [P:1.81 mmol/L (0.77-1.39 mmol/L) and red blood cells, RBC: 4.14 1012/L (4.40-7.10 1012)] aligned with the TB to the upper limit. On ultrasound abdominal imaging, enlarged and hyper-echogenic liver was observed. Based on the clinical evaluation, a condition of hepatic lipidosis was diagnosed, requiring dedicated nutritional treatment to solve the extreme emaciation along with the metabolic disorder in support of medical therapy. A two-step feeding protocol was planned to support treatments aiming at immediate re-hydration (Ringer lactate solution 2 ml/kg/8 h). The nutritional objectives were meant at first to restart the voluntary feed intake. Gradual increasing energy provision through a palatable hay-based diet was planned to cover one fourth of daily metabolizable energy requirement calculated on the expected metabolic weight, adjusted according to the daily intake of feed and clinical condition. At the conclusion of this first 7-day phase, circulating blood parameters were closer to the reference values and the BCS moved from 3 to 4 out of 9. Bowel motility was restored, and faecal score improved (4/8). In the second phase, allowance to pasture and a combination diet with compound mixed feed were designed. Within four weeks of starting the nutritional plan, blood parameters were re-established to reference values. The gradual feed provision calculated in this two-phase approach proved successful in support of the overall clinical improvement observed after four weeks of treatment, in a severely undernourished jenny with compromised liver functions.

The dual Aurora kinase inhibitor ZM447439 prevents anaplastic thyroid cancer cell growth and tumorigenicity.

The anaplastic thyroid cancer (ATC) is among the most aggressive human tumors which fail to respond to all the currently available therapeutic approaches. As a consequence most patients die within a few months from diagnosis. In the present preclinical study, the effects of the ZM447439, a functional inhibitor of Aurora kinases, on the growth and tumorigenicity of a panel of ATC derived cell lines (CAL-62, 8305C, 8505C and BHT-101) were evaluated. The treatment of the different ATC cells with ZM447439 inhibited proliferation in a time- and dose-dependent manner, with IC50 comprised between 0.5 mM and 5 mM. Moreover, the drug remarkably impaired the formation of colonies in soft agar of all the cell lines. Consistently with Aurora inhibition, immunofluorescence and immunoblotting experiments demonstrated that Aurora auto-phosphorylation following drug treatment was completely abrogated, and treated cells were characterized by the presence of multiple spindles with short microtubules. In the same experiments we observed the loss of histone H3 phosphorylation on Ser10, specifically due to Aurora-B, after ZM447439 treatment. Time-lapse videomicroscopy and flow cytometric analysis demonstrated that in presence of ZM447439 the cells were able to enter mitosis but not to complete it, becoming polyploid. Almost all the ATC cell lines studied showed increased apoptosis after only 48 h of treatment. In conclusion, our data demonstrate that ZM447439 is effective in reducing cell growth and tumorigenicity of different ATC derived cell lines, and further investigations are needed to exploit its potential therapeutic value for ATC treatment.

SARS-CoV-2 infection and H1N1 vaccination: does a relationship between the two factors really exist? A retrospective analysis of a territorial cohort in Ferrara, Italy.

