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Background: Few data are available on the pharmacoeconomic aspects of immunotherapy., Objective: To evaluate, from the health care system and societal perspectives, the costs and consequences of sublingual immunotherapy (SLIT) added to pharmacotherapy compared with drugs alone for respiratory allergy., Methods: This study compared costs, clinical outcomes, and cost-effectiveness ratios of 2 strategies in the management of allergic rhinitis and asthma, namely, SLIT associated with pharmacotherapy and pharmacotherapy alone (no SLIT). A decision tree was developed and populated with epidemiologic and resource utilization data concerning approximately 2,200 patients. Direct costs included visits, tests, pharmacotherapy, immunotherapy, and hospitalizations. Indirect costs and out-of-pocket drugs were also included. Outcome was calculated as the number of improved patients and asthma cases avoided at 6 years. Sensitivity analysis was performed by varying costs and epidemiologic data., Results: SLIT improved the symptoms of 399 of 1,000 patients and prevented asthma in 229 of 1,000 patients compared with drugs alone. For SLIT added to pharmacotherapy and pharmacotherapy alone, the direct cost per patient at more than 6 years was Euro2,400 and Euro3,026, whereas the indirect cost was Euro1,913 and Euro3,400., Conclusion: From both perspectives and for both effectiveness end points, SLIT is less expensive and more effective than pharmacotherapy alone.

The efficacy and safety of sublingual immunotherapy (SLIT) are currently supported by clinical trials, meta-analysis and post-marketing surveys. Practice parameters for clinical use of SLIT are proposed here by a panel of Italian specialists, with reference to evidence based criteria. Indications to SLIT include allergic rhinoconjunctivitis, asthma, and isolated conjunctivitis (strength of recommendation: grade A). As to severity of the disease, SLIT is indicated in moderate/severe intermittent rhinitis, persistent rhinitis and mild to moderate asthma (grade D). SLIT may be safely prescribed also in children aged three to five years (grade B), and its use in subjects aged more than 60 years is not prevented when the indications and contraindication are ascertained (grade D). The choice of the allergen to be employed for SLIT should be made in accordance with the combination of clinical history and results of skin prick tests (grade D). Polysensitisation, i.e. the occurrence of multiple positive response does not exclude SLIT, which may be done with the clinically most important allergens (grade D). As to practical administration, co-seasonal, pre co-seasonal, and continuous schedules are available, being the latter recommended for perennial allergens or for pollens with particularly prolonged pollination, such as Parietaria (grade D). For pollens with relatively short pollination, such as grasses and trees (cypress, birch, alder, hazelnut, olive) the pre co-seasonal and perennial schedules are preferred (grade C). The build-up phases suggested by manufacturers can be safely used (grade A), but they can be modified according to the patient's tolerance (grade C). A duration of SLIT of 3-5 years is recommended to ensure a long-lasting clinical effect after the treatment has been terminated (grade C).

Background: Wheat is believed to be an uncommon cause of food allergy in adults; the number of studies that address IgE mediated wheat allergy in adults is all too few., Objective: Determine how many subjects with a history of wheat allergy have real allergy by double-blind, placebo-controlled food challenge; identify the symptoms manifested during the challenge; determine the lowest provocation dose; determine the performance characteristics of wheat skin prick test and specific IgE; identify subjects with real wheat allergy for potential immunoblotting studies., Methods: Patients underwent skin test with commercial wheat extract; specific wheat IgE was determined. Subjects were challenged with 25 g wheat. Subjects who were positive to raw wheat challenge underwent cooked wheat challenge., Results: Thirty-seven double-blind placebo-controlled wheat challenges were performed on 27 patients. A total of 13 of 27 (48%) patients had a positive result. Eleven subjects with positive raw wheat challenge underwent cooked wheat challenge: 10 were positive. The provocation dose range was 0.1 to 25 g. Twenty-seven percent of the subjects allergic to wheat had a provocation dose that was < or =1.6 g., Conclusion: Wheat causes real food allergy in adults. More than a quarter of the patients allergic to wheat reacted to less than 1.6 g wheat. Specific IgE was more sensitive than skin test for wheat; however, specificity and predictive values were low for both tests. Thus, these tests should not be used to validate diagnosis of wheat allergy.

Adverse reactions to foods, aside from those considered toxic, are caused by a particular individual intolerance towards commonly tolerated foods. Intolerance derived from an immunological mechanism is referred to as Food Allergy, the non-immunological form is called Food Intolerance. IgE-mediated food allergy is the most common and dangerous type of adverse food reaction. It is initiated by an impairment of normal Oral Tolerance to food in predisposed individuals (atopic). Food allergy produces respiratory, gastrointestinal, cutaneous and cardiovascular symptoms but often generalized, life-threatening symptoms manifest at a rapid rate-anaphylactic shock. Diagnosis is made using medical history and cutaneous and serological tests but to obtain final confirmation a Double Blind Controlled Food Challenge must be performed. Food intolerances are principally caused by enzymatic defects in the digestive system, as is the case with lactose intolerance, but may also result from pharmacological effects of vasoactive amines present in foods (e.g. Histamine). Prevention and treatment are based on the avoidance of the culprit food.

