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Influence Of H1, H2 And H3 Antagonists On Apoptosis Of Lymphoid Cells
S178 Abstracts
697
Changes in Cytokine Levels in The First Year of Life in WHEALS Cohort Infants K. R. Bobbitt, K. J. Woodcroft, S. Havstad, L. K. Williams, C. C. Johnson; Henry Ford Hospital, Detroit, MI. RATIONALE: The immune response to pathogens is predominated by either a Th1 or Th2 response. Each is characterized by a specific cytokine profile, Th1 with IL-2, IL-12 and IFN-g, Th2 with IL-4, IL-5, and IL-13. Deregulation of the Th2 response has been reported in atopic individuals and characterized by an increase in Th2 associated cytokines. We examined a sample of whole blood specimens obtained from infants in a Detroit area population-based prospective birth cohort (WHEALS) to identify cytokine profiles in the first two years of life. METHODS: Whole blood specimens were obtained from infants at birth, 6 months, and 1 year. Immune cells were stimulated and detection of IL-4, IL-10, IL-12, and IFN-g was performed using the Flex Set Cytometric Bead Arrays (CBA) with flow cytometry. RESULTS: Low concentrations of both IL-4 and IL-12 cytokines were observed. In contrast, IL-10 and IFN-g levels were increased in the six month samples relative to cord blood. This was generally followed by a decrease in the one year samples. CONCLUSIONS: Only IL-10 and IFN-g were repeatedly elevated in the six month samples. However, concentrations of each cytokine decreased in the 1 year samples. Although the trends observed were similar for all infants the relative changes were different. Future investigation into the role played by the degree of IFN-g and IL-10 synthesis may provide a useful tool in predicting risk for later development of atopy. Funding: NIAID
698
MONDAY
Effect Of Nasal And Bronchial Grass Pollen Challenges On Plasma Cytokine Levels C. Barnig1, B. Uring-Lambert2, A. Purohit1, C. Donnay1, A. Casset1, S. Bahram2, F. de Blay1; 1Asthma and Allergy Division, Department of Chest Diseases, University Hospital, Strasbourg, FRANCE, 2Laboratory of Immunology, University Hospital, Strasbourg, FRANCE. RATIONALE: Few studies have performed nasal and bronchial allergen challenges in the same patients. The aim of our study was to assess the effect of the allergen delivery route (nasal and bronchial) on plasma cytokine levels in grass pollen sensitized patients. METHODS: Twenty-one subjects with seasonal grass pollen rhinitis and/ or intermittent asthma underwent outside the grass pollen season one nasal provocation test (NPT) and one bronchial provocation test (BPT) with a grass pollen extract in a randomized order. BPT, performed by using a dosimeter jet nebulizer (Spira Electro 2) producing 10 mm particles, was considered positive when FEV 1 dropped by 20%. NPT, performed by means of a nasal pump spray in both nostrils, was considered positive when nasal flow measured by anterior-rhinometry dropped by 40% or symptom score was 5. Plasma cytokine levels (IL-4, IL-5, IL-10, IL-2, TNF-a and IFN-g) were measured before and 3, 6 and 24h after allergen challenge by flow cytometry using a bead array immunoassay. RESULTS: A significant increase in plasma levels of IL-5 was found 24 hours following NPT (0.78 to 2.50 pg/ml; p50.008) and 6 and 24 hours following BPT (0.95 to 40.69 pg/ml; p50.029). The increase of IL-5 after BPT was significantly higher than after NPT (p50.034). IL-4, IL-10, IL-2, TNFa and IFN-g plasma levels were undetectable in most of the samples. CONCLUSIONS: Our results suggest that introducing allergens through the bronchi induces significantly higher concentrations of IL-5 than through the nose. Funding: Schering-Plough
699
Influence Of H1, H2 And H3 Antagonists On Apoptosis Of Lymphoid Cells I. Kaidashev1, I. Yastremskaya1, N. Kutzenko1, L. M. DuBuske2; 1Ukrainian Medical Stomatological Academy, Poltava, UKRAINE, 2Immunology Research Institute of New England, Gardner, MA. RATIONALE: Histamine induced responses mediated by H1 and H2 receptors may modulate immune responses to antigens and regulate
J ALLERGY CLIN IMMUNOL JANUARY 2007