OBJECTIVE: SARS-CoV-2 has been compared with other strains of coronaviruses, SARS-CoV and MERS-CoV, and with the flu viruses: all of them manifest themselves with respiratory symptoms and, although their genetic patterns are similar, the spread of SARS-CoV-2 infection has quickly reached global dimensions, demonstrating that SARS-CoV-2 is a virus with greater spreading capacity, albeit less lethal. Compared with influenza viruses, coronaviruses have a longer incubation period and the patients with coronaviruses' syndromes develop more severe diseases requiring frequent hospitalizations and intensive care admissions. The aim was to explore the relationships between seasonal influenza vaccination and coronavirus infection and to understand whether this hypothetic role by the flu vaccines modifies SARS-CoV-2 infection's outcomes. PATIENTS AND METHODS: In this retrospective, multicenter study, we enrolled 952 patients diagnosed with SARS-CoV-2 infection; 448 were admitted to our two main hospitals in Ferrara territory, while the remaining 504 were isolated at home. We compared the group of patients who had been vaccinated for influenza in the previous 12 months to that of unvaccinated patients. RESULTS: Significant differences were found for both the need for hospitalization and 30-day mortality between vaccinated and unvaccinated patients. We found age to be the only independent risk factor for a worse 30-day prognosis, while gender, influenza vaccinations and age itself were independent risk factors for undergoing hospitalization. CONCLUSIONS: In our groups of patients, we found a relationship between seasonal influenza vaccinations and SARS-CoV-2 infection. Age seems to be the main risk factor for short-term mortality in COVID-19 inpatients, while the influenza vaccination is, together with gender and age itself, a determining factor in predicting the need for hospitalization.

Triggering through CD16 or phorbol esters enhances adhesion of NK cells to laminin via very late antigen 6.

Very late antigens VLA-1, VLA-2, VLA-3, and VLA-6, belonging to the beta 1 subfamily of integrins, have been identified as receptors for different binding domains of laminin (LM). We have detected VLA-6, but not VLA-1 and VLA-2 on a subset (50-70%) of fresh peripheral blood CD3-, CD16+, CD56+ human natural killer (NK) cells by immunofluorimetric and biochemical analysis. Binding assays performed on LM-coated plates showed that 10-15% of NK cells spontaneously adhere to LM, and this adhesion is mediated by VLA-6. Activation of NK cells through CD16 triggering or by phorbol ester results in a rapid increase of adhesion to LM, which is still mediated by VLA-6. The enhanced adhesiveness is not associated with changes in beta 1 LM receptor expression, while it correlates with changes in the phosphorylation status of alpha 6 subunit. The expression of VLA-6 on NK cells and the modulation of its avidity by activating stimuli may be relevant for NK cell migration and tissue location during inflammation or immune response.

UVB radiation induced effects on cells studied by FTIR spectroscopy.

We have made a preliminary analysis of the results about the effects on tumoral cell line (lymphoid T cell line Jurkat) induced by UVB radiation (dose of 310 mJ/cm(2)) with and without a vegetable mixture. In the present study, we have used two techniques: Fourier transform infrared spectroscopy (FTIR) and flow cytometry. FTIR spectroscopy has the potential to provide the identification of the vibrational modes of some of the major compounds (lipid, proteins and nucleic acids) without being invasive in the biomaterials. The second technique has allowed us to perform measurements of cytotoxicity and to assess the percentage of apoptosis. We already studied the induction of apoptotic process in the same cell line by UVB radiation; in particular, we looked for correspondences and correlations between FTIR spectroscopy and flow cytometry data finding three highly probable spectroscopic markers of apoptosis (Pozzi et al. in Radiat Res 168:698-705, 2007). In the present work, the results have shown significant changes in the absorbance and spectral pattern in the wavenumber protein and nucleic acids regions after the treatments.

Meta-analysis on the utility of radiotherapy for the treatment of Ocular Melanoma

Introduction: Uveal melanoma is the most common intraocular tumor in the adult population. It can affect any part of the uveal tract: the iris, ciliary body, and choroid. Historically, enucleation has been the mainstay of treatment for primary melanoma. In the last decade, however, radiotherapy has acquired an increasingly important role and has now become our first-line modality. However, it is still widely debated what is the most effective radiotherapy technique for this tumor. Purpose to perform a literature review on the utility of radiotherapy for primary ocular melanoma and determine the most effective radiotherapy technique Materials and Methods: We included all systematic and narrative reviews on the topic, published between September 2007 and November 2017 on PubMed and SCOPUS. Two independent reviewers assessed the eligibility criteria for each article using the PRISMA checklist. The methodological quality of narrative and systematic reviews was evaluated with the INSA and AMSTAR checklists, respectively Results: Our study analyzed a total of 23 studies, including 18 narrative reviews and 5 systematic reviews. Radiotherapy with Brachytherapy, Proton Therapy, SRS/SRT with gamma knife and cyber knife, are the most common choices for the treatment of primary ocular melanoma. These techniques allow for excellent lesion spread control, eye, and vision conservation, and improve overall patients' quality of life. Among the narrative reviews, the highest INSA score was 5/7, the lowest 2/7, the mean was 3.83/7 and median was 4/7. Among the systematic reviews, the highest AMSTAR score was 9/12, the lowest 4/12, the mean 5.6/7 and median 4/7 Conclusion: The number of studies available on this topic is scarce. Among those published, the methodological quality is modest, as assessed with the INSA and AMSTAR checklists. As a result, we are not able to determine what the most effective radiotherapy technique is

