Oral myeloid cells uptake allergoids coupled to mannan driving Th1/Treg responses upon sublingual delivery in mice.

BACKGROUND: Polymerized allergoids coupled to nonoxidized mannan (PM-allergoids) may represent novel vaccines targeting dendritic cells (DCs). PM-allergoids are better captured by DCs than native allergens and favor Th1/Treg cell responses upon subcutaneous injection. Herein we have studied in mice the in vivo immunogenicity of PM-allergoids administered sublingually in comparison with native allergens. METHODS: Three immunization protocols (4-8 weeks long) were used in Balb/c mice. Serum antibody levels were tested by ELISA. Cell responses (proliferation, cytokines, and Tregs) were assayed by flow cytometry in spleen and lymph nodes (LNs). Allergen uptake was measured by flow cytometry in myeloid sublingual cells. RESULTS: A quick antibody response and higher IgG2a/IgE ratio were observed with PM-allergoids. Moreover, stronger specific proliferative responses were seen in both submandibular LNs and spleen cells assayed in vitro. This was accompanied by a higher IFNγ/IL-4 ratio with a quick IL-10 production by submandibular LN cells. An increase in CD4+ CD25high FOXP3+ Treg cells was detected in LNs and spleen of mice treated with PM-allergoids. These allergoids were better captured than native allergens by antigen-presenting (CD45+ MHC-II+ ) cells obtained from the sublingual mucosa, including DCs (CD11b+ ) and macrophages (CD64+ ). Importantly, all the differential effects induced by PM-allergoids were abolished when using oxidized instead of nonoxidized PM-allergoids. CONCLUSION: Our results demonstrate for the first time that PM-allergoids administered through the sublingual route promote the generation of Th1 and FOXP3+ Treg cells in a greater extent than native allergens by mechanisms that might well involve their better uptake by oral antigen-presenting cells.

Allergenome characterization of the mosquito Aedes aegypti.

BACKGROUND: Saliva and muscle-derived mosquito allergens have been purified and characterized. However, the complete set of allergens remains to be elucidated. In this study, we identified and characterized IgE-binding proteins from the mosquito species Aedes aegypti. METHODS: Serum was obtained from 15 allergic individuals with asthma and/or rhinitis and sensitized to mosquito. IgE binding was determined by ELISA. Total proteins from freeze-dried bodies of A. aegypti were extracted and IgE-reactive proteins were identified by 2D gel electrophoresis, followed by Western blot with pooled or individual sera. IgE-reactive spots were further characterized by mass spectrometry. RESULTS: Twenty-five IgE-reactive spots were identified, corresponding to 10 different proteins, some of which appeared as different variants or isoforms. Heat-shock cognate 70 (HSC-70) and tropomyosin showed IgE reactivity with 60% of the sera, lysosomal aspartic protease, and "AAEL006070-PA" (Uniprot: Q177P3) with 40% and the other proteins with <33.3% of the sera. Different variants or isoforms of tropomyosin, arginine or creatine kinase, glyceraldehyde-3-phosphate dehydrogenase (GPDH), calcium-binding protein, and phosphoglycerate mutase were also identified. The mixture of three allergens (Aed a 6, Aed a 8, and Aed a 10) seems to identify more than 80% of A. aegypti-sensitized individuals, indicating that these allergens should be considered when designing of improved mosquito allergy diagnostic tools. CONCLUSIONS: The newly identified allergens may play a role in the pathophysiology of mosquito allergy in the tropics, and some of them might be important arthropod-related proteins involved in cross-reactivity between A. aegypti and other allergenic arthropods.

Safety of immunotherapy with glutaraldehyde modified allergen extracts in children and adults.

