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In Italy, according to the International Study on Asthma and Allergies in Childhood study, the prevalence of current asthma, allergic rhinoconjunctivitis, and atopic eczema in 2006 was 7.9%, 6.5%, and 10.1% among children aged 6-7 and 8.4%, 15.5%, and 7.75% among children aged 13-14 yr. University education in this field is provided by the Postgraduate Schools of Pediatrics and those of Allergology and Clinical Immunology, as well as several annual Master courses. The Italian Society of Pediatric Allergology and Immunology (SIAIP) was founded in 1996 and counts about 1000 members. SIAIP promotes evidence-based management of allergic children and disseminates information to patients and their families through a quite innovative website and the National Journal 'Rivista Italiana di Allergologia Pediatrica'. In the last decade, four major regional, inter-regional, and national web-based networks have been created to link pediatric allergy centers and to share their clinical protocols and epidemiologic data. In addition, National Registers of Primary Immune-deficiencies and on Pediatric HIV link all clinical excellence centers. Research projects in the field of pediatric allergy and immunology are founded by the Italian Ministry of Education, University and Research (MIUR) and by the National Research Council (CNR), but the overall investments in this research area are quite low. Only a handful Italian excellence centers participate in European Projects on Pediatric Allergy and Immunology within the 7th Framework Program. The European Academy of Allergy and Clinical Immunology currently hosts two Italians in its Executive Committee (EC) and one in the EC of the Pediatric Section; moreover, major European Academy of Allergy and Clinical Immunology meetings and courses in the area of pediatrics (e.g., PAAM, Venice, 2009) have been held in Italy in the last 3 yr. Italian hallmarks in the management of allergic diseases in childhood are a quite alive and spread interest in Molecular Allergology and a remarkable predominance of sublingual (SLIT) compared to the subcutaneous (SCIT) immunotherapy.

Subjects with Down syndrome (DS) have a high mortality due to infections and a high risk of developing malignancies. These observations, together with the demonstration of a frequent occurrence of HBsAg carrier state and of autoantibodies, have prompted investigations of the immune function in DS. Thymic morphological and functional abnormalities have been demonstrated. Peripheral blood mononuclear cells of DS subjects have been shown to include a high number of T lymphocytes with low avidity for sheep erythrocytes and a very high percentage of cells with an NK phenotype. However, NK activity is low in DS. Production of some important cytokines, such as IL-2, is depressed, thus contributing to T-cell derangement. Abnormalities of the B-cell compartment were also demonstrated, with a tendency towards high IgG and low IgM serum levels. Controversial results have been obtained with regard to antigen-specific antibody response. Also phagocytes of DS subjects display some characteristic functional impairments, with low chemotactic ability and reduced production of oxygen radicals. Despite the clearly established and rather detailed evidence of immune derangements, therapeutic trials have been anecdotal and resulted in marginal effects. HBV vaccination is highly advisable in DS because of the high risk of becoming chronic HBV carriers once infected.

Cytokines play a major role in lymphoid development. Defects of the common gamma chain (gamma(c)) or of the JAK3 protein in humans have been shown to result in a severe combined immune deficiency (SCID), with a profound defect in T and natural killer (NK)-cell development, whereas B-cell generation is apparently unaffected (T-B+NK-SCID). While extensive molecular and biochemical analysis of these patients has been instrumental in understanding better the biological properties of the gamma(c) and JAK3 protein, an unexpected phenotypic heterogeneity of gamma(c) and JAK3 deficiency has emerged, indicating the need for appropriate and extensive investigations even in patients with atypical presentations. At the same time, characterization of the defects has been instrumental in the development of novel therapeutic approaches, from in utero hematopoietic stem cell transplantation to gene therapy.

Background Cow's milk allergy is quite frequent in the first years of human life. When breast-feeding is not possible, a cow's milk substitute must be provided for allergic subjects. Different alternatives to cow's milk have been suggested as protein sources (soy, hydrolysed proteins, goat's milk, etc.), but all these dietetic solutions are not without risks for polyallergic or more sensitive subjects. Objective To obtain new information on the suitability of other mammalian milks for allergic children, we evaluated the cross-reactivity between milk proteins from different animal species. Methods Milk samples were analysed by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). To detect antibody–antigen complexes, immunoblotting was performed by using sera from children allergic to cow's and ewe's milk (RAST class ≥ 4) and monoclonal antibodies (MoAb) specific for bovine proteins (caseins and β-lactoglobulin). Results IgEs from children allergic to cow's milk are capable of recognizing most part of milk proteins from mammals bred in European countries (ewe, goat, buffalo), while no serum used in this study contains IgEs reacting with camel's milk proteins. Camel's milk was also not recognized from circulating IgEs from a child specifically allergic to ewe's milk. Specific antibovine monoclonal antibodies cross-reacted with proteins from other mammalian species, apart from those of camel. Conclusions Homologies in amino acidic composition could justify the cross-reactivity observed between proteins from different animal species. On the other hand, the phylogenetic difference could be responsible for the failed recognition of camel's proteins by circulating IgEs and monoclonal antibodies.

