Immunological and Metabolomic Impacts of Administration of Cry1Ab Protein and MON 810 Maize in Mouse

We have investigated the immunological and metabolomic impacts of Cry1Ab administration to mice, either as a purified protein or as the Cry1Ab-expressing genetically modified (GM) MON810 maize. Humoral and cellular specific immune responses induced in BALB/cJ mice after intra-gastric (i.g.) or intra-peritoneal (i.p.) administration of purified Cry1Ab were analyzed and compared with those induced by proteins of various immunogenic and allergic potencies. Possible unintended effects of the genetic modification on the pattern of expression of maize natural allergens were studied using IgE-immunoblot and sera from maize-allergic patients. Mice were experimentally sensitized (i.g. or i.p. route) with protein extracts from GM or non-GM maize, and then anti-maize proteins and anti-Cry1Ab–induced immune responses were analyzed. In parallel, longitudinal metabolomic studies were performed on the urine of mice treated via the i.g. route. Weak immune responses were observed after i.g. administration of the different proteins. Using the i.p. route, a clear Th2 response was observed with the known allergenic proteins, whereas a mixed Th1/Th2 immune response was observed with immunogenic protein not known to be allergenic and with Cry1Ab. This then reflects protein immunogenicity in the BALB/c Th2-biased mouse strain rather than allergenicity. No difference in natural maize allergen profiles was evidenced between MON810 and its non-GM comparator. Immune responses against maize proteins were quantitatively equivalent in mice treated with MON810 vs the non-GM counterpart and no anti-Cry1Ab–specific immune response was detected in mice that received MON810. Metabolomic studies showed a slight “cultivar” effect, which represented less than 1% of the initial metabolic information. Our results confirm the immunogenicity of purified Cry1Ab without evidence of allergenic potential. Immunological and metabolomic studies revealed slight differences in mouse metabolic profiles after i.g. administration of MON810 vs its non-GM counterpart, but no significant unintended effect of the genetic modification on immune responses was seen

Interleukin-10 Mediated Autoregulation of Murine B-1 B-Cells and Its Role in Borrelia hermsii Infection

B cells are typically characterized as positive regulators of the immune response, primarily by producing antibodies. However, recent studies indicate that various subsets of B cells can perform regulatory functions mainly through IL-10 secretion. Here we discovered that peritoneal B-1 (B-1P) cells produce high levels of IL-10 upon stimulation with several Toll-like receptor (TLR) ligands. High levels of IL-10 suppressed B-1P cell proliferation and differentiation response to all TLR ligands studied in an autocrine manner in vitro and in vivo. IL-10 that accumulated in cultures inhibited B-1P cells at second and subsequent cell divisions mainly at the G1/S interphase. IL-10 inhibits TLR induced B-1P cell activation by blocking the classical NF-κB pathway. Co-stimulation with CD40 or BAFF abrogated the IL-10 inhibitory effect on B-1P cells during TLR stimulation. Finally, B-1P cells adoptively transferred from the peritoneal cavity of IL-10−/− mice showed better clearance of Borrelia hermsii than wild-type B-1P cells. This study described a novel autoregulatory property of B-1P cells mediated by B-1P cell derived IL-10, which may affect the function of B-1P cells in infection and autoimmunity

Effect of different sensitizing doses of antigen in a murine model of atopic asthma

The dose of antigen is assumed to be one of the important factors in the polarized development of helper T cell subsets, i.e. Th1 or Th2 cells. We investigated the effect of the sensitizing antigen dose in a murine model of atopic asthma, which involved sensitization with ovalbumin (OVA) followed by repeated exposure to OVA aerosols. BALB/c mice were primed with varying doses of OVA (0, 10, 100 and 1000 μg) plus Al(OH)3 on days 0, 7 and 14, and were challenged with OVA aerosols (50 mg/ml for 20 min) on days 15–20. There were striking antigen dose-related differences in OVA-specific antibodies: high IgE and low IgG2a titres were found in mice sensitized at 10 μg, while low IgE and high IgG2a titres were seen at 1000 μg. The sensitizing dose was inversely correlated with the total cell count and the eosinophil count in bronchoalveolar lavage fluid (BALF), as well as with the extent of histological changes such as goblet cell hyperplasia of the bronchial epithelium and cellular infiltration into bronchovascular bundles. Antigen-induced bronchial hyper-responsiveness (BHR) to methacholine was observed with sensitization at 10 μg but not at 1000 μg. Splenic mononuclear cells (SMNC) obtained from mice sensitized at either dose showed proliferation in response to OVA. Production of IL-4 and IL-5 by OVA-stimulated SMNC was inversely correlated with the dose of sensitizing antigen. High-dose sensitization resulted in general suppression of cytokine production by SMNC, including interferon-gamma (IFN-γ). The BALF levels of IL-4 and IL-5 were increased by low-dose sensitization, whereas IFN-γ and IL-12 levels were increased by high-dose sensitization. These results suggest that the dose of sensitizing antigen defines the phenotypic changes in the present murine asthma model, presumably by influencing the pattern of cytokine production