development of tolerance versus inflammation. H3/H4 receptors may also regulate inflammatory cell activity. This study investigates the influence of the H1antagonist desloratadine(D), the H2 antagonist famotidine(F) and the H3 antagonist thioperamide(T) on apoptosis of rat lymphoid cells. METHODS: D was administered to 40 Wistar rats in doses of 0.007, 0.07 and 0.7 mg/kg for 10 days. F was administered to 40 Wistar rats in doses of 0.06, 0.6 and 6 mg/kg for 10 days. T was given as an intraperitoneal injection in the doses 10 mg/kg to a similar number of rats. 10 untreated rats comprised the control group. Animals were sacrificed by cervical dislocation and suspensions of lymphoid cells from thymus, lymph nodes, spleen and peripheral blood were obtained. The level of apoptosis was estimated by flow cytometry with annexin V-FITC and propidium iodide (PI) staining and morphologically by MGG staining. Results were statistically calculated using Wilcoxon-Mann-Whitney criteria. RESULTS: D inhibited apoptosis of all lymphoid cells investigated including thymocytes, spleenocytes, lymph node cells and peripheral blood mononuclear cells. F stimulated apoptosis in all doses investigated in all cell types. This stimulation was in a dose-dependent manner, the largest doses increasing the percentage of apoptotic cells more significantly. F stimulated development of both early and late stages of lymphoid cell apoptosis. The mononuclear cells from peripheral blood were the most susceptible to apoptosis stimulated by F. T induced the late stages of lymphoid cell apoptosis, increasing the amount of annexin-V and PI double-positive cells. CONCLUSIONS: The H1 antagonist inhibited the apoptosis of lymphoid cells and the H2 antagonist stimulated the apoptosis of lymphoid cells, while the H3 antagonist induced late stages of apoptosis. Peripheral blood mononuclear cells were the cells most susceptible to these effects. Modulation of lymphoid cell apoptosis by H1, H2 and H3/H4 histamine antagonists may be exploited in future therapeutic applications of these agents as therapeutic agents for immunologic diseases.
700
Leukemoid Reaction in Children: Lymphocytes Flow Cytometry Characterization and Clinical Etiologies A. Beigelman1,2, J. Kapelushnik1, G. Shubinsky1, A. Haim1, A. M. Moser1; 1Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, ISRAEL, 2Washington University School of Medicine, Saint Louis, MO. RATIONALE: Leukemoid reaction (LR) is defined as a white blood cell (WBC) count higher than 503109/L. We aimed to define the etiologies of pediatric LR and to determine the lymphocytes cell surface markers of the lymphoid LR. METHODS: Twenty-one children with LR were prospectively studied. Excluded from the study were children with any previous medical history (including history of prematurity). LR was subdivided to lymphoid or myeloid according to the predominant causative cell lineage. Clinical investigation included blood count and morphology; Mycoplasma pneumoniae (M. pneumoniae), Epstein-Barr Virus (EBV), and Cytomegalovirus serology; bacterial cultures and chest X-ray. Lymphocytes cell surface markers were determined by Flow Cytometry (FC). RESULTS: Fourteen children had lymphoid LR, six had neutrophilic LR, and one had eosinophilic LR. Four children with lymphoid LR had gastroenteritis, three had M. pneumoniae; one child each had: EBV, Cytomegalovirus, Giardia lamblia, E. coli UTI, or Tinea capitis infection. The final diagnosis was obscured in two children. All children with neutrophilic LR had lobar pneumonia. The eosinophilic-LR child had scabies. FC analysis demonstrated expansion of both the B-cells and the T-cell lineage. All but two of the patients in this group also had a profound over-expression of CD101/ CD191 B-cells. One patient with EBV infection demonstrated over-expression of CD4/CD8 double positive and double negative T-lymphoid cells. CONCLUSIONS: Leukemoid reaction represents a benign acute response to an underlying disorder (mostly infectious); M. pneumoniae is a common, previously unreported etiology. FC analysis revealed that lymphoid leukemoid reaction is a combined B-cells and T-cells response, with over-expression of immature B-cells. Funding: Goldman Fund of the Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, Israel.
