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Primary immunodeficiencies (PIDs) are heterogeneous disorders, characterized by variable clinical and immunological features. National PID registries offer useful insights on the epidemiology, diagnosis, and natural history of these disorders. In 1999, the Italian network for primary immunodeficiencies (IPINet) was established. We report on data collected from the IPINet registry after 20 years of activity. A total of 3352 pediatric and adult patients affected with PIDs are registered in the database. In Italy, a regional distribution trend of PID diagnosis was observed. Based on the updated IUIS classification of 2019, PID distribution in Italy showed that predominantly antibody deficiencies account for the majority of cases (63%), followed by combined immunodeficiencies with associated or syndromic features (22.5%). The overall age at diagnosis was younger for male patients. The minimal prevalence of PIDs in Italy resulted in 5.1 per 100.000 habitants. Mortality was similar to other European registries (4.2%). Immunoglobulin replacement treatment was prescribed to less than one third of the patient cohort. Collectively, this is the first comprehensive description of the PID epidemiology in Italy.

Background X-linked agammaglobulinemia (XLA) is the prototype of primary humoral immunodeficiencies. Long-term follow-up studies regarding disease-related complications and outcome are scarce. Objective Our aim was to describe the natural history of XLA. Methods A nationwide multicenter study based on the Italian Primary Immunodeficiency Network registry was established in 2000 in Italy. Affected patients were enrolled by documenting centers, and the patients' laboratory, clinical, and imaging data were recorded on an annual base. Results Data on the patients (N = 168) were derived from a cumulative follow-up of 1370 patient-years, with a mean follow-up of 8.35 years per patient. The mean age at diagnosis decreased after establishment of the Italian Primary Immunodeficiency Network registry (84 months before vs 23 months after). Respiratory, skin, and gastrointestinal manifestations were the most frequent clinical symptoms at diagnosis and during long-term follow-up. Regular immunoglobulin replacement treatment reduced the incidence of invasive infections. Affected patients developed chronic lung disease over time (47% after 40 years of follow-up) in the presence of chronic sinusitis (84%). Malignancies were documented in a minority of cases (3.7%). Overall survival for affected patients was significantly reduced when compared with that for the healthy male Italian population, and it further deteriorated in the presence of chronic lung disease. Conclusions This is the first detailed long-term follow-up study for patients with XLA, revealing that although immunoglobulin replacement treatment reduces the incidence of invasive infections, it does not appear to influence the development of chronic lung disease. The overall survival of affected patients is reduced. Further studies are warranted to improve patients' clinical management and increase awareness among physicians.

Sediment samples from 24 tributaries of the Po River (Italy) were screened for selected trace elements (Cd, Cu, Hg, Pb, and Zn) and extractable organic compounds; a proxy for contamination by organic microcontaminants. The toxicity of sediment extracts was evaluated using a battery of biotests (Dugesia gonocephala, Paracentrotus lividus, and Tamnocephalus platyurus). Contamination by trace elements (including very high Hg pollution – 4 to 16ppm total Hg – in one sub-basin) reached potentially harmful levels only in the sediments of four tributaries; while contamination by organic microcontaminants was present in most sub-basins. Sediments from most study sites did actually show signs of anthropogenic stress and were able to elicit a toxic response. A more detailed evaluation of sediment quality in the Po River tributaries seems to be urgently needed for developing the necessary remediation strategies. Research priorities should include more thorough testing of sediment toxicity, determination of metal background levels in the various sub-basins and a more detailed identification of the organic micropollutants of possible concern.

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.

We assessed the knowledge of Italian mothers of adolescent girls about HPV and HPV vaccination, their willingness to immunize their daughters, and their perception of the role of different medical specialists in the HPV immunization strategy by a telephone interview. Fifty-four percent of the 807 interviewed mothers reported to have ever heard about HPV, and 84% of them were willing to immunize their daughters. Pediatricians most frequently provided information on HPV vaccine (31%), and were perceived as the preferred immunization providers (77%). Acceptance of HPV immunization was high and was not associated with knowledge of HPV.