HTLV-V: a new human retrovirus isolated in a Tac-negative T cell lymphoma/leukemia.

A new human retrovirus was isolated from a continuous cell line derived from a patient with CD4+ Tac- cutaneous T cell lymphoma/leukemia. This virus is related to but distinct from human T cell leukemia/lymphoma virus types I and II (HTLV-I and HTLV-II) and human immunodeficiency virus (HIV-1). With the use of a fragment of provirus cloned from one patient with T cell leukemia, closely related sequences were found in DNA of the cell line and of tumor cells from seven other patients with the same disease; these sequences were only distantly related to HTLV-I. The phenotype of the cells and the clinical course of the disease were clearly distinguishable from leukemia associated with HTLV-I. All patients and the wife of one patient showed a weak serological cross-reactivity with both HTLV-I and HIV-1 antigens. None of the patients proved to be at any apparent risk for HIV-1 infection. The name proposed for this virus is HTLV-V, and the date indicate that it may be a primary etiological factor in the major group of cutaneous T cell lymphomas/leukemias, including the sporadic lymphomas known as mycoses fungoides.

Chlamydia pneumoniae induces T cell apoptosis through glutathione redox imbalance and secretion of TNF-alpha.

Chlamydia pneumoniae persistent infection has been implicated in the pathogenesis of several chronic inflammatory diseases including atherosclerosis, and we hypothesized that modulation of the apoptosis of macrophages and/or T cells by C. pneumoniae infection may contribute to the development of such diseases. We therefore evaluated apoptosis, cytokine response, and redox status in human primary T cells and macrophages infected with C. pneumoniae. In addition, co-cultures of T cells and macrophages infected with C. pneumoniae were also carried out. Apoptosis, and levels of glutathione (GSH), glutathione disulfide (GSSG), and tumour necrosis factor (TNF)-alpha were measured by flow cytometry, high performance liquid chromatography and enzyme-linked immunosorbent assay. C. pneumoniae induced apoptosis in T cells as well as in co-cultures of T cells and infected macrophages by marked decrease in GSH/GSSG ratio and increased production of TNF-alpha, respectively. The results demonstrate that interaction of C. pneumoniae with T cells and/or macrophages characterized by interference with redox status, and secretion of tumour necrosis factor-alpha culminates in the induction of T cell apoptosis and survival of infected macrophages. In conclusion, the inappropriate T cell response against C. pneumoniae and survival of infected macrophages could explain the persistence of this intracellular obligate pathogen in the host-organism; it may contribute to the development of chronic inflammatory diseases, although further studies are needed to clarify such a complex mechanism.

Ultrasound delivery of Surface Enhanced InfraRed Absorption active gold-nanoprobes into fibroblast cells: a biological study via Synchrotron-based InfraRed microanalysis at single cell level.