697
Changes in Cytokine Levels in The First Year of Life in WHEALS Cohort Infants K. R. Bobbitt, K. J. Woodcroft, S. Havstad, L. K. Williams, C. C. Johnson; Henry Ford Hospital, Detroit, MI. RATIONALE: The immune response to pathogens is predominated by either a Th1 or Th2 response. Each is characterized by a specific cytokine profile, Th1 with IL-2, IL-12 and IFN-g, Th2 with IL-4, IL-5, and IL-13. Deregulation of the Th2 response has been reported in atopic individuals and characterized by an increase in Th2 associated cytokines. We examined a sample of whole blood specimens obtained from infants in a Detroit area population-based prospective birth cohort (WHEALS) to identify cytokine profiles in the first two years of life. METHODS: Whole blood specimens were obtained from infants at birth, 6 months, and 1 year. Immune cells were stimulated and detection of IL-4, IL-10, IL-12, and IFN-g was performed using the Flex Set Cytometric Bead Arrays (CBA) with flow cytometry. RESULTS: Low concentrations of both IL-4 and IL-12 cytokines were observed. In contrast, IL-10 and IFN-g levels were increased in the six month samples relative to cord blood. This was generally followed by a decrease in the one year samples. CONCLUSIONS: Only IL-10 and IFN-g were repeatedly elevated in the six month samples. However, concentrations of each cytokine decreased in the 1 year samples. Although the trends observed were similar for all infants the relative changes were different. Future investigation into the role played by the degree of IFN-g and IL-10 synthesis may provide a useful tool in predicting risk for later development of atopy. Funding: NIAID
698
MONDAY
Effect Of Nasal And Bronchial Grass Pollen Challenges On Plasma Cytokine Levels C. Barnig1, B. Uring-Lambert2, A. Purohit1, C. Donnay1, A. Casset1, S. Bahram2, F. de Blay1; 1Asthma and Allergy Division, Department of Chest Diseases, University Hospital, Strasbourg, FRANCE, 2Laboratory of Immunology, University Hospital, Strasbourg, FRANCE. RATIONALE: Few studies have performed nasal and bronchial allergen challenges in the same patients. The aim of our study was to assess the effect of the allergen delivery route (nasal and bronchial) on plasma cytokine levels in grass pollen sensitized patients. METHODS: Twenty-one subjects with seasonal grass pollen rhinitis and/ or intermittent asthma underwent outside the grass pollen season one nasal provocation test (NPT) and one bronchial provocation test (BPT) with a grass pollen extract in a randomized order. BPT, performed by using a dosimeter jet nebulizer (Spira Electro 2) producing 10 mm particles, was considered positive when FEV 1 dropped by 20%. NPT, performed by means of a nasal pump spray in both nostrils, was considered positive when nasal flow measured by anterior-rhinometry dropped by 40% or symptom score was 5. Plasma cytokine levels (IL-4, IL-5, IL-10, IL-2, TNF-a and IFN-g) were measured before and 3, 6 and 24h after allergen challenge by flow cytometry using a bead array immunoassay. RESULTS: A significant increase in plasma levels of IL-5 was found 24 hours following NPT (0.78 to 2.50 pg/ml; p50.008) and 6 and 24 hours following BPT (0.95 to 40.69 pg/ml; p50.029). The increase of IL-5 after BPT was significantly higher than after NPT (p50.034). IL-4, IL-10, IL-2, TNFa and IFN-g plasma levels were undetectable in most of the samples. CONCLUSIONS: Our results suggest that introducing allergens through the bronchi induces significantly higher concentrations of IL-5 than through the nose. Funding: Schering-Plough
699
Influence Of H1, H2 And H3 Antagonists On Apoptosis Of Lymphoid Cells I. Kaidashev1, I. Yastremskaya1, N. Kutzenko1, L. M. DuBuske2; 1Ukrainian Medical Stomatological Academy, Poltava, UKRAINE, 2Immunology Research Institute of New England, Gardner, MA. RATIONALE: Histamine induced responses mediated by H1 and H2 receptors may modulate immune responses to antigens and regulate
J ALLERGY CLIN IMMUNOL JANUARY 2007