Mannose binding lectin (MBL) is a soluble pattern recognition receptor of innate immunity that binds a wide range of pathogens and exerts opsonic effects. We investigated the association between serum MBL levels and development of sepsis in infants admitted to neonatal intensive care units (NICUs). Serum MBL levels on admission were measured by enzyme-linked immunosorbent assay (ELISA) in 206 neonates consecutively admitted to an NICU of whom 138 did not develop hospital-acquired sepsis and 68 did. Of these 68, 40 had confirmed sepsis with positive blood cultures, 19 clinically suspected sepsis, with negative blood cultures, and nine had clinically suspected sepsis with blood culture yielding coagulase-negative staphylococci (CoNS). Serum MBL levels on admission were significantly lower in infants with sepsis [0.45 microg/mL; interquartile range (IQR) 0.09-1.68], particularly in those with confirmed sepsis (0.17 microg/mL; IQR 0.05-0.96), compared with infants without sepsis (1.45 microg/mL; IQR 0.43-3.52), and infants with CoNS-positive blood culture (1.70 microg/mL: IQR 0.85-3.60). After adjusting for duration of exposure gestational age (GA) and birth weight (BW), the association of low MBL levels with development of sepsis was maintained [odds ratio (OR) = 0.52; 95% confidence interval (CI): 0.36-0.75]. The measurement of serum MBL levels on admission in NICU may help to identify neonates at higher risk of developing sepsis.

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.

We retrospectively analyzed the outcome of hematopoietic stem cell transplantations (HSCT) performed at our Center between 1991 and 2002 in 11 unselected patients with Omenn syndrome, a variant of severe combined immunodeficiency. The patients' mean age at the time of the first HSCT was 8.4 months. Two patients received two, and one patient three, HSCT procedures. The resulting 15 HSCT derived in seven cases from HLA-haploidentical parents, in four patients from matched unrelated donors, in three cases from an HLA phenotypically identical related donor, and in one case from an HLA genotypically identical family donor. Nine out of 11 patients are alive and immunoreconstituted 30-146 months after transplantation. At the time of the most recent evaluation, all of the nine survivors had normal T-cell function, and eight of them had developed normal antibody production. This study demonstrates an overall mortality of 18.2%, which is substantially lower than previously reported. Early recognition of OS, rapid initiation of adequate supportive treatment and HSCT lead to improved outcome for this otherwise fatal disease, regardless of the origin and matching of hematopoietic stem cells.

We report a family with pyogenic sterile arthritis, pyoderna and acne syndrome (PAPA). The proband presented several episodes of sterile pyogenic arthritis and became unresponsive to glucocorticoids. After treatment with the tumor necrosis factor inhibitor etanercept, the disease underwent rapid and sustained clinical remission. Production of tumor necrosis factor-α by mononuclear cells of the proband and of the affected relatives was abnormally elevated.

Objective: To describe the treatment and long term outcome after immunosuppressive treatment of children with myocarditis. Methods and results: 114 patients with newly diagnosed dilated cardiomyopathy were divided into three groups, according to the histological pattern: group A, acute myocarditis; group B, borderline myocarditis; and group C, non-inflammatory cardiomyopathy. Groups A and B were treated with cyclosporine and prednisone in addition to conventional treatment. Survivors of the whole cohort were analysed for 13 year transplant-free survival and assessed for left ventricular function. Event-free survival at 13 years was 97 (3)% for group A, 70 (8)% for group B, and 32 (7)% for group C (p < 0.0001). It was 96 (4)% at one year and 83 (5)% at 13 years for the cumulated myocarditis group (A and B). Cardiac function recovered completely in 79% of survivors in group A, 64% in group B, and 36% in group C. The rate of complete recovery in the cumulated group (A and B) was 70%. Conclusions: The high long term survival rate of this cohort of children with myocarditis is probably due to the effect of short term immunosuppression. This result differs from previously published series of conventionally treated children, whose survival probability at one year was about 60%.

Grain-size distribution, major elements, nutrients and trace metals were determined in bed sediments and suspension collected at 10 representative sites along the river Po under normal and high flow conditions. Grain-size distribution and major element composition of suspension highlighted the presence of two distinct particle populations in the upper-middle Po (coarser particles, lower carbonate content) and in the lower Po (finer particles, higher carbonate content). This change partly reflects the geological differences between the two parts of the basin, and also the presence of a hydroelectric power plant at Isola Serafini (Piacenza). With respect to environmental quality issues, bed sediments and suspension provide similar results. A moderate nutrient pollution is found in all but the uppermost parts of the river basin, while the most significant inputs of trace metals appear to originate from the urban areas of Turin and Milan. Calculation of sediment enrichment factors identifies Cd, Cu, Hg and Zn as the most impacted elements by human activities. On the other hand, the high levels of Ni and Cr throughout the river seem to derive mainly from the presence of basic rocks in the upper and middle parts of the basin. Both nutrient and trace metal particulate concentrations substantially decrease under high flow conditions possibly due to "flushing" of contaminated bed sediments and resuspension of coarser material. Under normal flow conditions, water hydrochemistry and concentrations of some elements (As, Ca, Cr, Cu, K, Mg, Mn, Na, Ni, and Pb) in the dissolved phase (