Ultrasound (US) induced transient membrane permeabilisation has emerged as a hugely promising tool for the delivery of exogenous vectors through the cytoplasmic membrane, paving the way to the design of novel anticancer strategies by targeting functional nanomaterials to specific biological sites. An essential step towards this end is the detailed recognition of suitably marked nanoparticles in sonoporated cells and the investigation of the potential related biological effects. By taking advantage of Synchrotron Radiation Fourier Transform Infrared micro-spectroscopy (SR-microFTIR) in providing highly sensitive analysis at the single cell level, we studied the internalisation of a nanoprobe within fibroblasts (NIH-3T3) promoted by low-intensity US. To this aim we employed 20 nm gold nanoparticles conjugated with the IR marker 4-aminothiophenol. The significant Surface Enhanced Infrared Absorption provided by the nanoprobes, with an absorbance increase up to two orders of magnitude, allowed us to efficiently recognise their inclusion within cells. Notably, the selective and stable SR-microFTIR detection from single cells that have internalised the nanoprobe exhibited clear changes in both shape and intensity of the spectral profile, highlighting the occurrence of biological effects. Flow cytometry, immunofluorescence and murine cytokinesis-block micronucleus assays confirmed the presence of slight but significant cytotoxic and genotoxic events associated with the US-nanoprobe combined treatments. Our results can provide novel hints towards US and nanomedicine combined strategies for cell spectral imaging as well as drug delivery-based therapies.

Fine tuning of the DNAM-1/TIGIT/ligand axis in mucosal T cells and its dysregulation in pediatric inflammatory bowel diseases (IBD).

De-regulated T-cell activation and functions are pivotal in the orchestration of immune-mediated tissue damage in IBD. We investigated the role of DNAM-1 (co-activating)/TIGIT (co-inhibitory)/ligand axis in the regulation of T-cell functions and its involvement in IBD pathogenesis. We show that DNAM-1 and TIGIT display a peculiar expression pattern on gut mucosa T-cell populations, in a microenvironment where their shared ligands (PVR and Nectin-2) are physiologically present. Moreover, DNAM-1 family receptor/ligand system is perturbed in IBD lesions, in a disease activity-dependent manner. The expression profile of CCR6 and CD103 mucosa addressins suggests that microenvironment-associated factors, rather than skewed recruitment of circulating T-cell populations, play a more relevant role in supporting the establishment of DNAM-1 and TIGIT expression pattern in mucosal T-cell populations, and may explain its alteration in IBD. Although both co-receptors mark functionally competent T cells, DNAM-1 and TIGIT segregate on T cells endowed with different proliferative potential. Moreover, their opposing role in regulating T-cell proliferation exquisitely depends on ligand availability. All together, our data propose a role for DNAM-1 and TIGIT in regulating mucosal T-cell activation and immune homeostasis, and highlight the involvement of an imbalance of this system in IBD.

Reduction of autophagy and increase in apoptosis correlates with a favorable clinical outcome in patients with rheumatoid arthritis treated with anti-TNF drugs.

BACKGROUND: Autophagy has emerged as a key mechanism in the survival and function of T and B lymphocytes, and its activation was involved in apoptosis resistance in rheumatoid arthritis (RA). To investigate whether the relationship between autophagy and apoptosis may impact the response to the therapy, we analyzed ex vivo spontaneous autophagy and apoptosis in patients with RA subjected to treatment with anti-tumor necrosis factor (TNF) drugs and in vitro the effects of TNFα and anti-TNF drugs on cell fate. METHODS: Peripheral blood mononuclear cells (PBMCs) from 25 RA patients treated with anti-TNF drugs were analyzed for levels of autophagy marker LC3-II by western blot and for the percentage of annexin V-positive apoptotic cells by flow cytometry. The same techniques were used to assess autophagy and apoptosis after in vitro treatment with TNFα and etanercept in both PBMCs and fibroblast-like synoviocytes (FLS) from patients with RA. RESULTS: PBMCs from patients with RA responsive to treatment showed a significant reduction in LC3-II levels, associated with an increased apoptotic activation after 4 months of therapy with anti-TNF drugs. Additionally, the expression of LC3-II correlated with DAS28. TNFα was able to induce autophagy in a dose-dependent manner after 24 h of culture in RA PBMCs and FLS. Moreover, etanercept caused a significant reduction of autophagy and of levels of citrullinated proteins. CONCLUSIONS: Our results show how the crosstalk between autophagy and apoptosis can sustain the survival of immune cells, thus influencing RA progression. This suggests that inhibition of autophagy represents a possible therapeutic target in RA.