BACKGROUND: Glutaraldehyde-modified natural allergen extracts show significant reduction in the IgE-binding capacity and proteolytic activity. This allows the administration of higher doses in a shorter period of time, and to mix different allergen extracts. OBJECTIVE: Evaluate the safety of different concentrations and mixtures of glutaraldehyde-modified allergen extracts in a large group of paediatric and adult patients undergoing specific immunotherapy treatment. MATERIALS AND METHODS: 1855 patients (1156 adults and 699 children), suffering from rhinoconjunctivitis and/or asthma, participated in an observational multicentre cohort study, to evaluate the safety of immunotherapy using vaccines containing modified allergen extracts. Patients were monosensitised, or polysensitised, and received a therapeutic vaccine containing polymerised allergen extracts adsorbed onto aluminium hydroxide. Safety was assessed by recording all side reactions related to immunotherapy. RESULTS: The clinically relevant local reactions totalled 120, (90 immediate and 30 delayed) (1.02% of injections). Of them, 31 (0.26% of injections) occurred in children (26 immediate and 5 delayed) and 89 (0.76% of injections) in adults (64 immediate and 25 delayed). There were 38 systemic reactions. Eleven reactions were immediate (9 of grade 1 and 2 of grade 2) and 27 delayed (22 of grade 1 and 5 of grade 2). There were seven grade 2 systemic reactions (0.06% of the injections). No differences (P>0.05) in the number of reactions were observed between adults and children and between treatments were found in systemic reactions. All systemic reactions were mild and resolved spontaneously without the need of medication. CONCLUSION: Specific immunotherapy using natural modified allergen vaccines is safe to treat allergic patients, even at higher doses and in mixtures of unrelated allergen extracts. The percentage of adverse reactions detected is lower than those reported in the literature with native unmodified allergen extracts.

Cluster immunotherapy with a glutaraldehyde-modified mixture of grasses results in an improvement in specific nasal provocation tests in less than 2.5 months of treatment.

BACKGROUND: Cluster immunotherapy is becoming increasingly used. It allows for a rapid build up phase and the administration of higher doses of allergen in a shorter period of time. OBJECTIVES: To evaluate the effect of short-term pre-seasonal immunotherapy using a glutaraldeyde-modified allergen vaccine in reducing specific nasal hyperreactivity in nasal challenge tests. MATERIALS AND METHODS: Thirty-three patients were selected. All patients had a positive history of allergic rhinitis and skin tests to grass pollen, although most of them (72.7%) were sensitized to other allergens as well. The study was conducted outside of the pollen season and the patients did not receive any pharmacological treatment during this period of time. Two randomized groups of patients were established; Group A: 22 patients (13 females and nine males) and Group B, 11 control patients (seven females and four males). Patients in Group A received immunotherapy with a vaccine containing 50% of the wild grasses Trisetum paniceum and Dactylis glomerata. All patients underwent titrated nasal provocation tests (NPT) before and after completion of the study (2.3 and 2.8 months for Groups A and B, respectively). The administration schedule consisted of 0.1 and 0.2 mL at day 1, followed by 0.3 and 0.5 mL at day 7, 0.5 mL after 2 weeks followed by 0.5 mL monthly. A single vial was used containing an allergen concentration of 10 000 TU/mL (105 microg of total protein and 24.6 microg of Group 1 plus 5 allergens/mL). A mean of 6.5 injections were administered to Group A patients between NPTs. RESULTS: There were no significant differences between both groups at the beginning of the study (P=0.48). At the end, only Group A patients needed significant greater threshold concentrations for a positive NPT than at the beginning (P=0.002). CONCLUSIONS: A short-term cluster pre-seasonal inmunotherapy with a modified vaccine containing a mixture of grass pollen is effective as determined by an objective measure after only a mean 2.3 months of treatment.

MV140, a sublingual polyvalent bacterial preparation to treat recurrent urinary tract infections, licenses human dendritic cells for generating Th1, Th17, and IL-10 responses via Syk and MyD88.