Background: Economic evaluations are increasingly relevant in order to provide support for decision makers when judging about alternative ways to allocate public funds. Aim of this study was to evaluate the economic effect of treatment with high dose sublingual immunotherapy (SLIT) in children., Methods: The study involved one allergy center, located in the north of Milan, Italy. From the existing records of patients seen for allergic disease, we extracted all children and adolescents with allergic disease, who had 1-year data prior to receiving SLIT and 3-year data on SLIT. Outcome measures (number of exacerbations, visits, absence from nursery or school), direct costs (euros spent on drugs, specialist visits, SLIT) and indirect costs (costs resulting from children school and parental work loss) were analysed. A second analysis compared a sub-group of allergic asthmatic children with a control group for costs, based on records of patients not SLIT-treated, extracted from a network-database of paediatricians., Results: 135 patients were extracted, of which 46 had perennial, and 89 had seasonal allergy with comparable gender and age distribution. A substantial reduction was found in all outcome measures during SLIT compared with the before period. The average annual cost/patient was 2672 before SLIT initiation and &#95;629/year during SLIT. Similar results were found for allergen subgroups. The asthma sub-analysis involved 41 children with SLIT and 35 controls. Again, SLIT patients showed a substantial reduction in outcome measures; the direct cost/patient over the whole follow-up (4 years) was &#95;1182 for SLIT patients and &#95;1100 for controls., Conclusion: High dose SLIT may be effective in reducing the cost of allergic rhinitis and asthma and comparably expensive to conventional treatment in children with allergic asthma over a 4 years follow-up.

Background: Walnut is the most common cause of allergic reactions to tree nuts, as reported by large population studies. Two major allergens of walnut have been identified up until now: a 2S albumin and a vicilin-like protein., Objective: This study was designed to identify the walnut major allergens in the Italian population and to compare the walnut IgE-binding profile in patients with or without pollen allergy., Methods: We selected 46 patients either with oral allergy syndrome confirmed by open oral challenge or with systemic symptoms after ingestion of walnut. These patients' sera were used for the immunoblotting of walnut extract; the identified allergens were purified by HPLC and sequenced. A peach-walnut cross-inhibition study was then performed., Results: The only major allergen recognized by our study population was a 9-kd lipid transfer protein (LTP), recognized by 37 patients. Two other minor allergens of approximately 9-kd molecular weight, both belonging to the vicilin family, were recognized by 10 patients. IgE binding to walnut LTP was completely inhibited by peach LTP., Conclusion: In Italian patients with walnut allergy confirmed by documented history of severe systemic reactions or by open oral food challenge, the major allergen is an LTP. The sensitization to this protein seems to be secondary to the sensitization to peach LTP, which acts as the primary sensitizer. LTP and vicilins were able to sensitize patients not allergic to pollen.

Although flow cytometry is increasingly used as a tool to diagnose hematologic malignancies, the reporting format of acute leukemia immunodiagnosis is still imprecise and sometimes vague, often reflecting old guidelines. Thus, the purpose of the present work was to make the reporting format for the immunological diagnosis of acute leukaemia easy and clear to understand. This work represents part of a more articulated series of technical guidelines that the Italian Society for Cytometry (GIC) is currently processing. Thirteen separate recommendations, covering all aspects of an acute leukemia cytometry report, are listed. According to our suggestions, the report must contain clear statements about: 1. demographic identification of patient; 2. identification of the hospital or division sending the sample; 3. type of specimen (bone marrow aspirate, peripheral blood, other biological fluids); 4. timing of observation (first diagnosis or follow-up); 5. diagnostic hypothesis made by the sender; 6. list of antigens and type of immunofluorescence analysis carried out; 7. absolute number of cells in the sample; 8. quality of the sample, in terms of viability; 9. general description of the gating procedure; 10. immunophenotype of blast cells; 11. description of cells surrounding blasts; 12. diagnostic conclusions; 13. definition of an antigen panel (when applicable) for the detection of minimal residual disease. As an example of a final report we present a case of acute myeloid leukaemia with t(8;21) translocation; in filling this report, we followed all the 13 points of the checklist described in the paper.

Background: In a previous study a 9-kd lipid-transfer protein (LTP) was identified as the major allergen of raw maize in a population of 22 anaphylactic patients. However, the stability of this protein in cooked maize is unknown., Objective: We investigated the allergenicity of 5 maize hybrids and its modification after different thermal treatments by using sera from anaphylactic patients and patients with positive double-blind, placebo-controlled food challenges., Methods: Five maize hybrids were extracted by using different methods, obtaining the water-soluble, zein, total zein, glutelin, and total protein fractions. The IgE-binding capacity of the different extracts, both raw and after thermal treatment, was investigated by means of SDS-PAGE immunoblotting. A 9-kd heat-stable allergen was purified by means of HPLC and sequenced. Changes in its secondary structure during and after heating from 25 degrees C to 100 degrees C were monitored by means of circular dichroism., Results: All raw maize hybrids showed similar protein and IgE-binding profiles. The SDS-PAGE of all the heat-treated hybrids demonstrated a decreased number of stained bands in respect to the raw samples. The IgE immunoblotting demonstrated that the major allergen of the water-soluble, total zein, total protein, and glutelin fractions was a 9-kd protein identified by means of amino acid sequence as an LTP and a sub-tilisin-chymotrypsin inhibitor (in total zein fraction). The IgE-binding capacity of this 9-kd protein remained unchanged after thermal treatments, even though circular dichroism demonstrated an altered secondary structure., Conclusions: Maize LTP maintains its IgE-binding capacity after heat treatment, thus being the most eligible candidate for a causative role in severe anaphylactic reactions to both raw and cooked maize.

Background: Few allergic reactions to grape are reported in the literature. In some cases an association with peach and cherry allergy was observed. No IgE-mediated reactions to wine have been described, and no grape major allergens have yet been identified., Objective: We describe several severe reactions to grape or wine. We characterized the grape major allergens and tried to identify the allergen in wine., Methods: We collected documented histories of allergic reactions to grape and wine. Grape allergens were identified by means of SDS-PAGE and immunoblotting and purified by means of HPLC. Using amino acid sequencing and mass spectrometry, we identified the family of proteins to which the allergens belong. Cross-reactivity with peach and cherry was evaluated by means of cross-wise inhibition experiments., Results: Eleven patients with reactions to grape and 3 with anaphylactic reactions to wine were recruited. The major allergens were an endochitinase 4A and a lipid-transfer protein (LTP) that was homologous to and cross-reactive with peach LTP. A 24-kd protein homologous to the cherry thaumatin-like allergen was a minor allergen. Endochitinase 4A is very likely the allergen in vino novello and in vino Fragolino., Conclusions: Grape and wine might cause severe allergic reactions in sensitive patients. The major allergens of grape are endochitinase 4A, which is also the allergen of wine, and an LTP cross-reacting with the peach major allergen.