Generation and characterization of two human alpha/beta T cell clones. Recognizing autologous breast tumor cells through an HLA- and TCR/CD3-independent pathway.

Cell-mediated immune response to breast tumor has only been marginally investigated. To gain insight into this issue we have developed two clones of distinct phenotype, CD3+ alpha/beta, CD4+, CD8-, CD16-, and CD3+ alpha/beta, CD4-, CD8+, CD16-, respectively, from peripheral blood lymphocytes (PBL) of a breast cancer patient. These effectors, selected on the basis of their cytolytic activity against autologous tumor cells and lack of lysis on NK-sensitive cell lines, preferentially recognize autologous tumor cells. The two clones' cytotoxic activity, while inhibited by anti-LFA-1 mAb, could not be abolished by mAbs to CD3, to class I and class II MHC molecules, and by mAbs to molecules involved in T cell function (i.e., CD4, CD8, CD2). The molecular structure of the alpha and beta T cell receptor chains of the two effector cells, confirmed their clonality and showed that, despite an overlapping killing pattern, they possess distinct TCR alpha and beta chains. These findings demonstrate that breast tumor-specific CTL clones can be generated through current technology and that a alpha/beta effector cell population operating through a HLA-unrestricted and TCR/CD3-independent pathway may be involved in the identification and killing of this tumor.

Differential effects on membrane permeability and viability of human keratinocyte cells undergoing very low intensity megasonic fields.

Among different therapeutic applications of Ultrasound (US), transient membrane sonoporation (SP) - a temporary, non-lethal porosity, mechanically induced in cell membranes through US exposure - represents a compelling opportunity towards an efficient and safe drug delivery. Nevertheless, progresses in this field have been limited by an insufficient understanding of the potential cytotoxic effects of US related to the failure of the cellular repair and to the possible activation of inflammatory pathway. In this framework we studied the in vitro effects of very low-intensity US on a human keratinocyte cell line, which represents an ideal model system of skin protective barrier cells which are the first to be involved during medical US treatments. Bioeffects linked to US application at 1 MHz varying the exposure parameters were investigated by fluorescence microscopy and fluorescence activated cell sorting. Our results indicate that keratinocytes undergoing low US doses can uptake drug model molecules with size and efficiency which depend on exposure parameters. According to sub-cavitation SP models, we have identified the range of doses triggering transient membrane SP, actually with negligible biological damage. By increasing US doses we observed a reduced cells viability and an inflammatory gene overexpression enlightening novel healthy relevant strategies.

Local activation of immune response in bladder cancer patients treated with intraarterial infusion of recombinant interleukin-2.

Tumor regression induced in cancer patients by i.v. infusion of interleukin-2 (IL-2) is often accompanied by severe side effects. To investigate whether local administration would affect immune response without the side effects, two 5-day cycles of continuous intraarterial [internal iliac artery] infusion of recombinant interleukin-2 (rIL-2) were performed in 12 patients with transitional cell carcinoma (tumor stage 1, node stage 0, metastasis stage 0, and grade 1-2) of the bladder. Four groups of 3 patients were treated at each of 4 escalating doses of rIL-2 (18 x 10(3), 18 x 10(4), 18 x 10(5), and 18 x 10(6) IU/m2/day) throughout the course of the two IL-2 cycles. This treatment was effective in inducing a marked intratumor inflammatory response, consisting mainly of T-lymphocytes and macrophages. A remarkable dose-dependent increase in the levels of soluble CD25 was observed in the urine of all patients, which was associated constantly with an enhanced number of intratumor CD25+ cells. Intratumor macrophages were often immunoreactive for interleukin-1 and/or tumor necrosis factor, suggesting an activated status. Increased levels of soluble CD25 and CD25+ lymphocytes were observed in peripheral blood only at the two highest doses of rIL-2, while increased percentages of circulating HLA-DR+ and CD71+ lymphoid cells and enhancement of CD3+/CD16+ T-lymphocytes were found at lower doses. Peripheral blood eosinophils were augmented in almost all patients but were rarely increased in situ. We provide evidence that continuous intraarterial infusion of rIL-2 activates host immune response, acting preferentially at the tissue level.

