Epigenetic Mechanisms of Inflammasome Regulation.

The innate immune system represents the host's first-line defense against pathogens, dead cells or environmental factors. One of the most important inflammatory pathways is represented by the activation of the NOD-like receptor (NLR) protein family. Some NLRs induce the assembly of large caspase-1-activating complexes called inflammasomes. Different types of inflammasomes have been identified that can respond to distinct bacterial, viral or fungal infections; sterile cell damage or other stressors, such as metabolic imbalances. Epigenetic regulation has been recently suggested to provide a complementary mechanism to control inflammasome activity. This regulation can be exerted through at least three main mechanisms, including CpG DNA methylation, histones post-translational modifications and noncoding RNA expression. The repression or promotion of expression of different inflammasomes (NLRP1, NLRP2, NLRP3, NLRP4, NLRP6, NLRP7, NLRP12 and AIM2) through epigenetic mechanisms determines the development of pathologies with variable severity. For example, our team recently explored the role of microRNAs (miRNAs) targeting and modulating the components of the inflammasome as potential biomarkers in bladder cancer and during therapy. This suggests that the epigenetic control of inflammasome-related genes could represent a potential target for further investigations of molecular mechanisms regulating inflammatory pathways.

Relationship between serum myostatin levels and carotid-femoral pulse wave velocity in healthy young male adolescents: the MACISTE study.

Serum myostatin (sMSTN) is a proteic compound that regulates skeletal muscle growth, adipogenesis, and production of extracellular matrix. Its relationship with functional and structural properties of the arterial wall is still understudied. We aimed at evaluating the association between sMSTN and carotid-femoral pulse wave velocity (cf-PWV), a measure of aortic stiffness, in a cohort of healthy male adolescents. Fifteen healthy male adolescents were recruited among the participants of the Metabolic And Cardiovascular Investigation at School, TErni (MACISTE) study, a cross-sectional survey conducted at the "Renato Donatelli" High School in Terni, Italy. sMSTN was measured through enzyme-linked immunosorbent assay. cf-PWV was measured through high-fidelity applanation tonometry. Muscle strength and body composition were measured through handgrip and bioimpedentiometry, respectively. sMSTN levels showed a skewed distribution (median: 6.0 ng/mL, interquartile range: 2.2-69.2 ng/mL). Subjects with sMSTN above median value showed higher values of brachial diastolic blood pressure and increased cf-PWV (6.1 ± 1.1 m/s vs. 4.6 ± 0.7 m/s, P < 0.01) values, compared with their counterparts. Such difference remained significant after controlling for age, mean BP, heart rate, body mass index z-score, waist-to-height ratio, body mass/lean mass ratio, and amount of physical activity (P = 0.02). The association between log-transformed sMSTN and cf-PWV was direct and linear, and independent from the effect of confounders at the multivariate analysis (P = 0.02). In this preliminary report, sMSTN was independently associated with cf-PWV, a measure of aortic stiffness, in healthy male adolescents. Our results shed lights on the potential role of myokines in the pathogenesis of systemic hypertension and atherosclerosis.NEW & NOTEWORTHY Serum myostatin, a proteic compound known to regulate skeletal muscle growth and production of extracellular matrix, is independently associated with increased aortic stiffness in healthy male adolescents. This result sheds lights on the potential novel role of myokines in the early development of systemic hypertension and early vascular aging, as well as on their inhibition as a hypothetical therapeutic strategy to counteract vascular aging at an early stage of physical development.

Serum IgE reactivity profiling in an asthma affected cohort.

BACKGROUND: Epidemiological evidence indicates that atopic asthma correlates with high serum IgE levels though the contribution of allergen specific IgE to the pathogenesis and the severity of the disease is still unclear. METHODS: We developed a microarray immunoassay containing 103 allergens to study the IgE reactivity profiles of 485 asthmatic and 342 non-asthmatic individuals belonging to families whose members have a documented history of asthma and atopy. We employed k-means clustering, to investigate whether a particular IgE reactivity profile correlated with asthma and other atopic conditions such as rhinitis, conjunctivitis and eczema. RESULTS: Both case-control and parent-to-siblings analyses demonstrated that while the presence of specific IgE against individual allergens correlated poorly with pathological conditions, particular reactivity profiles were significantly associated with asthma (p<10E-09). An artificial neural network (ANN)-based algorithm, calibrated with the profile reactivity data, correctly classified as asthmatic or non-asthmatic 78% of the individual examined. Multivariate statistical analysis demonstrated that the familiar relationships of the study population did not affect the observed correlations. CONCLUSIONS: These findings indicate that asthma is a higher-order phenomenon related to patterns of IgE reactivity rather than to single antibody reactions. This notion sheds new light on the pathogenesis of the disease and can be readily employed to distinguish asthmatic and non-asthmatic individuals on the basis of their serum reactivity profile.

An antigen microarray immunoassay for multiplex screening of mouse monoclonal antibodies.

The mouse monoclonal antibody (mAb) technology still represents a key source of reagents for research and clinical diagnosis, although it is relatively inefficient and expensive and therefore unsuitable for high-throughput production against a vast repertoire of antigens. In this article, we describe a protocol that combines the immunization of individual mice with complex mixtures of influenza virus strains and a microarray-based immunoassay procedure to perform a parallel screening against the viral antigens. The protocol involves testing the supernatants of somatic cell hybrids against a capture substratum containing an array of different antigens. For each fusion experiment, we carried out more than 25,000 antigen-antibody reactivity tests in less than a week, a throughput that is two orders of magnitude higher than that of traditional antibody detection assays such as enzyme-linked immunosorbent assays and immunofluorescence. Using a limited number of mice, we can develop a vast repertoire of mAbs directed against nuclear and surface proteins of several human and avian influenza virus strains.

Influence of amino acids on low-density Escherichia coli responses to nutrient downshifts.

A vast bibliography on nutrient effects on high-density cultures exists, while it has been overlooked that low densities of starving cells are often the rule in natural environments. By means of a novel sensitive beta-galactosidase assay, we examined Escherichia coli transitions to minimal media when the cell concentration was 100 to 10,000 cells per ml. As in high-density cultures, the enzyme activity depended on amino acid availability and was subject to catabolite repression and stringent control. In all conditions tested, despite the presence of other nutrient sources, the relationship between beta-galactosidase activity and the l-amino acid pool was hyperbolic. The affinity constant when the amino acid pool was the only nutrient source averaged 14 muM after 90 min and increased up to 222 muM after 4.5 h. While investigating the transition from lag phase to exponential phase, we observed that the cells did not enter into starvation mode in the presence of amino acids, even when the nutrient amount was insufficient to support full survival. Based on these premises, the switch from starvation to hunger was investigated in relation to the amino acid pools. A critical range of concentrations at which E. coli linearly synthesized beta-galactosidase despite, at the same time, suffering a large decrease in cell viability was then recognized. Since both beta-galactosidase production and the dilution rate were reduced by more than half in the absence of leucine, we examined the contribution of leucine to cell recovery capabilities.