Omenn's syndrome (OS) is characterized by erythrodermia, hepatosplenomegaly, lymphadenopathy, hypereosinophilia and elevated IgE levels associated with increased susceptibility to severe infections. Peripheral blood T cells, though usually present in normal number, show an activated phenotype (including an increased expression of CD95/Fas), a Th2 pattern of cytokine secretion and defective proliferative response to mitogens. In this report, we demonstrate that T cells from patients with OS undergo an excessive cell death in vitro resulting from two mechanisms. First, a substantial number of peripheral blood lymphocytes from OS patients die in unstimulated cultures (p = 0.009 vs. healthy controls). This spontaneous apoptosis is associated with reduced expression of bcl-2 gene product (p < 0.05) and can be prevented by addition of interleukin (IL)-2 (which also prevents down-modulation of bcl-2), while is independent from CD95 signaling. Second, lymphocytes from OS patients are highly susceptible to activation-induced cell death (AICD) induced with mitogens. This mechanism is largely independent from IL-2, while it can be significantly inhibited blocking CD95 with an IgG2b monoclonal antibody (mAb). The dependence of AICD from signals transduced via CD95 was confirmed showing that cross-linking CD95 with an IgM mAb induces a higher cell death in purified CD4+ CD45R0+ cells from OS patients than in controls (comparable for CD95 expression). Both mechanisms of cell death observed in this study result from lymphocyte hyperactivation occurring in vivo in these patients and may contribute to functional T cell defects of OS.

Background Hydrolysed casein and whey protein formulas have been developed with the aim of preventing sensitization in infants at high risk of cow milk allergy. Subsequently these products have also been used for treatment of children with cow milk allergy. However, severe reactions have occurred in some allergic infants fed with these formulas raising doubts about their absolute safety and suggest the need for developing in vitro techniques for detection of eventual residual allergenic activity in such preparations. Objectives Our purpose was to evaluate the usefulness of monoclonal and polyclonal antibodies against casein components (alpha, beta and kappa casein) as reagents for the detection of the residual antigenic activity of casein components in several hydrolysed formulas. Methods The monoclonal and polyclonal antibodies were produced according to standard procedures by immunizing female Balb/c mice with casein fraction (a mixture of alpha, beta and kappa casein). ELISA assays were developed to test the specificity of the antibodies and to detect and evaluate the amount of residual antigenic activity of the casein components in hydrolysed formulas. Results Use of polyclonal antiserum specific for casein allowed detection of residual antigenic activity of casein components in all partial hydrolysates and in the two extensive whey protein hydrolysates in the amounts ranging from 0.05 to 0.67% of total protein. No such activity was detectable in either the two extensive casein hydrolysates tested or the aminoacid based formula. The polyclonal antiserum proved to be more suitable than monoclonals for detecting residual antigenic activity in the hydrolysates. The monoclonal antibodies were directed against epitopes expressed on different casein components. Conclusions In this study the ELISA inhibition assay with polyclonal antibodies specific for casein components of cow milk proved to be a sensitive method for estimating residual antigenicity in the hydrolysed formulas commercially available for infants with cow milk allergy suggesting their potential application for the quality control of hypoallergenic infant formulas.

Background: Common Variable Immunodeficiency (CVID) is characterized by defective antibody production. This has been variably attributed to intrinsic B-cell defects or to T-cell disfunctions. Recently, it has been reported that the expression of the CD40 Ligand (CD40L), a T-cell surface molecule that plays a critical role in the cell-contact-mediated helper signals provided to B-cells, is defective in a subset of patients with CVID. Methods: To demonstrate that the defective expression is due to intrinsic functional abnormalities of CD4+ lymphocytes, CD4+ cells were purified from eight patients with CVID and eight age-paired controls, stimulated with PMA+Ionomycin. and studied for CD40L expression by flow cytome-try using specific monoclonal antibodies. Results and conclusions: The percentage of CD4+ cells expressing CD40L after optimal stimulation was correlated with age both in patients with CVID (r: 0. 74: p: 0.04) and in healthy controls (r: 0. 73; p: 0.04). The percentage of CD40L+ cells was reduced in patients with CVID compared to that of controls (p: 0.02 when data are paired for age) with a reduced density of expression (p: < 0.01). The defect was variable in different patients and in some cases it was marginal.