The biological activity of polysaccharides from the mycelia of 40 Basidiomycetes was studied using an uncommon toxicity test technique, the planaria bioassay, and the better known potato disk bioassay. The results showed the utility of this duo of 'in vitro' tests as a preliminary screening of the toxicity of substances that are present in aqueous fungal extracts.

An in utero paternal CD34(+) cell transplant was performed in a T-B+NK+ SCID fetus. We report here the results of the 3-year humoral immune reconstitution study. The methods used were ApoB VNTR typing, flow cytometry, nephelometry, hemagglutination, ELISA, ELISPOT and lymphoproliferative assays. The T cells were of donor origin whereas monocytes, B and NK cells were of host origin. Peripheral B cell counts and IgM levels were normal since birth. IVIG therapy was required at 5 months of age until 2 years old. IgA levels > or =20 mg/dl were detected from month 17 post transplantation. Isohemagglutinins were present since month 8 post transplantation, the highest titers (anti-A:1/128, anti-B:1/32) were obtained at month 33 post-transplantation. After immunization with rHBsAg, circulating anti-HBsAg IgG secreting cells and a 7.8-fold increase in serum anti-HBsAg Ab were detected. We conclude that split chimerism following in utero haploidentical BMT allows complete humoral immune reconstitution in a T-B+NK+ SCID patient.

Children with Down syndrome (DS) have increased susceptibility to infections and a high frequency of leukemia and autoimmune disorders, suggesting that immunodeficiency and immune dysfunction are integral parts of the syndrome. A reduction in B-cell numbers has been reported, associated with moderate immunodeficiency and normal immunoglobulin levels. Here, we compared B-cell populations of 19 children with DS with those in healthy age-matched controls. We found that all steps of peripheral B-cell development are altered in DS, with a more severe defect during the later stages of B-cell development. Transitional and mature-naive B-cell numbers are reduced by 50% whereas switched memory B cells represent 10–15% of the numbers in age-matched controls. Serum IgM levels were slightly reduced, but all other immunoglobulin isotypes were in the normal range. The frequency of switched memory B cells specific for vaccine antigens was significantly lower in affected children than in their equivalently vaccinated siblings. In vitro switched memory B cells of patients with DS have an increased ability to differentiate into antibody-forming cells in response to TLR9 signals. Tailored vaccination schedules increasing the number of switched memory B cells may improve protection and reduce the risk of death from infection in DS.

CD40 is a member of the tumor necrosis factor receptor superfamily, expressed on a wide range of cell types including B cells, macrophages, and dendritic cells. CD40 is the receptor for CD40 ligand (CD40L), a molecule predominantly expressed by activated CD4+T cells. CD40/CD40L interaction induces the formation of memory B lymphocytes and promotes Ig isotype switching, as demonstrated in mice knocked-out for eitherCD40LorCD40gene, and in patients with X-linked hyper IgM syndrome, a disease caused byCD40L/TNFSF5gene mutations. In the present study, we have identified three patients with an autosomal recessive form of hyper IgM who fail to express CD40 on the cell surface. Sequence analysis ofCD40genomic DNA showed that one patient carried a homozygous silent mutation at the fifth base pair position of exon 5, involving an exonic splicing enhancer and leading to exon skipping and premature termination; the other two patients showed a homozygous point mutation in exon 3, resulting in a cysteine to arginine substitution. These findings show that mutations of theCD40gene cause an autosomal recessive form of hyper IgM, which is immunologically and clinically undistinguishable from the X-linked form.