Background: Almond proteins can cause severe anaphylactic reactions in susceptible individuals. The aim of this study was the identification of IgE-binding proteins in almonds and the characterisation of these proteins by N-terminal sequencing., Methods: Five sera were selected from individuals with a positive reaction to food challenge. Sodium dodecylsulphate-polyacrylamide gel electrophoresis and immunoblotting were performed on almond seed proteins. Purified IgE-binding proteins were tested for immunoblot inhibition with sera pre-incubated with extracts of hazelnut and walnut., Results: N-terminal sequences of the 12-, 30- and 45-kD proteins were obtained. The 45- and 30-kD proteins shared the same N terminus, with 60% homology to the conglutin gamma heavy chain from lupine seed (Lupinus albus) and to basic 7S globulin from soybean (Glycine max). The sequences of the N-terminal 12-kD protein and of an internal peptide obtained by endoproteinase digestion showed good homology to 2S albumin from English walnut (Jug r 1). Immunoblot inhibition experiments were performed and IgE binding to almond 2S albumin and conglutin gamma was detected in the presence of cross-reacting walnut or hazelnut antigens., Conclusions: Two IgE-binding almond proteins were N-terminally sequenced and identified as almond 2S albumin and conglutin gamma. Localisation and conservation of IgE binding in a 6-kD peptide obtained by endoproteinase digestion of 2S albumin was shown., (Copyright 2002 S. Karger AG, Basel)

Background: The hazelnut major allergens identified to date are an 18-kd protein homologous to Bet v 1 and a 14-kd allergen homologous to Bet v 2. No studies have reported hazelnut allergens recognized in patients with positive double-blind, placebo-controlled food challenge (DBPCFC) results or in patients allergic to hazelnut but not to birch., Objective: We characterized the hazelnut allergens by studying the IgE reactivity of 65 patients with positive DBPCFC results and 7 patients with severe anaphylaxis to hazelnut., Methods: Hazelnut allergens were identified by means of SDS-PAGE and IgE immunoblotting. Further characterization was done with amino acid sequencing, evaluation of the IgE-binding properties of raw and roasted hazelnut with enzyme allergosorbent test inhibition, assessment of cross-reactivity with different allergens by means of immunoblotting inhibition, and purification by means of HPLC., Results: All the sera from the patients with positive DBPCFC results recognized an 18- and a 47-kd allergen; other major allergens were at molecular weights of 32 and 35 kd. Binding to the 18-kd band was inhibited by birch extract, indicating its homology with the birch major allergen, and abolished in roasted hazelnut. The 47-kd allergen is a sucrose-binding protein, the 35-kd allergen is a legumin, and the 32-kd allergen is a 2S albumin. Patients with severe anaphylactic reactions to hazelnut showed specific IgE reactivity to a 9-kd allergen, totally inhibited by purified peach lipid-transfer protein (LTP), which was heat stable and, when purified, corresponded to an LTP., Conclusions: The major allergen of hazelnut is an 18-kd protein homologous to Bet v 1, and the 9-kd allergen is presumably an LTP. Other major allergens have molecular weights of 47, 32, and 35 kd.

We report an example of metastatic metanephric adenoma containing foci of papillary carcinoma in the primitive tumor arising in the left kidney of an 11-year-old girl. Histology revealed a monomorphous population of small cells with bland cytology arranged in pseudoglandular, tubular, papillary, and glomeruloid structures with frequent psammoma bodies. Intermixed there were foci of and small cavities lined by larger cells with eosinophilic cytoplasm and larger nuclei with small nucleoli. A regional lymph node contained metastatic deposits with the former histologic pattern. By immunohistochemistry the small cells were negative for carcinoembryonic antigen (CEA) and keratin 7 while these antibodies reacted positively in the larger cells. A fluorescent in-situ hybridization (FISH) study for chromosome 17 in imprints from the primitive tumor revealed 3 signals in about 10% of the nuclei while the rest was disomic. Disaggregated cells from the metastatic lymph node consistently revealed 2 signals for chromosomes 7, 16, and l7. Histology of the primitive tumor resembled the epithelial component of the so-called metanephric adenofibroma while the metastatic lymph node exhibited histologic and FISH genomic features of metanephic adenoma. Int J Surg Pathol 9(3):241-247, 2001

Background: Cereals are the most important nutritional component in the human diet. Food-induced allergic reactions to these substances therefore have serious implications, and exhaustive diagnosis is required. Such diagnosis is still difficult because of the incomplete knowledge about major cereal allergens. In particular, few food-induced allergic reactions to maize have been reported, and no information on the allergenic proteins is available., Objective: Having observed several anaphylactic reactions to maize, we planned a study to identify maize major allergens and cross-reactivity with other cereals, as well as to peach because the majority of patients also reacted to Prunoideae fruits., Methods: Twenty-two patients with systemic symptoms after maize ingestion and positive skin prick test responses and serum-specific IgE antibodies to maize were selected. The IgE-reactivity pattern was identified by SDS-PAGE and immunoblotting. The major allergen identified was then purified by HPLC and characterized by mass spectrometry, determination of the isoelectric point value, and N-terminal amino acid sequencing., Results: Sera from 19 (86%) of the 22 patients recognized a 9-kd protein, thus confirming this as the maize major allergen. This protein had an isoelectric point of greater than 9, a molecular mass of 9047.0 d, and no glycosylation. Determination of its N-terminal sequence showed that it was a lipid transfer protein (LTP). By using immunoblotting-inhibition experiments, we demonstrated that the LTP cross-reacts completely with rice and peach LTPs but not with wheat or barley LTPs. N-terminal sequence of the 16-kd allergen (recognized by 36% of patients) showed it to be the maize inhibitor of trypsin. This protein cross-reacts completely with grass, wheat, barley, and rice trypsin inhibitors., Conclusion: The major allergen of maize is an LTP with a molecular weight of 9 kd that is highly homologous with the peach LTP, the major allergen of the Prunoideae subfamily.