In an attempt to define a biological marker of monocyte hyperactivation in the course of infantile histiocytosis, the spontaneous nitroblue tetrazolium (NBT) reduction assay was applied to monocytes from 13 children with Langerhans cell histiocytosis (LCH), familial haemophagocytic lymphohistiocytosis (FHL), juvenile xanthogranuloma or malignant histiocytosis. Significant increase in NBT reduction was observed in the patients with both active LCH and FHL in comparison with control subjects, who were either healthy or affected by different conditions. A close relationship between spontaneous reduction rate and clinical condition of the patients was evident in patients tested at diagnosis, during remission and during the course of disease reactivation. Interleukin-1 (IL-1) production by monocytes was also evaluated: the patients with LCH and FHL displayed a significant increase in in vitro IL-1 production by lipopolysaccharide-stimulated monocytes. In our experience the spontaneous NBT reduction assay was a sensitive, quite specific, low-cost and reproducible test for the evaluation of children with histiocytosis. Useful information may be obtained at diagnosis but also during the clinical course of disease by using this marker of monocyte spontaneous activation.

Summary Secretory-IgA (SIgA) concentrations were determined in whole saliva, unstimulated or stimulated by lemon juice, of thirty-eight children with atopic dermatitis, which comprised three adolescents, sixteen with IgE detected by RAST to one or more common allergen and twenty-two without specific IgE by RAST. There were thirty healthy controls matched for age and sex. The mean amount of total IgE was significantly greater in the RAST-positive than in the RAST-negative group. The mean SIgA concentration in unstimulated saliva of the RAST-positive atopic dermatitis group was less than that of the RAST-negative atopic dermatitis group and control groups, though the mean concentrations of SIgA of stimulated saliva were not significantly different in the three groups. It is suggested that the pathogenesis of atopic dermatitis may differ in children with or without specific-IgE antibodies; in those who were RAST-positive deficient exclusion of allergen by the intestinal barrier contributed to the pathogenesis, but not in those who were RAST-negative.

In 20 healthy premature infants, 10 of whom were administered a total dose of 120 mg/kg vitamin E intramuscularly during the first 13 days after birth, polymorphonuclear leukocyte (PMN) bactericidal activity, frequency and index of phagocytosis, NBT reduction, random movement, chemotaxis and metabolic activity were evaluated within the first 48 hours and again at 5, 14 and 30 days of age. PMN function was also assessed in 30 adult controls. In the treated and untreated infants no differences were found in PMN function before treatment with vitamin E; however phagocytosis, bactericidal activity and chemotaxis were significantly lower than in the adult controls. At 5 days of age, in the untreated infants both index and frequency of phagocytosis remained low but in the treated groups increased significantly. At 14 and 30 days phagocytosis was normal in both treated and untreated infants. No differences were found in the bactericidal activity, NBT reduction, random movement, chemotaxis or metabolic activity of the treated and untreated infants at the ages studied. This preliminary report suggests that vitamin E may be used in premature newborns for accelerating normalization of phagocytic function in the neonatal period.

We studied the interaction of lipopolysaccharide (LPS) and concanavalin A (Con A) with regard to IgM and IgG production in in vitro cultures of human peripheral blood lymphocytes (PBL). In our system LPS alone over a wide range of concentrations did not stimulate detectable IgM or IgG production, while Con A at optimal (6 microgram/ml) and suboptimal (0.6 microgram/ml) mitogenic concentrations induced synthesis of small amounts of Ig. A marked enhancing effect was present when both Con A and LPS were added to the cultures. The different doses of LPS has similar effects on both classes of Ig, and typical dose-response curves were obtained. To evaluate the cellular basis of this synergism, the effect on cell proliferation was studied under identical experimental conditions in normal subjects and patients with X-linked agammaglobulinaemia (X-LA). Parallel cultures were set up after monocyte depletion by adherence on Petri dishes. On day 3, increasing doses of LPS were associated with progressive decreases in 3H-thymidine (3H-TdR) incorporation. Similar results were obtained with normal lymphocytes and those from X-LA patients. Monocyte depletion did not substantially alter the lymphocyte response pattern. The preferential induction of helper activities, either directly by helper stimulation or indirectly by suppressor inhibition, is suggested as a possible mechanism of the interaction observed.

A simple technique has been worked out for the simultaneous assessment of cell proliferation, generation of plasma cells, plaque-forming cells and immunoglobulin production from single cultures of 1 X 10(6) human peripheral blood lymphocytes stimulated with pokeweed mitogen (PWM). Kinetic studies showed that number of cells, thymidine incorporation rate, number of plaque-forming cells against fluorescein-isothiocyanate-haptenated sheep erythrocytes and concentration of IgM and IgG in the supernatant peaked on the average between day 5 and 7 of culture. This technique is particularly suitable for the analysis of hypogammaglobulinaemias in infancy.