Inflammation research : official journal of the European Histamine Research Society ... [et al.] · 2025
Antigen-specific reduced IgG inhibits cellular response in rat basophilic leukemia cells activated with multivalent antigen.
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Iwase Akemi, Suzuki Ruriko, Yokawa Satoru, Furuno Tadahide
Аннотация
OBJECTIVE: The aim of this study was to investigate whether antigen-specific reduced IgG inhibits mast cell activation evoked by the aggregation of the high-affinity IgE receptor (FcεRI) in a manner similar to intact IgG.
MATERIALS: Rat basophilic leukemia (RBL-2H3) cells are used for a mast cell model.
TREATMENT: Monovalent mouse anti-trinitrophenyl (TNP) IgG1 (75 kDa) was prepared by reducing the disulfide bond between the heavy chains using cysteamine hydrochloride. IgE-sensitized RBL-2H3 cells were stimulated with TNP-OVA, and reduced IgG1 was added 5 min after stimulation.
METHODS: Degranulation and IL-4 secretion were measured 30 min and 3 h after TNP-OVA stimulation by β-hexosaminidase assay and ELISA, respectively. The intracellular distribution of SH2-containing inositol 5'-phosphatase 1 (SHIP1) was determined using immunostaining. Group differences were analyzed using the Tukey-Kramer test.
RESULTS: Reduced IgG1 significantly inhibited degranulation and IL-4 secretion in antigen-stimulated RBL-2H3 cells to an extent similar to intact IgG1. Intracellular SHIP1 localized to the plasma membrane 5 min after the addition of reduced IgG1, mirroring the effect of intact IgG1.
CONCLUSIONS: These results suggest that antigen-specific reduced IgG1 (monovalent) inhibits antigen-induced mast cell activation by activating SHIP1 through co-ligation of FcεRI and the low-affinity IgG receptor (FcγRIIB).
MATERIALS: Rat basophilic leukemia (RBL-2H3) cells are used for a mast cell model.
TREATMENT: Monovalent mouse anti-trinitrophenyl (TNP) IgG1 (75 kDa) was prepared by reducing the disulfide bond between the heavy chains using cysteamine hydrochloride. IgE-sensitized RBL-2H3 cells were stimulated with TNP-OVA, and reduced IgG1 was added 5 min after stimulation.
METHODS: Degranulation and IL-4 secretion were measured 30 min and 3 h after TNP-OVA stimulation by β-hexosaminidase assay and ELISA, respectively. The intracellular distribution of SH2-containing inositol 5'-phosphatase 1 (SHIP1) was determined using immunostaining. Group differences were analyzed using the Tukey-Kramer test.
RESULTS: Reduced IgG1 significantly inhibited degranulation and IL-4 secretion in antigen-stimulated RBL-2H3 cells to an extent similar to intact IgG1. Intracellular SHIP1 localized to the plasma membrane 5 min after the addition of reduced IgG1, mirroring the effect of intact IgG1.
CONCLUSIONS: These results suggest that antigen-specific reduced IgG1 (monovalent) inhibits antigen-induced mast cell activation by activating SHIP1 through co-ligation of FcεRI and the low-affinity IgG receptor (FcγRIIB).