Peripheral blood regulatory T cell measurements correlate with serum vitamin D level in patients with psoriasis.

OBJECTIVE: Vitamin D is the precursor of a hormone (1,25-dihydroxyvitamin D3), which has many biological effects in the skin. The immune modulator properties of vitamin D are mediated in part through effects on regulatory T cells (T-reg). Currently, in psoriasis, the relationship between vitamin D and T-reg has not well elucidated. We assess whether vitamin D status is correlated with circulating T-reg in patients affected by psoriasis and if there is a correlation with the severity of the disease evaluated with Psoriasis Area Severity Index (PASI) score. PATIENTS AND METHODS: For each patient we have analyzed, PASI-score, serum levels vitamin D and regulatory T cell percentages. Spearmen's coefficient was used between serum vitamin D levels and the predictors. Subsequently, the independent predictive factors were assessed by Multiple Regression. RESULTS: A total of 26 patients were included in our analysis. Using no parametric Spearman's Coefficient test between serum levels of vitamin D and the single variables, we found an association with T-reg population (p < 0.001) and with PASI-score (p = 0.04). CONCLUSIONS: While vitamin D treatment induces a cytokine profile known to favor the differentiation of T cells with suppressive activity, at the same time, several studies showed how vitamin D can prime for tolerogenic dendritic cells able to favor the differentiation of Treg from T naïve cells. Low levels of vitamin-D may decrease the number of circulatory T-reg, disrupting the immunological homeostasis in psoriatic patients and encouraging the inflammatory activity.

Effects of cadmium on lymphocyte activation.

The effects of cadmium (Cd) on phytohemoagglutinin or phorbol myristate acetate-induced lymphocyte activation were investigated and a dose-dependent inhibition of cell proliferation was found. Kinetic studies revealed that the Cd-sensitive step is an early event of T cell stimulation. Failure of IL2 secretion and reduction of IL2 receptor expression in the Cd-treated cells are also reported. Regardless of which mechanism is responsible for Cd effects, our studies show that the inhibition of lymphocyte activation is associated with reduced [3H]phorbol dibutyrate binding to Ca2+-phospholipid-dependent protein kinase and altered breakdown of phosphatidylinositols. Thus, Cd interferes with two biochemical events which play a critical role in lymphocyte signal transduction and activation.

Heavy-metal modulation of the human intercellular adhesion molecule (ICAM-1) gene expression.

The intercellular adhesion molecule 1 (ICAM-1) can be induced on many different cell types by a set of various modulators (IL1 beta, TNF, LPS, IFN-gamma), which are released during the inflammatory process. We have investigated the possibility that other factors, related to the stress and biophysical perturbations of the inflammatory response, may also modulate ICAM-1. Here, we report that heavy metals, in particular zinc, can enhance the expression of the ICAM-1 gene on cells actively involved at different levels during inflammation. Kinetic studies of ICAM-1 gene expression shows a maximum level of induction 4 h after treatment with metals, followed by a rapid decrease to basal levels within 12 h. The effect on enhanced gene expression is mostly due to a rapid increase of the transcriptional rate as shown by nuclear run-on experiments. In B lymphoblastoid cells, but not in fibroblasts, the increase in RNA expression seems significantly greater that the subsequent increase in protein expression, suggesting that a further point of post-transcriptional regulation of ICAM-1 occurs and may be linked to the cellular specificity. may be linked to the cellular specificity.