Severe combined immunodeficiency (SCID) comprises a heterogeneous group of primary immunodeficiencies, a proportion of which are due to mutations in either of the 2 recombination activating genes (RAG)-1 and -2, which mediate the process of V(D)J recombination leading to the assembly of antigen receptor genes. It is reported here that the clinical and immunologic phenotypes of patients bearing mutations in RAGs are more diverse than previously thought and that this variability is related, in part, to the specific type of RAG mutation. By analyzing 44 such patients from 41 families, the following conclusions were reached: (1) null mutations on both alleles lead to the T-B-SCID phenotype; (2) patients manifesting classic Omenn syndrome (OS) have missense mutations on at least one allele and maintain partial V(D)J recombination activity, which accounts for the generation of residual, oligoclonal T-lymphocytes; (3) in a third group of patients, findings were only partially compatible with OS, and these patients, who also carried at least one missense mutation, may be considered to have atypical SCID/OS; (4) patients with engraftment of maternal T cells as a complication of a transplacental transfusion represented a fourth group, and these patients, who often presented with a clinical phenotype mimicking OS, may be observed regardless of the type of RAG gene mutation. Analysis of the RAG genes by direct sequencing is an effective way to provide accurate diagnosis of RAG-deficient as opposed to RAG-independent V(D)J recombination defects, a distinction that cannot be made based on clinical and immunologic phenotype alone.

Background: Regurgitation and vomiting are common manifestations of cow's milk protein allergy (CMPA) in infants and are usually ascribed to gastroesophageal reflux (GER). Gastric anaphylaxis can induce antral dysmotility in the rat, and therefore the hypothesis for the current study was that cow's milk in sensitized infants may impair antral motility, thereby promoting GER and reflex vomiting. Methods: Seven vomiting infants with CMPA and nine with primary GER underwent a challenge with cow's milk formula. Electrogastrography (EGG) was used to measure the spectral frequency (bradygastria = 1.5-2.4 cycles per minute [cpm], normogastria = 2.5-3.9 cpm, tachygastria = 4.0-9.0 cpm) and the postprandial-to-fasting power ratio of gastric electrical activity, whereas gastric half-emptying time (T1/2) was measured by electrical impedance tomography (EIT). Results: In CMPA and GER, respectively, during fasting, the frequency distribution (mean ± SD) of the EGG was as follows: normogastria 47.9% ± 12.5% versus 52.2% ± 9.8%, bradygastria 24.1% ± 5.7% versus 22.8% ± 8.3%, and tachygastria 28.0% ± 8.5% versus 25.0% ± 8.3%. In contrast, after the cow's milk challenge, the difference between the two groups was statistically significant: normogastria 33.1% ± 8.8% versus 70.6°% ± 8.6% (P < 0.0001), bradygastria 38.0% ± 15.5% versus 15.7% ± 5.2% (P = 0.002), and tachygastria 28.9% ± 10.6% versus 13.4% ± 4.6% (P = 0.001. The postprandial/ fasting power ratio (mean ± SD) was 3.2 ± 1.9 in CMPA and 8.1 ± 2.1 in GER (P < 0.0001). Gastric T1/2 (mean ± SD) of the cow's milk meal was 89.0 ± 26.3 minutes versus 54.0 ± 12.6 minutes (P 0.003). In infants with GER all EGG parameters and gastric T1/2 were similar to that in 10 healthy control infants. Conclusions: In sensitized infants, cow's milk induces severe gastric dysrhythmia and delayed gastric emptying, which in turn may exacerbate GER and induce reflex vomiting. Electrogastrography and EIT can be useful in the assessment of vomiting, GER, and CMPA in infants.

Polyarteritis or panarteritis nodosa (PAN) is a necrotizing, focal segmental vasculitis that affects predominantly medium-sized arteries in many different organ systems. It is extremely rare in childhood. Involvement of the oral mucosa at diagnosis is uncommon in PAN. Here, we report a case of a pediatric patient with tongue necrosis.

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.

The activation-induced cytidine deaminase (AID) gene, specifically expressed in germinal center B cells in mice, is a member of the cytidine deaminase family. We herein report mutations in the human counterpart of AID in patients with the autosomal recessive form of hyper-IgM syndrome (HIGM2). Three major abnormalities characterize AID deficiency: (1) the absence of immunoglobulin class switch recombination, (2) the lack of immunoglobulin somatic hypermutations, and (3) lymph node hyperplasia caused by the presence of giant germinal centers. The phenotype observed in HIGM2 patients (and in AID−/− mice) demonstrates the absolute requirement for AID in several crucial steps of B cell terminal differentiation necessary for efficient antibody responses.