development of tolerance versus inflammation. H3/H4 receptors may also regulate inflammatory cell activity. This study investigates the influence of the H1antagonist desloratadine(D), the H2 antagonist famotidine(F) and the H3 antagonist thioperamide(T) on apoptosis of rat lymphoid cells. METHODS: D was administered to 40 Wistar rats in doses of 0.007, 0.07 and 0.7 mg/kg for 10 days. F was administered to 40 Wistar rats in doses of 0.06, 0.6 and 6 mg/kg for 10 days. T was given as an intraperitoneal injection in the doses 10 mg/kg to a similar number of rats. 10 untreated rats comprised the control group. Animals were sacrificed by cervical dislocation and suspensions of lymphoid cells from thymus, lymph nodes, spleen and peripheral blood were obtained. The level of apoptosis was estimated by flow cytometry with annexin V-FITC and propidium iodide (PI) staining and morphologically by MGG staining. Results were statistically calculated using Wilcoxon-Mann-Whitney criteria. RESULTS: D inhibited apoptosis of all lymphoid cells investigated including thymocytes, spleenocytes, lymph node cells and peripheral blood mononuclear cells. F stimulated apoptosis in all doses investigated in all cell types. This stimulation was in a dose-dependent manner, the largest doses increasing the percentage of apoptotic cells more significantly. F stimulated development of both early and late stages of lymphoid cell apoptosis. The mononuclear cells from peripheral blood were the most susceptible to apoptosis stimulated by F. T induced the late stages of lymphoid cell apoptosis, increasing the amount of annexin-V and PI double-positive cells. CONCLUSIONS: The H1 antagonist inhibited the apoptosis of lymphoid cells and the H2 antagonist stimulated the apoptosis of lymphoid cells, while the H3 antagonist induced late stages of apoptosis. Peripheral blood mononuclear cells were the cells most susceptible to these effects. Modulation of lymphoid cell apoptosis by H1, H2 and H3/H4 histamine antagonists may be exploited in future therapeutic applications of these agents as therapeutic agents for immunologic diseases.
700
Leukemoid Reaction in Children: Lymphocytes Flow Cytometry Characterization and Clinical Etiologies A. Beigelman1,2, J. Kapelushnik1, G. Shubinsky1, A. Haim1, A. M. Moser1; 1Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, ISRAEL, 2Washington University School of Medicine, Saint Louis, MO. RATIONALE: Leukemoid reaction (LR) is defined as a white blood cell (WBC) count higher than 503109/L. We aimed to define the etiologies of pediatric LR and to determine the lymphocytes cell surface markers of the lymphoid LR. METHODS: Twenty-one children with LR were prospectively studied. Excluded from the study were children with any previous medical history (including history of prematurity). LR was subdivided to lymphoid or myeloid according to the predominant causative cell lineage. Clinical investigation included blood count and morphology; Mycoplasma pneumoniae (M. pneumoniae), Epstein-Barr Virus (EBV), and Cytomegalovirus serology; bacterial cultures and chest X-ray. Lymphocytes cell surface markers were determined by Flow Cytometry (FC). RESULTS: Fourteen children had lymphoid LR, six had neutrophilic LR, and one had eosinophilic LR. Four children with lymphoid LR had gastroenteritis, three had M. pneumoniae; one child each had: EBV, Cytomegalovirus, Giardia lamblia, E. coli UTI, or Tinea capitis infection. The final diagnosis was obscured in two children. All children with neutrophilic LR had lobar pneumonia. The eosinophilic-LR child had scabies. FC analysis demonstrated expansion of both the B-cells and the T-cell lineage. All but two of the patients in this group also had a profound over-expression of CD101/ CD191 B-cells. One patient with EBV infection demonstrated over-expression of CD4/CD8 double positive and double negative T-lymphoid cells. CONCLUSIONS: Leukemoid reaction represents a benign acute response to an underlying disorder (mostly infectious); M. pneumoniae is a common, previously unreported etiology. FC analysis revealed that lymphoid leukemoid reaction is a combined B-cells and T-cells response, with over-expression of immature B-cells. Funding: Goldman Fund of the Faculty of Health Sciences, Ben-Gurion University of the Negev, Beer-Sheva, Israel.