Recurrent urinary tract infections (RUTIs) are one of the most common bacterial infectious diseases, especially in women. Antibiotics remain the mainstay of treatment, but their overuse is associated with antibiotic-resistant infections and deleterious effects in the microbiota. Therefore, alternative approaches are fully demanded. Sublingual immunization with MV140 (Uromune), a polyvalent bacterial preparation (PBP) of whole heat-inactivated bacteria, demonstrated clinical efficacy for the treatment of RUTIs, but the involved immunological mechanisms remain unknown. Herein, we demonstrated that MV140 endorses human dendritic cells (DCs) with the capacity to generate Th1/Th17 and IL-10-producing T cells by mechanisms depending on spleen tyrosine kinase (Syk)- and myeloid differentiation primary response gene 88 (MyD88)-mediated pathways. MV140-induced activation of nuclear factor κB (NF-κB) and p38 in human DCs is essential for the generated Th1/Th17 and IL-10 immune responses whereas c-Jun N-terminal Kinase (JNK) and extracellular-signal regulated kinase (ERK) contribute to Th1 and IL-10 responses, respectively. Sublingual immunization of BALB/c mice with MV140 also induces potent systemic Th1/Th17 and IL-10 responses in vivo. We uncover immunological mechanisms underlying the way of action of MV140, which might well also contribute to understand the rational use of specific PBPs in other clinical conditions with potential high risk of recurrent infections.

Involvement of IFNbeta on IFNgamma and nitric oxide (NO) production by bone marrow (BM) cells in response to lipopolysaccharide.

Bone marrow (BM) cells fractioned in Percoll gradients yield a low-density fraction (Fr3) highly enriched in suppressor activity. Previously, it has been demonstrated that BM associated suppressor activity was mediated by early myeloid cells, through a mechanism dependent on endogenous IFNgamma and nitric oxide production after bacterial stimuli, e.g. lipopolysaccharide (LPS). However, the mechanism(s) through which the IFNgamma is produced in BM has not yet been fully elucidated. Therefore, in the present study we investigated the involvement of IL-12, IL-18 and IFNbeta on the production of IFNgamma and nitric oxide in cultures of BM Fr3 cells, and characterized the IFNgamma-producing cells, in response to LPS. The results show that both IL-12 and IFNbeta, but not IL-18, are involved on IFNgamma production. However, only IFNbeta appears to be critical on nitric oxide production. Furthermore, we found that cells of the Thy1.2+CD3+ phenotype produce IFNgamma and are tightly involved on nitric oxide production by BM Fr3 cells. In conclusion, IFNbeta appears to be critical on IFNgamma- and nitric oxide production by BM cells in response to LPS, through a mechanism that is dependent on Thy1.2+CD3+ IFNgamma-producing cells.

Human colon adenocarcinomas express a MUC1-associated novel carbohydrate epitope on core mucin glycans defined by a monoclonal antibody (A10) raised against murine Ehrlich tumor cells.

A monoclonal antibody (mAb; A10) raised against murine Ehrlich tumor cell surface carbohydrates was tested for reactivity with human normal and malignant tissues. A10 reacted strongly, with a high proportion of adenocarcinomas arising from colon and other tissues but not with breast carcinomas or other malignant tumors. Normal tissues were virtually A10 unreactive, except for the duct cells from breast and pancreas and some bronchial mucosae. Ultrastructural studies showed mAb A10 immunolabeling of both microvilli and mucin droplets in colon cancer cells but not in normal absorptive or globet cells. A10 reacted strongly with mucin-enriched fractions from colon cancer tissues and HT-29 xenografts but not from normal colon tissues. A10 epitope was carried on MUC1 derived from colon adenocarcinomas and probably on other mucin species, although not on MUC2 molecules. A10 epitope was resistant to exoglycosidases and periodate oxidation but sensitive to the Smith's degradation and beta-elimination, suggesting the involvement of O-linked carbohydrates in nonterminal reducing positions. A mucin-type glycosidic linkage was supported because of the lack of A10 reactivity with HT-29 cells grown with phenyl-N-acetyl-alpha-D-galactosaminide. Deglycosylation studies with trifluoromethanesulfonic acid pointed to the involvement of core mucin glycans in the A10 epitope. This epitope was resistant to protease, O- and N-glycanase treatments carried out on trifluoromethanesulfonic acid-deglycosylated mucins. Inhibition studies with core 1, core 2, core 3, and core 6 suggested the latter [GlcNAcbeta(1-6)GalNAc] as being involved in A10 epitope. Taken together, the present results point to A10 defining a core 6-related epitope on core mucin glycans expressed by colon cancer MUC1 not previously associated with human cancer.