Oral complications are a significant cause of morbidity and potential mortality for children undergoing hematopoietic cell transplant (HCT). Oral complications can occur at all stages of HCT and can interfere significantly with transplant recovery. Mucosal disease caused by conditioning regimen toxicity and infection are frequent clinical problems. Untreated dental caries and periodontal disease may result in severe infections of the mouth and/or life-threatening systemic spread of the microbial pathogens. In the course of chronic graft-versus-host disease (GVHD), which can complicate HCT, lichenoid and ulcerative lesions of the mucosa are observed. Furthermore, total-body irradiation utilized in the conditioning regimens can cause early xerostomia and consequent dental decay and also result in significant dental and skeletal developmental anomalies. The dental health care team should have a key role in the support of HCT patients. The team's primary responsibilities are those related to the prevention of severe infections originating in the mouth, which includes providing instruction on oral prophylaxis and hygiene as well as direct intervention. Prevention and/or diagnosis and management of oral complications of HCT by the dental team can improve the success of a transplant by reducing morbidity, improving the quality of life, and reducing the cost of care. The authors present specific protocols for the diagnosis and prevention and for the management of oral complications in pediatric HCT.

Background It is generally believed that the elimination of certain foods from the diet of mothers during the lactation period produces a significant improvement in breast-fed children who develop allergic symptoms. Several studies have shown the presence of food proteins in human milk; on the other hand, no study has been able to correlate unequivocally the presence of these allergens in human milk with newborn sensitization. Objective The aim of this study was to evaluate the presence of bovine proteins in breast milk. Methods Milk samples were separated by sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS-PAGE). To detect bovine proteins in human milk, immunoblotting was performed by using monoclonal antibodies (MA) specific for β-lactoglobulin and bovine caseins. Results The results of this study do not confirm the presence of bovine proteins in breast milk suggested by other authors and shows unequivocally that the conflicting results reported in the literature about the presence of β-lactoglobulin in human milk are due to cross-reactivity between bovine milk proteins and human proteins. Conclusions Components other than bovine β-lactoglobulin or caseins could be involved in the induction of allergic symptoms in exclusively breast-fed children.

Takayasu arteritis (TA) is a chronic vasculitis of unknown etiology. Experience with anti-tumor necrosis factor alpha (anti-TNF) agents in difficult-to-treat patients with TA is limited and refers to adult patients. Here, we present two cases of pediatric TA treated with infliximab, in which clinical remission was observed. Anti-TNF treatment represents a useful therapy in pediatric Takayasu arteritis too, especially to avoid the risk of long-term corticosteroids toxicity.

Mutations in the human RAG genes that impair, but do not abolish, recombination activity lead to Omenn syndrome, a severe primary immune deficiency that is associated with clinical and pathological features of graft-versus-host disease and oligoclonal expansion of activated, autologous T cells. We have analyzed the mechanisms accounting for peripheral oligoclonality of the T-cell repertoire. Predominance of few T-cell receptor clonotypes (both within TCRAB- and within TCRGD-expressing lymphocytes) is already detectable in the thymus and is further selected for in the periphery, with a different distribution of clonotypes in different tissues. These data indicate that oligoclonality of the T-cell repertoire in Omenn syndrome is due both to intrathymic restriction and to peripheral expansion. Moreover, the RAG genes defect that causes Omenn syndrome directly affects early stages of V(D)J recombination, but does not alter the process of double-strand-break DNA repair, including N and P nucleotide insertion.

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.

We have studied the regeneration of T cell subsets and function after BMT in 21 children affected by combined immunodeficiency after BMT. In the first months, the striking predominance of CD4+ cells displayed the primed CD45R0+ phenotype and a high number of activated (HLA-DR+) T cells were observed. Regeneration of naive CD4+CD45RA+ cells correlated with the recovery of proliferative responses to mitogens (r = 0.64, P

