Attenuation of diet-induced obesity and induction of white fat browning with a chemical inhibitor of histone deacetylases.

BACKGROUND/OBJECTIVES: In the last decade, a strict link between epigenetics and metabolism has been demonstrated. Histone deacetylases (HDACs) have emerged as key epigenetic regulators involved in metabolic homeostasis in normal and pathologic conditions. Here we investigated the effect of the class I HDAC inhibitor MS-275 in a model of obesity induced by a high-fat diet (HFD). METHODS: C57BL6/J male mice were fed HFD for 17 weeks and then randomized in two groups, treated intraperitoneally with vehicle dimethylsulfoxide (DMSO) or with the class I selective HDAC inhibitor MS-275 every other day for 22 days. Glucose tolerance test and measurement of body temperature during cold exposure were performed. Adipose tissues and liver were phenotypically characterized through histological analysis. Gene and protein expression analysis of brown and white adipose tissues (WATs) were performed. RESULTS: MS-275 treated mice showed 10% reduction of body weight, lower adipocyte size and improved glucose tolerance. Inhibition of class I HDAC determined reduction of adipocyte size and of fat mass, paralleled by higher expression of adipose functionality markers and by increased rate of lipolysis and fatty acid ß-oxidation. MS-275 also promoted thermogenic capacity, related to 'browning' of visceral and subcutaneous WAT, showing increased expression of uncoupling protein 1. In brown adipose tissue, we observed limited effects on gene expression and only reduction of brown adipocyte size. CONCLUSIONS: This study provides evidence that class I HDAC inhibition stimulated functionality and oxidative potential of adipose tissue, improving glucose tolerance and ameliorating the metabolic profile in diet-induced obese mice.

Shrimp Allergy: Analysis of Commercially Available Extracts for In Vivo Diagnosis.

BACKGROUND AND OBJECTIVE: Skin prick testing (SPT) with commercial extracts is the first step in the diagnosis of shrimp allergy, although its clinical efficiency is unknown. Objective: To analyze the clinical usefulness of all commercial crustacean extracts available for SPT in Italy. METHODS: We performed a multicenter study of 157 shrimp-allergic patients who underwent SPT with 5 commercial crustacean extracts and with house dust mite (HDM) extract. Commercial extracts were analyzed using SDS-PAGE and compared with a freshly prepared in-house shrimp extract. IgE to Pen a 1/Pen m 1, Pen m 2, and Pen m 4 was determined, and immunoblot analysis was performed on a large number of sera. RESULTS: The skin reactions caused by commercial crustacean extracts were extremely heterogeneous, resulting in 32 clinical profiles, with marked differences in protein content and missing proteins at molecular weights corresponding to those of major shrimp allergens. Only strong Pen a 1/Pen m 1 reactors reacted to both HDM and all 5 commercial extracts in SPT. Most patients, including those who were tropomyosin-negative, reacted to HDM. Patients reacted to a large and variable array of proteins, and IgE reactivity was common at high molecular weights (>50 kDa). CONCLUSIONS: The in vivo diagnosis of shrimp allergy must continue to be based on SPT with fresh material. Shrimp-allergic patients frequently react to a number of ill-defined high-molecular-weight allergens, thus leaving currently available materials for component-resolved diagnosis largely insufficient. Mites and crustaceans probably share several allergens other than tropomyosin.

Theory and preliminary experimental verification of quantitative edge illumination x-ray phase contrast tomography.

X-ray phase contrast imaging (XPCi) methods are sensitive to phase in addition to attenuation effects and, therefore, can achieve improved image contrast for weakly attenuating materials, such as often encountered in biomedical applications. Several XPCi methods exist, most of which have already been implemented in computed tomographic (CT) modality, thus allowing volumetric imaging. The Edge Illumination (EI) XPCi method had, until now, not been implemented as a CT modality. This article provides indications that quantitative 3D maps of an object's phase and attenuation can be reconstructed from EI XPCi measurements. Moreover, a theory for the reconstruction of combined phase and attenuation maps is presented. Both reconstruction strategies find applications in tissue characterisation and the identification of faint, weakly attenuating details. Experimental results for wires of known materials and for a biological object validate the theory and confirm the superiority of the phase over conventional, attenuation-based image contrast.

Phase stability of Li-Mn-O oxides as cathode materials for Li-ion batteries: insights from ab initio calculations.