Inhibition of in vivo tumor growth by a monoclonal IgM antibody recognizing tumor cell surface carbohydrates.

We have shown previously that IgM from Ehrlich tumor (ET)-immunized mice, recognizing ET cell surface carbohydrates, protects control mice to a subsequent tumor challenge. The factors involved in such IgM-mediated protection were unknown, since it was independent of complement activation. Here, we have extended these in vivo studies by means of monoclonal IgM antibodies. Two of them (A10 and E1), strongly recognizing ET cells and with specificity to ET cell surface carbohydrates, were selected. The results show that A10 (but not E1 or unrelated IgM antibodies) is able to protect nonimmunized mice against ET growth. Protection by A10 was also seen by reducing 800-fold the initial dose; however, E1 was unsuccessful whatever the dose used. A10-mediated protection was observed in C3-defective mice (cobra venom factor treated) or in C5-deficient DBA/2, but not in silica-treated animals. Endotoxin removal did not affect the protection afforded by A10 while specific IgM depletion prevented any protective effect. In addition, the relationship between natural antibodies of IgM isotype recognizing ET cell surface carbohydrates and mouse strain resistance to this tumor is established. Similarly, this natural resistance seems to be complement independent but macrophage mediated. Therefore, these results indicate that some IgM molecules recognizing cell surface carbohydrates may participate in in vivo tumor suppression by a macrophage-dependent mechanism.

Ehrlich tumor stimulates extramedullar hematopoiesis in mice without secreting identifiable colony-stimulating factors and without engagement of host T cells.

Tumor growth is associated with neutrophilia, thrombocytosis, and extramedullar hematopoiesis. The mechanism(s) accounting for these phenomena is unclear, although granulocyte-macrophage colony-stimulating factor (GM-CSF) and/or granulocyte colony-stimulating factor (G-CSF) released by tumor cells have been involved. We studied whether CSF released by Ehrlich tumor (ET) may play a role. A comparative study was performed with two cell variants (ET and ET/0) growing in euthymic, nude, and SCID mice. Extramedullar hematopoiesis was assessed in the spleen by scoring organ enlargement, wheat germ agglutinin ve+ cells, and interleukin 3-dependent granulocyte-macrophage colony-forming unit (GM-CFU). Both cell lines showed the same cytokine profile by reverse transcriptase polymerase chain reaction, including GM-CSF, G-CSF, and macrophage colony-stimulating factor (M-CSF); yet, only ET cells produced detectable colony-stimulating activity in vitro, mainly due to GM-CSF. No differences in tumorigenicity were noted between ET and ET/0 cells inoculated to normal or immunodeficient mice. An increase in extramedullar hematopoiesis, accompanied by neutrophilia and thrombocytosis, was associated with tumor progression irrespective of the cell line. A strong correlation was obtained between the increase in splenic GM-CFU and tumor mass (r = 0.96, p < 0.0001) that was independent on the tumor cell line, strain of mice, or stage of tumor development. The results point against CSF released by tumor cells and/or reactive host T cells as the only factors involved in the extramedullar hematopoiesis in this tumor model. The remarkable correlation between splenic GM-CFU and the tumor mass still suggests that a factor(s) of tumor origin may play a critical role.

A 50-gene signature is a novel scoring system for tumor-infiltrating immune cells with strong correlation with clinical outcome of stage I/II non-small cell lung cancer.