Invasive bacterial infections cause severe diseases, especially in children younger than 5 years of age. Although several risk factors are well known, most sporadic cases remain apparently unexplained. We set up a pilot study to explore new determinants of invasive bacterial diseases (IBDs). We recruited a sample of paediatric patients with IBDs admitted to our hospital between 2004 and 2007. Their families were asked to reply to a questionnaire on epidemiological data and vaccination history. A 5 ml blood sample was obtained for blood cell count, evaluation of total haemolytic complement (CH50) and a functional evaluation of IgM memory B cells and toll-like receptors. Patients also underwent ultrasound examination to determine the presence or absence of the spleen. To date, we have recruited 48 patients with an age range of 1 month to 14 years. Fifty six per cent were male, 62% had meningitis and 69% sepsis. Most invasive infections were associated with Neisseria meningitidis (33%). Epidemiological and clinical data analyses are still in progress. Immunological, epidemiological or bacterial factors may explain cases of IBD in apparently immunocompetent patients. A larger study will be necessary to identify the highest risk profile candidates for specific prevention strategies.

Mutations of the Janus family kinase JAK3 have been found to be responsible for autosomal recessive severe combined immunodeficiency (SCID) in humans. We report here the analysis of four new unrelated patients affected by JAK3-deficient SCID. The genetic defects were heterogeneous and included a large intragenic deletion as well as different point mutations, leading to missense substitutions, early stop codons, or splicing defects. We performed a series of studies of the biochemical events induced by cytokines on lymphoblastoid B-cell lines obtained from these patients. Abnormalities in tyrosine phosphorylation of JAK3 in response to interleukin-2 (IL-2) and IL-4 were present in all patients. Accordingly, IL-2–mediated phosphorylation of STAT5 was also absent or barely detectable. On the contrary, in all cases, we could show reduced but clear phosphorylation of STAT6 upon IL-4 stimulation. In one patient carrying a single amino acid change (Glu481Gly) in the JH3 domain of JAK3, we observed partially conserved IL-2 responses resulting in reduced but detectable levels of JAK3 and STAT5 phosphorylation. Interestingly, the patient bearing this mutation developed a substantial number of circulating CD4+/CD45RO+ activated T lymphocytes that were functionally impaired. In two cases, patients' cells expressed JAK3 proteins with mutations in the JH2 pseudo-kinase domain. A single cysteine to arginine substitution (Cys759Arg) in this region resulted in high basal levels of constitutive JAK3 tyrosine phosphorylation unresponsive to either downregulation by serum starvation or cytokine-mediated upregulation. The characterization of the genetic defects and biochemical abnormalities in these JAK3-deficient patients will help define the role of JAK3 in the ontogeny of a competent immune system and may lead to a better understanding of the JAK3 functional domains.

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.

The Wiskott-Aldrich syndrome (WAS), X-linked severe combined immunodeficiency (SCIDX1), and X-linked agammaglobulinemia (XLA) are severe congenital immunodeficiencies with X-linked inheritance. Although rare, they are all associated with severe infections from early in life, and high morbidity and mortality. Female carriers of these diseases can be identified by a non-random pattern of X-chromosomal inactivation in cell lineages targeted by each gene defect. For patients with WAS, SCIDX1 or XLA, the demonstration of non random X-Chromosome inactivation in their mothers can be used to confirm clinical diagnosis. Furthermore, analysis of X-Chromosome inactivation in at risk females allows preconceptional carrier detection, thus representing an important aid in genetic counseling. For each disease we established a PCR-based, non radioactive assay at the human androgen receptor (HUMARA) locus, that allows analysis of X-Chromosome inactivation in the affected cell types and in tissue specific controls to exclude the issue of skewed X-chromosomal inactivation. In our study, 50 females with a known family history of XLA [19], WAS [18], and SCIDX1 [13],were examined. A carrier status was established in 19 females (7 XLA, 6 WAS, 6 SCIDX1) and excluded in 29 ( 11 XLA, 11 WAS, 7 SCIDX1). Only in 2 cases (4%) the assay was not informative.

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.

The study was performed to evaluate the effects of influenza and pneumococcal vaccines administered alone or in combination. 124 elderly subjects living in community were vaccinated either with influenza split vaccine or with pneumococcal 23-valent or with both vaccines at the same time in different sites. Sera were tested for hemoagglutination inhibiting antibodies for influenza and for antibodies against 23-valent vaccine for streptococcus pneumoniae. No side effects were observed in the vaccinated population. Serological results indicated that influenza vaccine increased significantly antibody levels. No difference was observed between the group which received influenza vaccine alone and that which received influenza and pneumococcal vaccines associated, considering either G.M.T or the percentages of protected individuals or the percentages of subjects who seroconverted. When pneumococcal vaccine was administered at the same time with influenza vaccine, there was a not statistically significant reduction in both mean antibody concentration and mean fold increase. It is concluded that the simultaneous administration of influenza and pneumococcal vaccines to elderly individuals, including subjects at risk, is safe, effective and economically advantageous.

