Evidence that FcRn mediates the transplacental passage of maternal IgE in the form of IgG anti-IgE/IgE immune complexes

Abstract

Background: The mechanism(s) responsible for acquisition of maternal antibody isotypes other than IgG are not fully understood. This uncertainty is a major reason underlying the continued controversy regarding whether cord blood (CB) IgE originates in the mother or fetus.

Objective: To investigate the capacity of maternal IgE to be transported across the placenta in the form of IgG anti-IgE/IgE immune complexes (ICs) and to determine the role of the neonatal Fc receptor (FcRn) in mediating this process.

Methods: Maternal and CB serum concentrations of IgE, IgG anti-IgE, and IgG anti-IgE/IgE ICs were determined in a cohort of allergic and non-allergic mother/infant dyads. Madin-Darby canine kidney (MDCK) cells stably transfected with human FcRn were used to study the binding and transcytosis of IgE in the form of IgG anti-IgE/IgE ICs.

Results: Maternal and CB serum concentrations of IgG anti-IgE/IgE ICs were highly correlated, regardless of maternal allergic status. IgG anti-IgE/IgE ICs generated in vitro bound strongly to FcRn-expressing MDCK cells and were transcytosed in an FcRn-dependent manner. Conversely, monomeric IgE did not bind to FcRn and was not transcytosed. IgE was detected in solutions of transcytosed IgG anti-IgE/IgE ICs, even though essentially all the IgE remained in complex form. Similarly, the majority of IgE in CB sera was found to be complexed to IgG.

Conclusions and clinical relevance: These data indicate that human FcRn facilitates the transepithelial transport of IgE in the form of IgG anti-IgE/IgE ICs. They also strongly suggest that the majority of IgE in CB sera is the result of FcRn-mediated transcytosis of maternal-derived IgG anti-IgE/IgE ICs. These findings challenge the widespread perception that maternal IgE does not cross the placenta. Measuring maternal or CB levels of IgG anti-IgE/IgE ICs may be a more accurate predictor of allergic risk.

Keywords: FcRn; IgG anti-IgE; allergy; autoantibodies; cord blood; immune complexes; infant; mother; placenta.

Figure 1

IgG anti‐IgE autoantibodies in maternal and CB sera. Data represent results obtained from 59 allergic and 85 non‐allergic mother/infant dyads. (a) Levels of IgG anti‐IgE were similar between allergic and non‐allergic pregnant women. Total serum IgE correlated with levels of IgG anti‐IgE in (b) allergic pregnant women but not in (c) non‐allergic pregnant women. (d) A combined analysis of allergic and non‐allergic mother/infant dyads demonstrated that maternal and CB levels of IgG anti‐IgE were highly correlated. ns, not significant. The horizontal line represents the median.

Figure 2

IgG anti‐IgE/IgE ICs in maternal and CB sera. Data represent results obtained from 59 allergic and 85 non‐allergic mother/infant dyads. (a) Levels of IgG anti‐IgE/IgE ICs were similar between allergic and non‐allergic pregnant women, when determined using the m107, HP6029, or rFcεRIα IC assays. Total serum IgE correlated with levels of IgG anti‐IgE/IgE ICs in (b) allergic pregnant women but not in (c) non‐allergic pregnant women, when ICs were measured using rFcεRIα. (d–f) A combined analysis of allergic and non‐allergic mother/infant dyads demonstrated that maternal and CB levels of IgG anti‐IgE/IgE ICs were highly correlated when determined using the m107, HP6029, or rFcεRIα IC assays. ns, not significant.

Figure 3

IgG subclass distribution of IgG anti‐IgE/IgE ICs in maternal and CB sera. Data represent results obtained from 10 allergic and 11 non‐allergic pregnant women with similar levels of total IgG anti‐IgE/IgE ICs, and their infants. (a) Total IgE levels were significantly greater in allergic as compared to non‐allergic pregnant women. (b) Levels of IgG₁‐containing ICs were significantly greater in allergic as compared to non‐allergic pregnant women. In corresponding infants, there were no significant differences in levels of CB IgE (c) or IgG subclass‐specific ICs (d) based on maternal history of allergy. (e) In allergic mother/infant dyads, combined levels of maternal IgG₁‐ and IgG₃‐containing ICs correlated significantly with levels of CB IgE. ns, not significant. ***P < 0.001, *P < 0.05, ns⁺ P = 0.11, ns⁺⁺ P = 0.15, ns^# P = 0.18, ns^## P = 0.30.

Figure 4

hFcRn binds IgE in the form of IgG anti‐IgE/IgE ICs. hFcRn‐expressing MDCK cells were incubated with (a) biotin‐conjugated mIgG anti‐hIgE_{C ε4}, (b) hIgE, (c) rIgG anti‐hIgE, or (d) rIgG anti‐hIgE/hIgE ICs, at (e) different pHs, or (f) in the presence of the hFcRn‐blocking antibody DVN24. Biotin‐conjugated mIgG anti‐rFcγ was used to detect bound rIgG anti‐hIgE, and biotin‐conjugated mIgG anti‐hIgE_{C ε4} was used to detect bound hIgE or rIgG anti‐hIgE/hIgE ICs. Visualization of bound antibodies was with SA‐APC, and the cells were analysed by flow cytometry. (g) Immunofluorescence microscopy of MDCK cells performed at pH 6.0 demonstrated co‐localization of hFcRn (green) and hIgE (in the form of rIgG anti‐hIgE/hIgE^{AF 555} ICs) (red). The blue colour represents nuclear staining by DAPI.

Figure 5

hFcRn facilitates the transcytosis of hIgE in the form of IgG anti‐IgE/IgE ICs. hFcRn‐expressing MDCK cells were plated on semipermeable Transwell filters and grown to confluence. Transcytosis assays were performed with the input chamber buffered to pH 6.0 and the output chamber buffered to 7.4. (a) rIgG anti‐hIgE and rIgG anti‐hIgE/hIgE ICs were transcytosed across MDCK cells, whereas monomeric IgE was not. Transcytosis of rIgG anti‐hIgE and rIgG anti‐hIgE/hIgE ICs was inhibited by DVN24 and RhoGAM. (b) Increasing concentrations of DVN24 added to the input chambers resulted in greater inhibition in the transport of rIgG anti‐hIgE/hIgE ICs. (c) hIgE detected in pooled solutions of transcytosed rIgG anti‐hIgE/hIgE ICs was significantly reduced after selective removal of rIgG (and rIgG anti‐hIgE/hIgE ICs) using protein A, indicating the majority of the hIgE remained in complex form. Transport data are representative of 4 individual experiments. ***P < 0.0001.

Figure 6

The majority of IgE in CB sera is bound by IgG. CB sera obtained from 20 different infants were pooled into five separate groups of 4. Symbols represent total IgE levels obtained for each group, before and after selective absorption of IgG (and IgG anti‐IgE/IgE ICs) using protein A chromatography. Total IgE in CB sera was significantly reduced after protein A exposure indicating the majority of IgE was bound by IgG. *P < 0.05. The horizontal line represents the mean.

Figure 7

CB IgE is predictive of basophil‐bound IgE in infants of allergic mothers, but not in infants of non‐allergic mothers. Data represent CB samples from 7 infants of allergic mothers and 11 infants of non‐allergic mothers, containing ≥ 0.5 kU/L total serum IgE (determined by ImmunoCAP) and a minimum of 200 basophils. Flow cytometry was used to calculate the percentages of CB basophils with surface‐bound IgE relative to an isotype control.