PURPOSE: To identify a novel system for scoring intratumoral immune response that can improve prognosis and therapy decisions in early stage non-small cell lung cancer (NSCLC). METHODS/PATIENTS: Eighty-four completely resected stage I/II NSCLC without adjuvant therapy were classified by expression profiling using whole genome microarrays. An external cohort of 162 tumors was used to validate the results. Immune cells present in tumor microenvironment were evaluated semiquantitatively by CD20, CD79, CD3, CD8, CD4 and CD57 immunostaining. Univariate and multivariate analyses of variables associated with recurrence-free survival were performed. RESULTS: Initial molecular classification identified three clusters, one with significantly better RFS. A reduced two-subgroup classification and a 50-gene predictor were built and validated in an external dataset: high and low risk of recurrence patients (HR = 3.44; p = 0.001). Analysis of the predictor´s genes showed that the vast majority were related to a B/plasma cell immune response overexpressed in the low-risk subgroup. The predictor includes genes coding for unique B lineage-specific genes, functional elements or other genes that, although non-restricted to this lineage, have strong influence on B-cell homeostasis. Immunostains confirmed increased B-cells in the low-risk subgroup. Gene signature (p < 0.0001) and CD20 (p < 0.05) were predictors for RFS, while CD79 and K-RAS mutations showed a tendency. CONCLUSIONS: Favorable prognosis in completely resected NSCLC is determined by a B-cell-mediated immune response. It can be differently scored by a 50-gene expression profile or by CD20 immunostaining. That prognosis information not reflected by traditional classifications may become a new tool for determining individualized adjuvant therapies.

Inhibition of bone marrow-derived natural suppressor activity by glucocorticoids and its reversal by IFN-gamma or IL-2.

Natural suppressor (NS) activity is mediated by cells (NS cells) of bone marrow origin with ability to suppress nonspecifically proliferative responses of lymphocytes. Here we show that pharmacologic concentrations (10(-6)-10(-8) M) of glucocorticoids (GC) greatly inhibit NS activity, as detected by coculturing bone marrow and spleen cells stimulated with B cell (LPS) or T cell (concanavalin A) mitogens. Progesterone antagonizes GC-mediated inhibition of NS activity, suggesting that GC were acting through a receptor-dependent mechanism. A prior treatment of NS cells with GC (10(-5) M) has no effect on the NS activity mediated by these cells. GC are required in culture during the first 24 hr of the suppressor assay. Addition of low amounts of IFN-gamma to GC-treated cultures fully reverses NS cell-mediated suppression. IL-2 produces a reversion as well, while addition of IL-3 or IL-4 does not prevent the GC effect. Neutralizing anti-IFN-gamma antibodies, but not anti-IL-2 or anti-TGF-beta, greatly inhibit NS activity in absence of GC. Taken together, these results indicate that GC inhibit NS activity by impairing endogenous cytokine production required to obtain successful NS cell activation, and not by acting directly on NS cells (i.e., inhibiting the secretion of putative NS factors). Among the cytokines involved in NS cell activation, IFN-gamma appears to be critical, since its addition readily overrides the GC effect and its neutralization results in strong inhibition of NS activity.

Pseudo-allergic reactions to corticosteroids: diagnosis and alternatives.

Two patients treated with parenteral paramethasone (Triniol) and dexamethasone (Sedionbel) are described. A few minutes after administration of the drugs, they presented urticaria (patients 1 and 2) and conjunctivitis (patient 1). The purpose of our study was to determine the cause of the patients' reactions, the immunological mechanisms involved and whether these patients would be able to tolerate any kind of corticoid. Clinical examinations and skin, oral and parenteral challenges with different corticosteroids and ELISA tests were performed. In the two patients, skin and ELISA tests with paramethasone were negative, as was the prick test with each of its excipients. A single-blind parenteral challenge with Triniol was positive in both patients after the administration of 1 ml of the drug, and negative with its excipients. We also carried out oral and parenteral challenges with other corticosteroids and found intolerance to some of them. These results suggest that paramethasone caused pseudoallergic reactions in our patients. Corticosteroids different from paramethasone also produced hypersensitivity reactions in these patients; however, a few of them were tolerated. The basic mechanisms of those reactions are not yet fully understood. To our knowledge, this is the first report of a pseudo-allergy caused by paramethasone.

Severe allergic conjunctivitis and chemosis caused by disodium cromoglycate.

A 63-year-old man with chronic, nonallergic rhinoconjunctivitis presented immediate adverse reactions, such as intense itching, burning, redness and severe swelling of both conjunctivae after using disodium cromoglycate eye drops. Skin prick tests and conjunctival provocation tests were positive with pure disodium cromoglycate. Circulating IgE-specific antibodies to disodium cromoglycate in serum were demonstrated by RAST. We suggest that the acute ocular reaction was caused by disodium cromoglycate and that the underlying mechanism was probably an IgE-mediated immunological reaction.