Background: Tree nuts are a common cause of food allergy in Europe. However, few studies deal with real food allergy to hazelnuts in subjects believed to be allergic to this food., Objective: We sought to select subjects with a history of allergic reactions on ingestion of hazelnut and determine how many of these have true allergy by means of the double-blind, placebo-controlled food challenge (DBPCFC)., Methods: Eighty-six subjects with a history of symptoms after hazelnut ingestion were recruited from 3 allergy centers (Milan, Zurich, and Copenhagen). All subjects underwent skin prick tests (SPTs) with aeroallergens and hazelnut, as well as having their specific hazelnut IgE levels determined. Diagnosis of clinical relevant food allergy was made on the basis of the DBPCFC., Results: Sixty-seven (77.9%) of 86 subjects had a positive DBPCFC result; 8 were placebo responders, and 11 were nonresponders. Of the 11 nonresponders, 4 had positive open-challenge test results. Of the DBPCFC-positive subjects, 87% also had positive skin test responses to birch pollen extract. Specific IgE determination for hazelnut (positive CAP response >/=0.7 kU/L [ie, class 2]) showed a sensitivity of 0.75, a positive predictive value (PPV) of 0.92, a specificity of 0.16, and a negative predictive value (NPV) of 0.05. Skin tests with commercial hazelnut extract produced a sensitivity of 0.89, a PPV of 0.92, a specificity of 0.05, and an NPV of 0.05. Skin tests with natural food produced a sensitivity of 0.88, a PPV of 0.94, a specificity of 0.27, and an NPV of 0.15., Conclusion: This study shows that hazelnut is an allergenic source that can cause real food allergy, as confirmed by DBPCFC. Skin and IgE tests demonstrated reasonable sensitivity and PPV but a very low specificity and NPV, thus implying that these should not be used to validate the diagnosis of food allergy to hazelnut.

Background: Apricots are widely grown in Europe, and allergic reactions are becoming more common, especially oral allergy syndrome. Apricot belongs to the botanical subfamily of Prunoideae, which includes peach, the major allergen of which was identified as a 9-kd protein, a lipid transfer protein (LTP)., Objective: The aim of the study was to evaluate the IgE reactivity pattern to an apricot extract in subjects with allergic reactions to apricot, as demonstrated by a positive oral challenge response., Methods: Thirty patients were investigated. All the patients displayed oral allergy syndrome (2 with systemic reactions) to apricot, with positive open food challenge responses, skin prick test responses, and serum-specific IgE antibodies to apricot. The IgE reactivity pattern to apricot extract was identified by using SDS-PAGE and immunoblotting. The major allergen, a 9-kd protein, was then purified by HPLC and characterized by periodic acid-Schiff stain, isoelectric point determination, and N-terminal amino acid sequencing., Results: The sera from all patients allergic to apricot recognized the 9-kd protein, whereas none of the other allergens, with molecular weights from 15 to 80 kd, acted as a major allergen. The 9-kd allergen has an isoelectric point of 8.7 and is not glycosylated. Determination of the N-terminal 34 amino acid sequence showed that it belongs to the LTP family, with a 94% homology with the LTP from peach. IgE blotting of the apricot extract was completely inhibited by the 9-kd purified LTP from peach., Conclusions: The major allergen of apricot is an LTP, which is highly cross-reactive with the LTP from peach.

Among vegetable foods peach (Prunus persica) has been recognized as a significant cause of allergy. The protein, which is considered to be the major peach allergen, has been named Pru p 1. Because peaches are consumed both as fresh fruits and after processing to obtain peach juice, nectar, jam, syrupy peach, etc., research was carried out to identify a technological process for production of hypo- or nonallergenic peach-based products. SDS-PAGE and immunoblotting analysis of extracts prepared from four commercial peach nectars showed that the Pru p 1 was not removed, and neither was its allergenic activity decreased by technological treatments carried out for nectar production. Some treatments oriented toward a removal of or, at least, a decrease in the allergenic power were assumed and verified at laboratory scale. A variable considered was heat treatment at 121 degrees C for 10 and 30 min: this treatment was not able to decrease the allergenicity of the Pru p 1 protein. Furthermore, the protein band was still present even after 60-min reaction with two different acidic proteases. The two technological treatments that were found to decrease the major allergen of peach were chemical lye peeling of fruits and ultrafiltration of juice through membranes with suitable cutoff. On the basis of the results obtained from this research, a processing flow sheet was defined to obtain hypoallergenic or probably nonallergenic limpid juices and nectars. These products may represent, besides finished foods, intermediates to obtain various products after addition of further ingredients such as pectins, sugars, and fiber.

The major protein allergen of peach (Prunus persica), Pru p 1, has recently been identified as a lipid transfer protein (LTP). The complete primary structure of Pru p 1, obtained by direct amino acid sequence and liquid chromatography-mass spectrometry (LC-MS) analyses with the purified protein, is described here. The protein consists of 91 amino acids with a calculated molecular mass of 9178 Da. The amino acid sequence contains eight strictly conserved cysteines, as do all known LTPs, but secondary structure predictions failed to classify the peach 9 kDa protein as an 'all-alpha type', due to the high frequency of amino acids (nine prolines) disrupting alpha helices. Although the sequence similarity with maize LTP is only 63%, out of the 25 amino acids forming the inner surface of the tunnel-like hydrophobic cavity in maize ns-LTP 16 are identical and 7 similar in the peach homolog, supporting the hypothesis of a similar function.