In this work, we present a density-functional theory (DFT) investigation of the phase stability, electrochemical stability and phase transformation mechanisms of the layered and over-lithiated Mn oxides. This study includes the thermodynamic stability of Li and oxygen vacancies, to examine the electrochemical activation mechanisms of these cathode materials. The DFT calculations provide phase diagrams of the Li-Mn-O system in both physical and chemical potential spaces, including the crystals containing vacancies as independent phases. The results show the ranges of electrochemical activity for both layered LiMnO2 and over-lithiated Li2MnO3. By using a thermodynamic model analysis, we found that the required temperature for oxygen evolution and Li vacancy formation is too high to be compatible with any practical synthesis temperature. Using solid-state transition calculations, we have identified the key steps in the phase transition mechanism of the layered LiMnO2 into the spinel phase. The calculated effects of pH on the Li-Mn-O phase stability elucidated the mechanism of Mn(2+) formation from the spinel phase under acidic conditions.

Comparison of single distance phase retrieval algorithms by considering different object composition and the effect of statistical and structural noise.

Phase retrieval is a technique for extracting quantitative phase information from X-ray propagation-based phase-contrast tomography (PPCT). In this paper, the performance of different single distance phase retrieval algorithms will be investigated. The algorithms are herein called phase-attenuation duality Born Algorithm (PAD-BA), phase-attenuation duality Rytov Algorithm (PAD-RA), phase-attenuation duality Modified Bronnikov Algorithm (PAD-MBA), phase-attenuation duality Paganin algorithm (PAD-PA) and phase-attenuation duality Wu Algorithm (PAD-WA), respectively. They are all based on phase-attenuation duality property and on weak absorption of the sample and they employ only a single distance PPCT data. In this paper, they are investigated via simulated noise-free PPCT data considering the fulfillment of PAD property and weakly absorbing conditions, and with experimental PPCT data of a mixture sample containing absorbing and weakly absorbing materials, and of a polymer sample considering different degrees of statistical and structural noise. The simulation shows all algorithms can quantitatively reconstruct the 3D refractive index of a quasi-homogeneous weakly absorbing object from noise-free PPCT data. When the weakly absorbing condition is violated, the PAD-RA and PAD-PA/WA obtain better result than PAD-BA and PAD-MBA that are shown in both simulation and mixture sample results. When considering the statistical noise, the contrast-to-noise ratio values decreases as the photon number is reduced. The structural noise study shows that the result is progressively corrupted by ring-like artifacts with the increase of structural noise (i.e. phantom thickness). The PAD-RA and PAD-PA/WA gain better density resolution than the PAD-BA and PAD-MBA in both statistical and structural noise study.

Heterogenity of IgE response to walnut and hazelnut in Italian allergic patients.

BACKGROUND: The prevalence of IgE reactivity against genuine walnut and hazelnut allergens is poorly defined. OBJECTIVE: The IgE response to walnut and hazelnut was investigated in Italian patients with primary allergy to these nuts. METHODS: Sera from 36 patients allergic to hazelnut and/or walnut, not reactive to PR-10, profilin, and LTP, underwent immunoblot analysis with extracts of both nuts. RESULTS: Most patients had a history of systemic symptoms following the ingestion of the offending food(s). Twelve patients were sensitized to both walnut and hazelnut, and 13 were sensitized to other nuts and seeds (cashew, peanut, sesame, pine nut, almond, Brazil nut, and pistachio). On walnut immunoblot, the 7 sera which scored positive showed much variability in their IgE profile. Two reacted uniquely at 10 kDa, and the others at 35 , 40, 45, 50, 67, and > 67 kDa. The profiles obtained under reducing and non-reducing conditions showed several differences. The 7 sera positive on hazelnut immunoblot under reducing conditions recognized sera at 10 kDa and at <10 kDa (n=1), 20 kDa (n=4), at about 22, 24, 30, 40, 43, 58, 60, and 90 kDa, and higher m.w. in other cases. Under non-reducing conditions IgE reactivity at 20, 28, 35, 40, 45, 60, 90, and 100 kDa, was detected. Only two sera scored positive under both conditions and showed an IgE profile that partly changed from one assay to another. CONCLUSION: The current list of walnut and hazelnut allergens is far from being complete. Both reducing and non-reducing conditions are needed to detect IgE reactivity in individual patients.

First application of computed radiology to mammography with synchrotron radiation.

