Maternal-fetal interaction: preconception immunization in mice prevents neonatal sensitization induced by allergen exposure during pregnancy and breastfeeding

Abstract

Allergen exclusion measures during pregnancy and lactation have been given consideration in studies of primary allergy prevention but complete avoidance of mother/neonatal allergen exposure has proven to be a difficult procedure. To evaluate a strategy to prevent allergen sensitization in early life in mice, we first established a neonatal model with ovalbumin sensitization through maternal allergen exposure during pregnancy or breastfeeding. The modulatory potential of preconception immunization was investigated on the neonatal development of subsequent allergic responses to maternal allergen exposure. Herein, we demonstrate that immunized mothers exposed to antigen during pregnancy or breastfeeding underwent intense vertical transmission of antibodies, including immunoglobulin G (IgG) in complex with ovalbumin and IgG1 antibody with anaphylactic function. It was further shown that maternal immunization efficiently decreased the passage of free antigens through breastfeeding and inhibited the enhanced IgE antibody response after postnatal antigen exposure. In addition, antenatal immunization decreased the antigen-specific proliferative response of immunized neonates, in parallel with profound downmodulatory effects on both the activation and differentiation of T and B cells after a non-specific stimulus and cytokine production. These findings showed that early life sensitization, subsequent to maternal allergen exposure during both the prenatal and postnatal periods, could be avoided by preventive vaccination of the mother.

Figure 1

Schematic representation of maternal immunization and exposure to OVA in prenatal or postnatal period. BALB/c mice were subcutaneously immunized with 200 μg OVA and i.p. boosted with 100 μg OVA after 10 and 20 days of immunization and mated one day after that. Immunized (Im) groups were orally administered with five 300 μg OVA doses during pregnancy (Im + Ag preg) or with three doses of 0·5 or 3·0 mg during the first week of breastfeeding (Im + Ag breast). Groups of non-immunized (n Im) pregnant mice exposed during pregnancy (Ag preg) or during the breastfeeding period (Ag breast) were also studied.

Figure 2

Evaluation of antibody transmission by immune mother exposed to OVA in the prenatal or postnatal period. Female BALB/c mice immunized or not with OVA were mated and orally administered OVA during (a) gestation or (b) the breastfeeding period. Placental serum and amniotic fluid were obtained from Caesarean section from full-term pregnant mice. Breast-milk samples were obtained from the stomach of 5-day-old newborn and offspring serum at 20 and 60 days old. Maternal serum was collected at (a, c) parturition time or (b) 20 days postpartum. Results represent the means of anti-OVA antibody titres ± SD for five or six animals analysed by ELISA and pooled sera from four to six animals for the evaluation of anaphylactic IgG1 antibody by PCA reaction. *P ≤ 0·05 compared to pups from immune mothers.

Figure 3

Determination of antigen and OVA-IgG immune complex transference. Placental serum, milk and pup sera (20-day-old) from immunized or non-immunized mothers exposed to OVA in the prenatal or postnatal period were obtained as described in the Materials and methods. (a) Measurement of OVA in breast milk; (b) detection of OVA–IgG immune complex analysed by ELISA represent OD from a pool of five samples per group (1 : 200). *P ≤ 0·05 compared to non-immune mother; **P ≤ 0·05 compared to mother exposed to antigen postnatally.

Figure 4

Cellular responsiveness of non-immunized offspring. (a) Spleen cells from non-immunized offspring (20-day-old) from immune mothers and/or those exposed to OVA during gestation or breastfeeding periods were cultured with OVA, anti-CD3 mAb or LPS and [³H]thymidine uptake are represented by the stimulation index (SI); (b) cytokine measurement in supernatants of spleen cell culture after 24 hr of stimulation with anti-CD3 mAb by ELISA. Results represent mean ± SD from five to eight mice/per group assayed individually in three different experiments. *P ≤ 0·05 compared to control group.

Figure 5

Evaluation of IgE response in the neonatal and weaning immunization. (a) Three-day-old or 25-day-old offspring of immune mothers and/or those exposed to OVA at prenatal or postnatal period were immunized with OVA and IgE antibody evaluated by means of PCA reaction; (b) Neonatal pups (3-day-old) from non-immunized mothers were injected with 1·0 mg or 0·4 mg of rabbit anti-OVA IgG antibody or control antibody, immunized with OVA and sera were evaluated for IgE by PCA reaction. *P ≤ 0·05 compared to non-immunized mother.

Figure 6

Effect of antenatal immunization with subsequent antigen exposure at postnatal period in the cellular responsiveness of immunized neonates. (a) Spleen cells of offspring immunized at the neonatal period born to immunized or non-immunized mothers exposed to OVA at postnatal period were cultured with OVA, anti-CD3 mAb or LPS and [³H]thymidine uptake are represented by the stimulation index (SI); (b) cytokine levels were evaluated in supernatants of spleen cell culture stimulated with anti-CD3 mAb for 24 hr by ELISA. Results represent mean ± SD from five to eight mice/per group assayed individually in three different experiments. *P ≤ 0·05 compared to pups of non-immunized mothers.