Background: Allergy to fresh fruits and vegetables is mostly observed in subjects with pollinosis, especially from birch, because of cross-reacting allergens in vegetable foods and pollens. However, allergic reactions to fruits, specifically Rosaceae fruits, have been reported in subjects without pollinosis., Objective: This study evaluated the pattern of IgE reactivity, identifying the allergen responsible in 2 groups of patients with oral allergy syndrome to peach with or without birch pollinosis., Methods: The allergenic components of peach were detected by SDS-PAGE and immunoblotting. The major peach allergen was purified by HPLC with a cation-exchange column followed by gel filtration chromatography. Its IgE-binding capacity and its homology with the protein of the crude extract were demonstrated by immunoblotting inhibition techniques. To better characterize this allergen, periodic acid-Schiff stain and isoelectrofocusing were used. The amino acid sequencing was done with a gas-phase sequencer., Results: SDS-PAGE and immunoblotting of the 15 patients allergic to peach, 8 without and 7 with birch pollinosis, showed that they all recognized a protein with a molecular weight of 9 kd. This was the only allergen recognized by patients not sensitized to pollen, whereas the birch pollen-sensitive patients had IgE binding to other allergenic proteins at higher molecular weights. The purified 9-kd protein retained its IgE-binding capacity, was negative to periodic acid-Schiff stain, and had an isoelectric point value of greater than 9. A search in the Swiss Prot Bank showed this was a lipid transfer protein, belonging to a group of molecules involved in the defensive system of plants., Conclusions: The major allergen of peach is a 9-kd protein belonging to the group of lipid transfer proteins. This is the only allergen recognized by patients allergic to peach but not sensitized to birch pollen.

Background: Allergic reactions to fruits and vegetables are among the most frequent food allergies in adults. Kiwi fruit (Actinidia chinensis) is commonly involved, causing local mucosal, systemic, or both types of symptoms by an IgE-mediated mechanism. In a previous study on 30 patients allergic to kiwi, we identified a major allergen of 30 kd against which all sera tested clearly reacted. Other allergens were detected at 12, 24, and 28 kd., Objective: The aim of this study was to fully characterize the major kiwi fruit allergen of 30 kd., Methods: Allergens were separated and purified by high-performance liquid chromatography with anion-exchange columns. The purity of the single proteins was checked by sodium dodecylsulfate-polyacrylamide gel electrophoresis, and their allergenicity was checked by immunoblotting with a pool of sera from patients allergic to kiwi. The allergens were characterized by isoelectrofocusing and amino acid sequencing, and periodic acid-Schiff stain was used to detect glycoproteins., Results: Proteins of 30, 28, 24, and 17 kd were purified by high-performance liquid chromatography. IgE binding indicated the 30 kd protein, which showed an isoelectric point of 3.5, as the major allergen of kiwi. Determination of its partial amino acid sequence and comparison with the Swiss Protein Bank showed that this was actinidin, the main protein component of kiwi. The 24 and 28 kd proteins had the same N-terminal sequence, which did not correspond to any known protein. The 17 kd protein had a blocked N-terminal sequence., Conclusions: These results demonstrate that the major allergen of kiwi fruit, Act c 1, is actinidin, a proteolytic enzyme belonging to the class of thiol-proteases. Two other allergens of 24 and 28 kd appear identical on amino acid sequencing.

We have been studying the immune system of healthy centenarians for many years, and they provide the best example of successful aging. They are people who have escaped major age-related diseases and reached the extreme limit of human life in good clinical condition. In most cases, histories of centenarians reveal them to be free of cancer, dementia, diabetes, cardiovascular diseases, and cataracts. Moreover, in order to reach such an advanced age, they should be equipped with well preserved and efficient immuno- and defense mechanisms, and optimal combinations of an appropriate lifestyle and genetic background. Using this approach, several paradoxes emerged as far as the immune system of centenarians is concerned, regarding: i) humoral immunity (increase in plasma immunoglobulins and nonorgan-specific autoantibodies, decrease in B cell number and lack of organ-specific autoantibodies); ii) cellular immunity (well preserved number of "virgin" T cells, a relatively intact T cell repertoire despite a thymus involuting since puberty, increased number of cells with markers of NK activity); iii) decreased peripheral blood lymphocyte tendency to programmed cell death, associated with a well preserved mitochondria functionality and intracellular bcl-2 levels. An age-related increase in the levels of adhesion molecule present on lymphocyte plasma-membrane, accompanied by a complex reshaping of the cytokine network, must be added to this scenario. All our data fit the hypothesis that a complex, unpredicted remodeling of the immune system occurs with age. In the present review it is underlined how flow cytometry has been used to study most of the above mentioned aspects of immunosenescence, and to establish new age-related reference values.