OBJECTIVE: The aim of this study was to evaluate the feasibility of phase-contrast mammography with synchrotron radiation using a high-resolution computed radiology (CR) system devoted to mammography. MATERIALS AND METHODS: The study was performed at the Synchrotron Radiation for Medical Physics (SYRMEP) beamline of the Elettra synchrotron radiation (SR) facility in Trieste (Italy); X-ray beams were in the range 16-22 keV with a high degree of monochromaticity and spatial coherence. The CR system evaluated is the FCR Profect CS by Fujifilm Global. The first images were obtained from test objects and surgical breast specimens. Images obtained using SR and both screen-film and the CR system were compared with images of the same samples acquired with digital mammography equipment. In view of the good quality of the results obtained, the CR system was used in two mammographic examinations with SR. RESULTS: Images acquired using SR and both screen-film and CR were obtained with the same level of delivered dose. Image quality obtained with CR was similar or superior to that of screen-film images. Moreover, the digital images obtained with SR were always better than those acquired using the digital mammography system. CONCLUSIONS: Phase-contrast mammography with SR using the studied CR system is a feasible option.

Multiscale X-ray phase-contrast tomography: From breast CT to micro-CT for virtual histology.

Phase-contrast imaging techniques address the issue of poor soft-tissue contrast encountered in traditional X-ray imaging. This can be accomplished with the propagation-based phase-contrast technique by employing a coherent photon beam, which is available at synchrotron facilities, as well as long sample-to-detector distances. This study demonstrates the optimization of propagation-based phase-contrast computed tomography (CT) techniques for multiscale X-ray imaging of the breast at the Elettra synchrotron facility (Trieste, Italy). Two whole breast mastectomy samples were acquired with propagation-based breast-CT using a monochromatic synchrotron beam at a pixel size of 60 µm. Paraffin-embedded blocks sampled from the same tissues were scanned with propagation-based micro-CT imaging using a polychromatic synchrotron beam at a pixel size of 4 µm. Images of both methodologies and of the same sample were spatially registered. The resulting images showed the transition from whole breast imaging with propagation-based breast-CT methodology to virtual histology with propagation-based micro-CT imaging of the same sample. Additionally, conventional histological images were matched to virtual histology images. Phase-contrast images offer a high resolution with low noise, which allows for a highly precise match between virtual and conventional histology. Furthermore, those techniques allow a clear discernment of breast structures, lesions, and microcalcifications, being a promising clinically-compatible tool for breast imaging in a multiscale approach, to either assist in the detection of cancer in full volume breast samples or to complement structure identification in paraffin-embedded breast tissue samples.

Shrimp allergy in Italian adults: a multicenter study showing a high prevalence of sensitivity to novel high molecular weight allergens.

BACKGROUND: Shrimp is a frequent cause of food allergy worldwide. Besides tropomyosin, several allergens have been described recently. OBJECTIVE: We investigated which allergens are involved in Italian shrimp-allergic adults. METHODS: Sera from 116 shrimp-allergic patients selected in 14 Italian allergy centers were studied. Skin prick tests with house dust mite (HDM) as well as measurements of IgE to Pen a 1 (shrimp tropomyosin) and whole shrimp extract were performed. All sera underwent shrimp immunoblot analysis, and inhibition experiments using HDM extract as inhibitor were carried out on some Pen a 1-negative sera. RESULTS: Immunoblots showed much variability. IgE reactivity at about 30 kDa (tropomyosin) was found in <50% of cases, and reactivity at about 67 kDa and >90 kDa was frequent. Further reactivities at 14-18, 25, 43-50, about 60 and about 80 kDa were detected. Most subjects had a history of shrimp-induced systemic symptoms irrespective of the relevant allergen protein. IgE to Pen a 1 were detected in sera from 46 (41%) patients. Skin reactivity to HDM was found in 43/61 (70%) Pen 1-negative subjects and inhibition studies showed that pre-adsorption of sera with HDM extract induced a marked weakening of the signal at >67 kDa. CONCLUSIONS: Several allergens other than tropomyosin are involved in shrimp allergy in adult Italian patients. Some hitherto not described high molecular weight allergens seem particularly relevant in this population and their cross-reactivity with HDM allergens makes them novel potential panallergens of invertebrates.

Fiber density index in the evaluation of the spinal cord in patients with multiple sclerosis.