Sensitization to indigenous pollen and molds and other outdoor and indoor allergens in allergic patients from saudi arabia, United arab emirates, and Sudan.

BACKGROUND: : Airborne allergens vary from one climatic region to another. Therefore, it is important to analyze the environment of the region to select the most prevalent allergens for the diagnosis and treatment of allergic patients. OBJECTIVE: : To evaluate the prevalence of positive skin tests to pollen and fungal allergens collected from local indigenous plants or isolated molds, as well as other outdoor and indoor allergens in allergic patients in 6 different geographical areas in the Kingdom of Saudi Arabia (KSA), the United Arab Emirates, and Sudan. MATERIAL AND METHODS: : Four hundred ninety-two consecutive patients evaluated at different Allergy Clinics (276 women and 256 men; mean age, 30 years) participated in this study. The selection of indigenous allergens was based on research findings in different areas from Riyadh and adjoining areas. Indigenous raw material for pollen grains was collected from the desert near the capital city of Riyadh, KSA. The following plants were included: Chenopodium murale, Salsola imbricata, Rumex vesicarius, Ricinus communis, Artiplex nummularia, Amaranthus viridis, Artemisia monosperma, Plantago boissieri, and Prosopis juliflora. Indigenous molds were isolated from air sampling in Riyadh and grown to obtain the raw material. These included the following: Ulocladium spp., Penicillium spp., Aspergillus fumigatus, Cladosporium spp., and Alternaria spp. The raw material was processed under Good Manufacturing Practices for skin testing. Other commercially available outdoor (grass and tree pollens) and indoor (mites, cockroach, and cat dander) allergens were also tested. RESULTS: : The highest sensitization to indigenous pollens was detected to C. murale (32%) in Khartoum (Sudan) and S. imbricata (30%) and P. juliflora (24%) in the Riyadh region. The highest sensitization to molds was detected in Khartoum, especially to Cladosporium spp. (42%), Aspergillus (40%), and Alternaria spp. (38%). Sensitization to mites was also very prevalent in Khartoum (72%), as well as in Abu Dhabi (United Arab Emirates) (46%) and Jeddah (KSA) (30%). CONCLUSIONS: : The allergenicity of several indigenous pollens and molds derived from autochthonous sources was demonstrated. Prevalence studies in different regions of KSA and neighbor countries indicate different sensitization rates to these and other outdoor and indoor allergens.

[Stimulation of cell division by Ehrlich carcinoma cell surface antibodies]. / Estimulación de la división celular por anticuerpos contra la superficie de las células del carcinoma de Ehrlich.

Some Ehrlich ascites cancer cells of the mouse show nuclear division as a response to cell surface signals. Antibodies to plasma membrane produce a membrane molecular redistribution, but only 25 per 100 proceed to nuclear division. This division is not associated to DNA synthesis, showing that the effect is produced in G0 phase cells originated from G2 phase cells.

IgM response and resistance to ascites tumor growth.

Antibody response and protection against Ehrlich ascites tumor (EAT) was studied in eight EAT-immunized strains of mice (AL/N, BALB/C, C57BL/6J, F1 (C57BL/6 x BALB/C), C57BL/10J, B10.BR, CBA/Ca, SW). The results showed a close association between IgM response and resistance to subsequent tumor challenge. Thus, protection was only achieved in those animals giving a measurable IgM response against EAT cell surface antigens, i.e., all inbred strains of mice tested, except CBA/Ca, and some outbred SW mice. The lack of IgM response to these antigens in CBA/Ca was not linked to the strain H-2 haplotype. Resistance could be passively transferred to nonimmunized mice by means of serum, or purified IgM, from protected immune animals. Moreover, complement depletion by cobra venom factor treatment did not modify the protection afforded to those mice. IgM reactivity to EAT cells was completely abolished by previous cell trypsinization. Trypsin removed but did not destroy the antigen(s) recognized by the IgM, since all its activity could be absorbed with the supernatant of the EAT cell trypsinization. Absorption assays with this supernatant treated with different agents, showed that lipids, simple peptides and nucleic acids were not important components of the antigenic determinants. On the contrary, its susceptibility to beta-galactosidase and particularly to a mild periodate oxidation, suggested that determinants recognized by the IgM against the EAT cell surface are carbohydrate in nature.