Patients with agammaglobulinemia may excrete enteroviruses, including vaccine-derived poliovirus, for prolonged periods of time. This poses a risk to the patients but it also may pose a risk to the population after eradication of poliovirus and the cessation of routine vaccination. To assess this risk, a pilot study was performed to identify potential poliovirus long-term excretors in a cohort of 38 patients with a definite/presumptive diagnosis of X-linked agammaglobulinemia (XLA). Stool samples were analyzed to detect any polio or other enteroviruses replicating in the gut and neutralizing antibodies against polioviruses were measured in the sera. No viruses were isolated from the stool samples and most sera had neutralizing antibody levels against all three poliovirus serotypes considered by the WHO to be protective in immunocompetent individuals. This suggests that long-term excretion of enteroviruses in patients with agammaglobulinemia is relatively uncommon.

Summary Background X-linked severe combined immunodeficiency (SCIDX1) is an inherited immune defect which leads to death in infancy from severe infections. The defect is caused by mutations of the IL-2RG gene that encodes for the common γ chain shared by several cytokine receptors. The disease is characterised by lack of T and NK cells with normal numbers of B cells. SCIDX1 can be cured by bone marrow transplantation (BMT) or prevented by abortion after prenatal diagnosis. Methods A male fetus was diagnosed as having SCIDX1 by molecular, immunophenotypic, and functional analyses. The fetus was injected intraperitoneally under ultrasound guidance with CD34 haematopoietic progenitor cells purified from paternal bone marrow and T-cell depleted by E resetting. Chimerism analysis was by HLA-DQα typing and γ-chain staining on cord blood. Findings A healthy 3·6 kg boy was delivered by caesarean section at 38 weeks of gestation with no clinical or laboratory signs of graft-versus-host disease. Engraftment of donor-derived CD2 cells was found at birth. At 3·5 months of age the infant is well and his T-cell counts and function are normal. Interpretation In-utero transplantation of haematopoietic progenitor cells allowed immune reconstitution of a fetus with SCIDXI and may be an alternative to elective abortion. Our report should encourage applications of this method to other inherited disorders curable by BMT.

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.

Summary Many clinical and experimental data strongly support the role of immune mechanisms in the pathogenesis of childhood epilepsy. Following Pechadre’s first observations with intramuscular immune globulin (IMIG), intravenous immune globulin (IVIG) has been employed in some forms of intractable childhood epilepsy (ICE), mainly ill West syndrome (WS) and Lennox Gastaut syndrome (LGS), with good results. So far, 373 children suffering from ICE have been treated in 29 studies and 174 have responded favourably. Although these studies are heterogeneous and controls are lacking, most authors report similar responsiveness ranging from 30% to 50%. Several mechanisms have been suggested to account for the efficacy of IVIG in ICE including antiviral effect, substitutive therapy in patients with concomitant humoral immunodeficiency, idiotype-anti-idiotype interaction or a neuromodulant effect. To better define the real efficacy of IVIG in ICE in paediatric patients, a randomized, multicenter, double-blind clinical trial was started in 1993. including only patients suffering from WS and LGS. To date, only one doubleblind trial had been carried out (with both adult and paediatric patients); it showed a clear trend in favour of IVIG treatment but lacked statistical significance, perhaps because of the small and heterogenous sample. Controlled multicentre studies on well-defined populations arc needed and patients with WS and LGS are probably the best candidates.

The CD40 ligand (CD40L) is a molecule expressed by activated T cells which plays a critical role in the regulation of B-cell responses, including differentiation into Ig-producing cells. Using the specific monoclonal antibody TRAP1 we have evaluated the ontogeny of CD40L expression in 97 normal individuals between birth and 50 years of age. The expression of CD40L is a function of age; it is severely reduced at birth, progressively increases during the first months of life, and reaches a plateau in the second decade. This progressive attainment of the ability to express CD40L is due to a process of maturation of the CD4 + subset, being significantly correlated with the expression of the CD45RO antigen.