PURPOSE: The aims of this study were to determine fractional anisotropy (FA) and the fibre density index (FDi) in the cervical spinal cord of patients with multiple sclerosis (MS) by using diffusion-tensor magnetic resonance imaging (DT-MRI) to identify possible differences between MS patients and controls. MATERIALS AND METHODS: We studied 27 patients with MS - nine with primary progressive (PPMS), nine with secondary progressive (SPMS) and nine with relapsing-remitting (RRMS) disease - and 18 healthy individuals as controls. Conventional and DTI sequences with diffusion gradients applied in 32 directions were obtained. The results were compared between healthy controls and patients, between healthy controls and individual forms of MS and between the three forms of MS. Statistical analysis was performed by analysis of variance (ANOVA) and Student's t test. RESULTS: The FDi in the three subgroups of patients and in controls showed a statistically significant difference. Using the t test, we found results from both PPMS and SPMS groups were different from controls. The correlation between FA and FDi was significant both in healthy controls and in MS patients evaluated as a single group. CONSLUCIONS: Despite the small group of patients, these findings suggest that FDi associated with FA is a sensitive parameter for assessing spinal cord damage in patients with MS.

Functional MRI during the execution of a motor task in patients with multiple sclerosis and fatigue.

PURPOSE: This study was undertaken to assess cortical activation during execution of a motor task in patients with multiple sclerosis (MS) and fatigue. MATERIALS AND METHODS: We enrolled 24 right-handed patients affected by relapsing-remitting MS and mild disability (12 with and 12 without fatigue) and 15 healthy volunteers. Magnetic resonance imaging (MRI) examination (1.5 T) was performed with conventional sequences and an echoplanar imaging (EPI) sequence for functional MRI (fMRI). The motor task consisted of sequential finger tapping performed with the right hand. Statistical maps of motor activation were obtained. Comparison between the two subgroups of patients and between patients and controls was performed with analysis of variance (ANOVA) statistical analysis (p<0.05). RESULTS: Compared with controls, patients without fatigue showed greater activation of the primary sensorimotor cortex bilaterally, of the right supplementary motor cortex, of the left premotor cortex, of the left cerebellum and of the superior parietal lobule bilaterally. Compared with patients without fatigue, patients with fatigue demonstrated greater activation of the right premotor area, of the putamen and the dorsolateral prefrontal cortex. CONCLUSIONS: Patients with fatigue have greater activation of the motor-attentional network when performing a simple motor task.

Optimization of a customized simultaneous algebraic reconstruction technique algorithm for phase-contrast breast computed tomography.

Objective.To introduce the optimization of a customized GPU-based simultaneous algebraic reconstruction technique (cSART) in the field of phase-contrast breast computed tomography (bCT). The presented algorithm features a 3D bilateral regularization filter that can be tuned to yield optimal performance for clinical image visualization and tissues segmentation.Approach.Acquisitions of a dedicated test object and a breast specimen were performed at Elettra, the Italian synchrotron radiation (SR) facility (Trieste, Italy) using a large area CdTe single-photon counting detector. Tomographic images were obtained at 5 mGy of mean glandular dose, with a 32 keV monochromatic x-ray beam in the free-space propagation mode. Three independent algorithms parameters were optimized by using contrast-to-noise ratio (CNR), spatial resolution, and noise texture metrics. The results obtained with the cSART algorithm were compared with conventional SART and filtered back projection (FBP) reconstructions. Image segmentation was performed both with gray scale-based and supervised machine-learning approaches.Main results.Compared to conventional FBP reconstructions, results indicate that the proposed algorithm can yield images with a higher CNR (by 35% or more), retaining a high spatial resolution while preserving their textural properties. Alternatively, at the cost of an increased image 'patchiness', the cSART can be tuned to achieve a high-quality tissue segmentation, suggesting the possibility of performing an accurate glandularity estimation potentially of use in the realization of realistic 3D breast models starting from low radiation dose images.Significance.The study indicates that dedicated iterative reconstruction techniques could provide significant advantages in phase-contrast bCT imaging. The proposed algorithm offers great flexibility in terms of image reconstruction optimization, either toward diagnostic evaluation or image segmentation.

Phase retrieval in quantitative x-ray microtomography with a single sample-to-detector distance.

Phase retrieval extracts quantitative phase information from x-ray propagation-based phase-contrast images. Notwithstanding inherent approximations, phase retrieval using a single sample-to-detector distance (SDD) is very attractive, because it imposes no setup complications or additional radiation dose compared to absorption-based imaging. Considering the phase-attenuation duality (ε=δ/ß, where ε is constant), a simple absorption correction factor is proposed for the modified Bronnikov algorithm in x-ray propagation-based phase-contrast computed tomography (PPCT). Moreover, a practical method for calculating the optimal ε value is proposed, which requires no prior knowledge of the sample. Tests performed on simulation and experimental data successfully distinguished different materials in a quasihomogeneous and weakly absorbing sample from a single SDD-PPCT data point.