Cyclophosphamide induces the development of early myeloid cells suppressing tumor cell growth by a nitric oxide-dependent mechanism.

Adoptive immunotherapy with cyclophosphamide (Cy) increases the host resistance against tumor growth. The precise mechanism(s) by which this therapy enhances tumor suppression is unclear. Cy induces the development of early myeloid cells that may be strongly antiproliferative through NO production. These cells are similar to the natural suppressor cells found in normal bone marrow with a potential antitumor effect. Here we have addressed whether the development of NO-producing cells may be involved in this tumor resistance in Cy-treated mice. The results show a synergism between Cy treatment and tumor-specific lymphocytes transferred systemically (i.v.) or locally (Winn's assay) that results in a strong tumor suppression. Inhibition of NO production by N(G)-monomethyl-L-arginine at the site of tumor inoculation results in a loss of the protection achieved by the combined therapy. Cy-treated mice develop splenic early myeloid (CD11b, Gr-1, CD31 (ER-MP12), ER-MP20, ER-MP54) cells producing large amounts of NO upon T cell-derived signals (IFN-gamma plus CD40 ligation) able to inhibit tumor cell growth in vitro. Early myeloid cells (ER-MP54(+)) and cells expressing inducible NO synthase are increased at the site of tumor challenge in mice treated with the combined therapy, but not in those treated with Cy or immune cell transfer alone. Thus, Cy induces the expansion of early myeloid cells, inhibiting tumor cell growth by a mechanism involving NO. Both the recruitment and the activation of these myeloid cells at the site of tumor challenge appear to be dependent on the presence of tumor-specific lymphocytes.

Bakers' asthma caused by alpha amylase.

Two bakers with bronchial asthma and two with rhinoconjunctivitis are described. Prick and RAST tests were positive with wheat flour in all of them, but the challenge test (nasal or bronchial) with wheat flour extract was positive only in one asthmatic baker. The prick test, RAST, and nasal or bronchial challenge done with alpha amylase extract (a glycolytic enzyme obtained from Aspergillus oryzae and used as a flour additive) were positive in all four patients. Our results support previous data indicating that alpha amylase used in bakeries is an important antigen that could cause respiratory allergy in bakers. It can function as sole causative allergen or in addition with other allergens used in the baking industry.

Prediction of annual variations in atmospheric concentrations of grass pollen. A method based on meteorological factors and grain crop estimates.

We performed an aerobiologic observation of the grasses present in Madrid for 14 years (1978-1991), using volumetric air samplers. The counts obtained show that the major grass pollen release period (average daily grass pollen counts greater than 50 grains/m3 of air) occurs in the months of May and June, although lower counts can occur some days from the end of January onward. There are wide year-to-year variations in total atmospheric grass pollen counts, expressed as the total sum of the mean daily concentrations from April 1st to July 30th (ranging from 2568 to 6624). A strong, statistically significant correlation, based on Spearman's rank test and/or simple and multiple linear regressions, was found between the total grass seasonal count and preseasonal rainfall from October to March (R2 = 0.64; P = 0.0429). The meteorological variable which gave the correlation with greatest statistical significance (R2 = 0.97; P = 0.0016) was the average monthly preseasonal humidity from October to March. A good correlation was also found between March estimates of wheat, rye and barley crops and the total grass count (R2 = 0.73; P = 0.006). A model was designed from the above mentioned humidity variable through a multilinear regression analysis, and it was possible to predict, at the beginning of April, total seasonal counts for 1989 (predicted = 5468; actual = 4410; average error = 24%), 1990 (5033; 6090; -17%) and 1991 (3930; 2568; 53%). These data may help clinicians to predict and prepare themselves for the intensity of the grass pollen season and to explain yearly variations in the severity of symptoms.