Objective: To study the V beta T-cell repertoire in peripheral blood lymphocytes (PBL) during acute HIV syndrome by using several anti-V beta monoclonal antibodies (MAb) and to analyse its functionality by stimulating PBL with superantigens (SAg) such as Staphylococcus aureus enterotoxins., Methods: Cytofluorimetric analysis of V beta T-cell-receptor expression was performed on PBL from eight patients with symptomatic, acute HIV-1 primary infection, showing a dramatic decrease of CD4+ PBL accompanied by a marked increase in activated/memory CD8+ T cells, and on 12 age- and sex-matched healthy controls. PBL were then isolated, stimulated with different SAg, anti-CD3 MAb or phytohaemagglutinin and cultured for 3 days. PBL capability to progress through cell cycle was studied by the classic cytofluorimetric method of bromodeoxyuridine incorporation and DNA staining with propidium iodide., Results: Despite the presence of a few expansions of some V beta families among CD8+ T lymphocytes, no gross alterations in T-cell repertoire were present in patients with acute HIV syndrome. Its functionality was maintained overall, as PBL responsiveness to SAg was well preserved. Interestingly, all CD8+ T cells, although bearing different V beta T-cell receptors, expressed marked signs of activation, i.e., CD45R0, CD38 and major histocompatibility complex class II molecules, and also high amounts of CD11a and CD18., Conclusions: Our data suggest, at least in the early phases and in the acute form of the infection, that HIV is not likely to act as a SAg. However, further studies are needed to analyse other sites, such as lymph nodes, where HIV could exert other, significant effects, and to study the expression of other V beta families than those investigated here.

Symptoms appear immediately at the site of contact with the food when a susceptible allergic patients eats the offending food. Allergy to fresh fruits and vegetables, associated with all kinds of pollinosis, is the more frequent cause of Oral Allergy Syndrome.

Background: Oral allergy syndrome in response to fruits and vegetables frequently occurs as clusters of hypersensitivity to members of the same botanical family, for which the immunologic basis lies in a number of common allergens, most of them still unidentified., Objective: This study was designed to assess the in vivo and in vitro cross-reactivity between fruits of the Prunoideae subfamily (i.e., peach, cherry, apricot, and plum) and to identify their major allergens and the cross-reactivity of the peach extract with grass and birch pollen., Methods: The in vivo study was conducted by skin prick tests and open food challenges with fresh fruits in 23 patients with oral allergy syndrome for peach and positive skin prick test and RAST results for the other Prunoideae. In vitro sodium dodecylsulfate-polyacrylamide gel electrophoresis was followed by immunoblotting and immunoblotting-inhibition., Results: A 13 kd component was identified as the only major allergen common to all the Prunoideae, the other major allergens were found at 14 kd in peach and at 30 kd in cherry. Immunoblotting inhibition showed wide cross-reactivity within the Prunoideae, whereas grass and birch pollen partially inhibited the peach blotting., Conclusions: Clinical cross-reactivity to Prunoideae is essentially due to a common 13 kd IgE-binding component, which seems to be the most important major allergen of this subfamily, not shared with grass and birch pollen.

The aim of this study was to assess the ability of nedocromil sodium (NS) to prevent the immediate asthmatic reaction and the increase in the serum level of heat stable neutrophil chemotactic activity (HS-NCA) induced by antigen inhalation. In a double-blind, cross-over study, 13 atopic subjects affected with seasonal asthma underwent a bronchial provocation test with a preselected dose of grass pollen allergen (enough to cause a decrease of > or = 20% in FEV1:FEV1 PD20) after pre-treatment with 4 mg NS or placebo. Serum samples were withdrawn from 11 subjects for HS-NCA determination. After NS administration the decrease in FEV1 was significantly less than after placebo administration at all time points after challenge (2 min P = 0.0004; 7 min, P = 0.0005; 17 min P = 0.0002 and 27 min P = 0.0005). The percentage increase in HS-NCA was significantly higher after placebo than after NS inhalation, both 10 (P = 0.0048) and 20 (P = 0.0068) min after challenge. Our study confirms previous investigations, showing that NS inhibits the immediate asthmatic response to allergen inhalation in atopic, asthmatic subjects and moreover it shows that this drug prevents in vivo the increase of the serum HS-NCA. This last finding has not been previously reported.

A double-blind, placebo-controlled study was conducted to evaluate the efficacy and safety of immunotherapy (IT) with a partially purified alginate-conjugated extract of Parietaria judaica (Conjuvac Parietaria, Dome/Hollister-Stier) in patients suffering from rhinoconjunctivitis caused by Parietaria pollen. Eighteen patients (10 women, 8 men, mean age 35 years) received active treatment and 17 (10 women, 7 men, mean age 42.5 years) received placebo. Actively treated patients had significantly lower nasal symptom/medication scores (running nose P = 0.0087 and sneezing P = 0.048) during the Parietaria pollen season. Significant decreases in specific skin (P < 0.01), nasal (P < 0.05), and conjunctival (P < 0.01) reactivity to the Parietaria extract and significant increases of specific IgG (P < 0.001), IgG1 (P < 0.001), and IgG4 (P < 0.001) in actively treated patients, but not in placebo, were found. IT was well tolerated, the active extract inducing five mild systemic reactions (four rhinitis and one urticaria) and placebo two (rhinitis). A significant correlation was found between low skin reactivity and high specific IgG (P = 0.0002) and IgG4 (P = 0.036). These findings indicate that IT with a partially purified P. judaica extract is an effective and safe treatment for Parietaria pollen allergy. The correlation between low immediate skin reactivity and high specific IgG and IgG4 suggests that, at least in the studied cutaneous model, these antibodies may exert a blocking effect.

The presence of a subset of T lymphocytes with an irregular phenotype has been identified in the peripheral blood of 22 subjects, selected among more than 5000 individuals evaluated for peripheral blood lymphocyte subset distribution as part of a routine procedure. By Southern Blot Analysis 13 out of 14 of the analysed samples presented additional non-germline bands, indicative of monoclonal or oligoclonal T cell expansions. Moreover, the cytometric analysis showed that 7 out of 19 analysed samples were restricted for a subfamily of the TCR variable regions. Thus, lymphocyte subsets with phenotypic irregularities could represent the clonally driven expansion of otherwise normally subpopulations, which may be present in peripheral blood below the limit of detection of routine phenotypic analyses. Such clonal populations could exert a regulatory activity either on the pathogenetic mechanisms of the disease or in maintaining homeostasis in healthy people.