Are IgE levels to foods other than rosaceae predictive of allergy in lipid transfer protein-hypersensitive patients?

BACKGROUND: Lipid transfer protein (LTP), the most frequent cause of primary food allergy in Italy, is a cross-reacting plant pan-allergen. Markers able to predict whether a patient sensitized to a certain food but not yet clinically allergic will develop allergy would be extremely helpful. OBJECTIVE: It was the aim of this study to investigate the relevance of IgE levels to some plant foods other than Rosaceae as predictors of either local or systemic allergic reaction in LTP-allergic subjects. METHODS: One hundred (40 males, 60 females , mean age 29 years) peach-allergic patients monosensitized to LTP seen at 14 Italian centres in 2009 were studied. Walnut, hazelnut, peanut, tomato, rice and/or maize allergy was ascertained by interview and confirmed by positive skin prick test. IgE levels to these foods and to rPru p 3 were measured. RESULTS: Higher levels of IgE to Pru p 3 were associated with a higher prevalence of allergy to hazelnut, peanut and walnut. For all study foods, except rice, median IgE levels in allergic subjects significantly exceeded those in tolerant subjects, though within single allergic groups, the differences between patients reporting systemic or local (oral) symptoms were not significant. Ninety-five percent cut-off IgE levels predictive of clinical allergy were established for study foods although the marked overlaps between allergic and tolerant subjects made them of limited usefulness. CONCLUSION: Specific IgE levels are only partially predictive of clinical allergy. The reasons why some individuals showing low specific IgE levels develop clinical allergy whereas others showing high IgE levels do not, despite similar exposure to the allergen, remain unclear.

Prediction of phonon thermal transport in thin GaAs, InAs and InP nanowires by molecular dynamics simulations: influence of the interatomic potential.

A number of different potentials are currently being used in molecular dynamics simulations of semiconductor nanostructures. Confusion can arise if an inappropriate potential is used. To illustrate this point, we performed direct molecular dynamics simulations to predict the room temperature lattice thermal conductivity λ of thin GaAs, InAs and InP nanowires. In each case, simulations performed using the classical Harrison potential afforded values of λ about an order of magnitude smaller than those obtained using more elaborate potentials (an Abell-Tersoff, as parameterized by Hammerschmidt et al for GaAs and InAs, and a potential of Vashishta type for InP). These results will be a warning to those wishing to use computer simulations to orient the development of quasi-one-dimensional systems as heat sinks or thermoelectric devices.

Magnetic cooperative effects in small Ni-Ru clusters.

We report ab initio calculations of the structures, magnetic moments, and electronic properties of Ni(7)-xRu(x) clusters (x = 0-7) using a density-functional method that employs linear combinations of pseudoatomic orbitals as basis sets, nonlocal norm-conserving pseudopotentials, and the generalized gradient approximation to exchange and correlation. The pure clusters Ni(7) and Ru(7) were predicted to have octahedral and cubic structures, respectively, and the binary clusters were found to be either decahedral (Ni(6)Ru, Ni(5)Ru(2), and Ni(4)Ru(3)) or cubic (Ni(3)Ru(4), Ni(2)Ru(5), and NiRu(6)). For Ni(5)Ru(2) and Ni(4)Ru(3) we found a magnetic cooperative phenomenon, which is due to both geometrical effects and electronic contributions through Ni-Ru hybridization.

Magnetism of substitutional Fe impurities in graphene nanoribbons.

Using the generalized gradient approximation to exchange and correlation, we perform density functional calculations on an Fe atom at a single vacancy of graphene nanoribbons. Our results show that, after relaxation, the Fe atom is magnetic, in contrast to the behavior recently found for Fe at a single vacancy of the graphene sheet.

Assessing cardiovascular risk in Mediterranean women with polycystic ovary syndrome.