Severe combined immune deficiency (SCID) represents a heterogenous group of hereditary diseases. Mutations in the common gamma-chain (gamma c), which is part of several cytokine receptors including those for interleukin (IL)-2, IL-4, IL-7, IL-9 and IL-15, are responsible for X-linked SCID, which is usually associated with a lack of circulating T cells and the presence of B lymphocytes (T- B+ SCID). The gene(s) responsible for autosomal recessive T- B+ SCID is still unknown. The Jak-3 protein kinase has been found to associate with the gamma c-chain-containing cytokine receptors. Therefore Jak-3 or other STAT proteins with which it interacts are candidate genes for autosomal recessive T- B+ SCID. Here we investigate two unrelated T- B+ SCID patients (both from consanguineous parents) who have homozygous mutations in the gene for Jak-3. One patient carries a mutation (Tyr100-->Cys) in a conserved tyrosine residue in the JH7 domain of Jak-3 which is absent in more than 150 investigated chromosomes. The other patient carries a homozygous 151-base-pair deletion in the kinase-like domain, leading to a frameshift and premature termination. Both mutations resulted in markedly reduced levels of Jak-3. These findings show that abnormalities in the Jak/STAT signalling pathway can account for SCID in humans.

The fetus can be compared to a haploidentical graft, sharing only half of the histocompatibility antigens of the “mother host”. As such, the fetus should be rejected by the immune system of the mother. At the same time, since the fetal immune system starts developing around the 9th week of gestation, the fetus should also mount a “graft-versus-host” reaction against those histocompatibility antigens that it has not inherited from the “mother host”. It follows that survival of the fetus - indeed of all mammalian species - is strictly dependent on a wide variety of mechanisms preventing graft rejection by the mother and graft-versus-host reaction by the fetus [1]. Among the latter, a major role is certainly played by the slow intrauterine maturation of the fetal immune function, ultimately resulting in the physiological immune deficiency of the neonate that involves both innate and adaptive immunity. This immune immaturity - more severe in preterm and in very low birth weight infants - results in a weak immune response to antigen challenge and in the production of a poor immunological memory [2].

to analyze the interrelationships between GH secretion and pattern of sleep.18 children (10 male, 8 female; mean age 9.1 yr, range 5.1-14.3 yr), with short stature (mean height standard deviation score (SDS) -2.52, range -3.86-(-)1.88; mean height velocity SDS -1.1, range -2.40-(-)0.08), including 9 children with genetic short stature and 9 with idiopathic short stature.blood samples were taken every 15 min from 2000 h-0800 h, and GH profiles were analyzed by the PULSAR computerized peak identification algorithm; simultaneous sleep was analyzed by electroencephalogram recording.no significant correlation was noted between GH secretion parameters and any of the electroencephalogram parameters evaluated: stage 1 (S1) percent, stage 2 (S2) percent, slow-wave sleep (SWS) percent, rapid eye movement sleep percent, wakefulness (W) percent, and sleep efficiency (EFF); there was no significant difference in GH secretion between children with EFF less than 76% and those with EFF more than 76% (P0.5). Maximal GH peak coincided 9 times (50%) with SWS, 3 times (17%) with S2, 3 times with W, twice (11%) with S1, and once (6%) with rapid eye movement sleep. First GH peak coincided 12 times (67%) with W, 3 times with S2, twice with SWS, and once with S1. There was no significant difference comparing the percentage of sleep stages occurring in the 15 min of maximal GH increment, in the 15 min preceding it, and in those following it; there was no significant difference comparing the percentage of sleep stages occurring in the 15 min preceding the onset of a GH peak and in those following it.GH secretion in short children seems independent of the sleep stage and efficiency; in children it is possible that GH secretion relates with sleep per se and with neurohormonal changes occurring at nighttime rather than with a specific sleep stage or sleep stage sequence.

A single oral dose (430 mg/kg) of atrazine, a widely employed s-triazine herbicide, was administered to young male rats. There was a significant increase of the in vivo elimination ofhexobarbital and a significant induction of the activity of 7-pentoxyresorufin-O-dealkylase, while cytochrome P-450 content and other mixed function oxidase activities remained unaltered. The administration of carbon tetrachloride (CCI,) to atrazine pretreated rats did not substantially augment the impairment of drug metabolizing enzymes brought about by CCLi alone. Results suggest that atrazine behaves like a relatively weak inducer of phenobarbital-inducible families of cytochrome P-450.