Allergy to fresh fruits and vegetables is IgE-mediated. Its main clinical features are local, such as oral pruritus and swelling of lips and tongue but systemic symptoms such as urticaria, asthma, or anaphylactic shock may occur. Clinical associations with allergic rhinitis due to cross-reactive antigens of pollens and foods are frequent.

The production of cytokines during aging, except interleukin (IL)-2, has been neglected in humans. We measured the in vitro production of IL-6, tumor necrosis factor (TNF)-alpha, interferon (IFN)-gamma and IL-1 beta by peripheral mononuclear cells from selected healthy young (mean age 26.8 years) and aged (mean age 80.2 years) subjects. Significant increases of IL-6, TNF-alpha and IL-1 beta levels were found in mitogen-stimulated cultures from aged donors, occurring at 24 to 72 h after stimulation. No significant differences were observed for IFN-gamma production. Proliferative capability of cells stimulated with PHA was not impaired in aged subjects. Since the amounts of all cytokines studied were similar in unstimulated cultures from young and aged subjects, and also serum levels of TNF-alpha did not differ, these data indicate that the cellular machinery for the production of these cytokines is well preserved in aging, and also that cells from old people are able to up-regulate their production in response to appropriate stimuli. The increases in cytokine synthesis were not dependent on changes in the number of monocytes, nor were they related to the significant rise of CD45RO+, and the concomitant decrease of CD45RA+, occurring in both CD4+ and CD8+ lymphocytes from aged subjects. The increased production of pro-inflammatory cytokines by stimulated mononuclear cells of healthy aged subjects may be relevant to several aspects of age-associated pathological events, including atherosclerosis, osteoporosis, fibrosis and dementia.

The reliability and safety of the oral challenge procedure were evaluated in the diagnosis and prevention of intolerance to nonsteroidal anti-inflammatory drugs (NSAIDs). 112 NSAID-intolerant patients were submitted to oral challenge with aspirin (acetylsalicylic acid), dipyrone, paracetamol, imidazole-hydroxybenzoate or nimesulide to confirm historic intolerance and to evaluate tolerance to other NSAIDs. A significant correlation was demonstrated between history of intolerance and the results of oral challenge in aspirin-intolerant patients (p < 0.001). Of 237 challenges with various NSAIDs and paracetamol in 101 patients, 19 challenges were positive and 2 patients developed anaphylactic shock. The ratio of positive: total challenges with alternative NSAIDs and paracetamol were as follows: 7 of 83 for paracetamol, 2 of 49 for imidazole-hydroxybenzoate and 0 of 30 for nimesulide. On the basis of these results, nimesulide was evaluated as an alternative NSAID in a second group of 284 NSAID-intolerant patients. Challenge with nimesulide elicited a positive response in 14 patients (in 6 patients the response was delayed), although adverse reactions were mild in all instances. In conclusion, a positive history of intolerance to a given NSAID is a sufficient reason to contraindicate its use even for diagnostic purposes, e.g. oral challenge. Oral challenge should therefore be restricted to the assessment of the tolerability of alternative NSAIDs. In the present study nimesulide appeared to be the safest currently available alternative in patients with NSAID intolerance.

Several immunological abnormalities have been observed in ataxia-telangiectasia (AT), the most consistent being defects of immunoglobulin isotypes, decreased T-cell numbers, and reduced proliferative responses to mitogens. We examined the distribution of T lymphocytes expressing distinctive surface Ag characteristic of "naive" (CD45RA+) and "memory" (CD29+, CD45RO+) T cells, in both CD4+ and CD8+ (bright and dim) lymphocytes from 13 AT patients, compared with healthy age-matched controls. We found that, irrespective of age, patients with AT had a severe deficiency of CD4+/CD45RA+ lymphocytes. This decrease accounted for the reduction of total CD4+ cells, since the absolute numbers of memory CD4+ cells were not significantly different in AT and in controls. Functional tests revealed poor proliferative responses to phytohemagglutinin and normal responses to soluble Ag (tetanus toxoid) in AT patients. These data fit with the distribution of naive and memory cells, which are known to respond predominantly to mitogens or to recall Ag, respectively. CD45RA molecules were normally expressed on CD8+ lymphocytes. This rules out a generalized defect of regulation or differential splicing as the cause of defective expression of CD45RA on CD4+ cells. The selective deficiency of CD4+CD45RA+ may provide a cellular basis for some functional T-cell abnormalities of AT patients. Furthermore, it might practically serve for an early, or even prenatal, diagnosis of this disease.

A study of 80 consecutive cutaneous malignant melanomas in 78 patients older than 60 years, with an average of 73 (60-93), is presented. The following data were considered: age, sex, localization, stage, other synchronous or metachronous malignant tumors, excluded basal cell epitheliomas, histological type and depth of cutaneous invasion according to Clark's levels and Breslow's measure. The follow up lasted 1 to 156 months with an average of 39 months. All patients were treated by surgical removal of the cutaneous lesion or the regional lymph node metastases, if they appeared. No prophylactic lymph node dissections were performed. There were 45 women and 33 men. The histological type was superficial spreading (ES) in 17 women and 7 men; nodular (NOD) in 18 and 16; lentigo maligna melanoma (LM) in 4 and 4 and acral in 7 and 7. Relations between sex, localization, histological type and actuarial survival curves can be observed in Figs. 5, 6, 7, 8 and Tables 1, 2. Even considering the shortcomings of this retrospective analysis and a relatively short follow up for melanomas, this study points out that the thickness of the lesion became prognostically significant in relation to survival, beyond 3.7 mm (p < 0.001); but when the tumors were thinner, this group of patients showed other causes of death (MPOC) with a greater frequency than those related to tumoral progression (MPM) (p < 0.001) and that maybe this should be taken into account when planning treatment for melanomas in elderly patients. Other malignant synchronous or metachronous tumors were found in 19% of the patients (Table 2).