BACKGROUND: Although dyslipidemia is common in the polycystic ovary syndrome (PCOS), current diagnostic guidelines suggest to evaluate only plasma HDL-cholesterol and triglyceride concentrations, assuming that, in this disorder, cardiovascular risk is mainly due to the presence of the metabolic syndrome (MS). In the US, MS has been found in up to 50% of PCOS, but in other countries its prevalence is lower. METHODS: We compared the prevalence of MS with that of increased LDL- and non-HDL-cholesterol levels in 350 Mediterranean PCOS women (244 anovulatory and 106 ovulatory), and 95 normo-weight and 90 body mass index (BMI)- matched controls. RESULTS: The prevalence of MS was 7.1% in PCOS, higher than normoweight and BMI-matched controls (2.4% and 3.5%, respectively, p < 0.05 for both). The prevalence of elevated LDL- and non-HDL-cholesterol levels in PCOS was respectively, 14.9% and 8.6%, higher than normoweight (2.1% and 1.0%, respectively, p < 0.01 for both) and BMI-matched controls (4.4% and 2.2%, respectively, p<0.05 for both). In PCOS, increased LDL-cholesterol was twice more prevalent than MS or non-HDL-cholesterol. Only a minority of PCOS with MS had increased LDL-cholesterol while increased non-HDL-cholesterol was generally associated to increased LDL-cholesterol. CONCLUSIONS: We have found that in Mediterranean PCOS the prevalence of MS is relatively low while elevated LDL-cholesterol levels are more prevalent. Therefore, beyond MS, a more comprehensive lipid evaluation, including LDL-cholesterol, is needed for a more effective assessment of cardiovascular risk in PCOS.

Epinephrine autoinjector prescription in food-allergic adults: symptom-based only or allergen-based also? An Italian multi-centre study.

BACKGROUND: Epinephrine is the treatment of choice for acute food-allergic reactions but existing guidelines state that it should be prescribed uniquely to patients who already experienced at least one food-induced anaphylactic episode. OBJECTIVE: We investigated whether in Italy epinephrine auto-injector is prescribed uniquely following the existing guidelines only, or is allergen-informed as well (i.e., based on the potential risk associated with sensitization to certain food allergens), and hence preventive. METHODS: 1110 adult patients (mean age 31 years; M/F 391/719) with food allergy seen at 19 allergy outpatient clinics were studied. Patients with a history of probable anaphylaxis were identified. Subjects were classified as having primary (type 1) and/or secondary (type 2) food allergy and were divided into several subgroups based on the offending allergen/food. Epinephrine prescriptions were recorded and analyzed both as a whole and by sensitizing allergen. RESULTS: Epinephrine was prescribed to 138/1100 (13%) patients with a significant difference between subjects with type-1 and type-2 food allergy (132/522 [25%] vs. 6/629 [1%]; p < 0.001). The epinephrine group included most patients with a history of anaphylaxis (55/62 [89%]) or emergency department visits 106/138 (77%). In some specific subsets, namely fish-, tree nuts-, and lipid trasfer protein (LTP)-allergic patients, epinephrine was prescribed to patients without a history of systemic allergic reactions. CONCLUSIONS: Italian allergy specialists prescribe epinephrine auto-injectors both on the basis of clinical history of severe reactions and on a critical analysis of the hazard associated with the relevant protein allergens, which suggests a good knowledge of allergens as well as acquaintance with the guidelines for prescription of emergency medication.

A prospective Italian survey on the safety of subcutaneous immunotherapy for respiratory allergy.

BACKGROUND: Subcutaneous immunotherapy is effective for the treatment of respiratory allergy, and it is largely used in Italy, but no systematic safety assessment has been carried out so far. OBJECTIVE: To assess prospectively the safety of injection immunotherapy in a multicentre, real-life survey. METHODS: Eleven Italian allergy departments recorded the clinical characteristics of systemic reactions (SRs) due to immunotherapy. Vaccines were prescribed according to guidelines; only standardized depot extracts were used. SRs were graded according to the EAACI recommendations, and were classified as immediate or delayed. RESULTS: One thousand seven hundred and thirty-eight patients (847 males, age range 5-71) received immunotherapy from eight different manufacturers, for a total of 2038 courses (300 patients received two extracts). A total of 60 785 injections were given over a mean immunotherapy duration of 3 years. Overall, 95 reactions were observed in 57 patients (3.28%), corresponding to 4.7% of the courses and 1.56/1000 injections. Twenty-five patients experienced more than one adverse event. There were 34 grade 2, 60 grade 3 and one grade 4 reactions and no fatality. SRs occurred more frequently in patients with asthma than in patients with rhinitis alone (4.1% vs. 1.1%), and were equally distributed between the build-up and the maintenance phase. Ragweed and grass extracts caused significantly more side effects than other allergens. CONCLUSION: In this large prospective study, the rate of SRs was low, thus confirming that injection immunotherapy has an acceptable risk/benefit ratio when prescribed and carried out according to recommendations.