The presence of immunoreactive ACTH molecules on phagocytic cells from the freshwater snails Planorbarius corneus and Lymnaea stagnalis was shown by cytofluorimetric analysis. The role of ACTH in phagocytosis and in the release of biogenic amines, two biological responses that may be taken as ancestral types of immune response and stress, respectively, has also been evaluated. ACTH markedly increased the phagocytosis of Staphylococcus aureus by P. corneus haemocytes and caused the release of biogenic amines from such cells into the serum. These data, as well as tracing the ancestral physiological role of ACTH, favour the hypothesis that immune and neuroendocrine systems share a common evolutionary origin.

The sensitivity and specificity of the Pharmacia CAP System, a new in vitro assay for measuring IgE antibodies, were evaluated in a multicentric study on 286 allergic patients and 243 normal subjects. Sensitivity was 95.5%, with results significantly better than RAST for some allergens, and specificity was excellent, 98.1%.

The peripheral T cell receptor repertoire is mainly controlled by the processes of positive and negative selection occurring in the thymus. Studies in normal or transgenic mice have provided compelling evidence for both negative and positive selection. Negative selection is characterized by partial or total disappearance from the periphery of T cells expressing certain C beta regions, which are normally present in the thymus. Positive selection is chiefly characterized, in the periphery, by an imbalanced ratio between CD4+ and CD8+ T cells expressing a given V domain. To date little information concerning positive and negative selection has been available in man. We studied the distribution of 4 V beta domains on CD4+ and CD8+ peripheral T cells of 34 healthy individuals with a wide age-range (0-96 years). One of the V beta domain studied, V beta 6.7, is preferentially expressed on CD4+ compared to CD8+ T cells (p greater than 0.001). No significant differences were observed using the other V beta domain-specific monoclonal antibodies (V beta 5.2-5.3, 8 or 12). These results provide evidence that a process of thymic education, similar to that described in murine animal models, may also take place in man.

Phenotype and proliferative ability of peripheral blood lymphocytes from 15 noninstitutionalized children affected with Down Syndrome (DS), in apparently good health, were studied and compared with those of 16 healthy control children of the same age. A complex derangement of all the major peripheral blood cell subsets, i.e., B cells, T cells, and natural killer (NK) cells, was present in DS children. A significant decrease of the absolute number of circulating lymphocytes, a marked and significant decrease of B lymphocyte absolute number and percentage, and dramatic modifications of the T-cell subsets were observed. The absolute number of CD4+ cells was significantly decreased, whereas CD8+ cells increased significantly in percentage but not in absolute number. A derangement of cells bearing markers associated with NK activity, such as CD57, CD16, and CD56, was observed. Among the most important alterations, the presence of a high number of CD57+, CD16- cells, of CD57+, CD8+ lymphocytes, and of CD3+, CD56+ lymphocytes was seen. Many of these alterations are similar to those characteristic of chromosomally normal subjects of advanced age. The hypothesis that the reduced thymic endocrine activity and the zinc deficiency characteristic of DS are responsible for the derangement of T and NK subsets is discussed.

Reference is made to a series of 620 cases of Raynaud's phenomenon observed at the University of Milan between 1951 and 1977. The following parameters are analysed: sex, age, aetiology (where known), incidence of involvement of the four limbs, presence or otherwise of a complete sign, percentage of trophic lesions, arteriographic picture, lab examinations performed. 534 thoracic sympathectomies were performed in 464 patients with Raynaud or Raynaud-like forms. The results are described.

In 15 subjects immunized with two doses of purified adjuvant-free tetanus toxoid, the behaviour of specific IgE and hemoagglutinative antibodies was studied. In the majority of the subject the presence of specific IgE and hemoagglutinative antibodies was observed upon a control carried out 15 days after immunization. After reintroduction of antigen an increase both of specific IgE and hemoagglutinative antibodies was observed. No subject who had presented specific IgE in the serum after the first inoculation of tetanus toxoid showed local or general reaction after reintroduction.

The aim of this prospective study was to evaluate the incidence of allergic reactions to drugs compared to other kinds of medical emergencies admitted to the main Hospital in Milan during a 6 months period. At the same time we drew a list of drugs most frequently involved in allergic reactions, and a list of the most frequent symptoms. Using special forms, the medical staff collected patients' data: age, history of atopy, identification of the drug causing the reaction, and any previous reactions. Among 11,407 cases of medical emergencies, we found 163 (1.43%) patients showing drug reactions: the mean age was 27.3; 58.90% were female; atopy was present in 16.56%. The drugs most frequently involved were: pyrazon group (22%); ASA (20.86%); penicillin and derivatives (9.20%); sulfa drugs (6.14%); group B vitamins (4.30%); tetanus toxoid (4.30%); hyposensitizing extracts (3.68%); propionic acid derivatives (2.46%); paracetamol (1.84%); indomethacin (1.23%); rifampicin (1.23%); erythromycin (1.23%); glafenine (1.23%); others (17.80%). Urticaria and/or angioedema were the most frequent symptoms (86.51%), then anaphylactic shock (9.81%) and asthma (3.68%) with regard to anaphylactic shock only 6.20% of the patients had had a previous reaction to the same drug. From these data we can see that the incidence of drug reactions is very low compared to other medical emergencies; penicillin evidenced fewer reactions than expected, while the pyrazon group and ASA confirmed the data from literature.

Twenty patients with chronic urticaria exacerbated by aspirin or tartrazine were challenged with the offending substance after pre-treatment with either oral sodium cromoglycate or placebo in a double-blind, crossover study. The sequential vascular response test (SVR) described by Fisherman and Cohen 1-4 was used to assess response to challenge. Eighteen patients were protected by oral sodium cromoglycate and one by placebo, taking as a positive response an increase in the bleeding time of 100% or more over control values.