Bet-specific iTreg ameliorated allergic symptoms in humanized
mouse models of allergen-induced airway and intestinal inflammation
Humanized mice are a remarkable investigative tool and preclinical study
system, which close the gap between exclusively murine and human
studies.49 Here, we used well-established and
standardized humanized mouse models of type I allergy to analyze the
function of iTregBet on allergic symptoms in
allergen-induced intestinal and airway
inflammation.41,50,51 An overview of the
reconstitution and challenge protocol is visualized in Figure 5A (see
also Methods and Supplementary Methods). Briefly, immunodeficient mice
(NOD.CB17-Prkdcscid/Jγc-/-) were
engrafted with human PBMC from birch pollen allergic donors with
associated hazelnut allergy and injected with birch pollen extract +/-
iTregBet as indicated.41,52 After
allergen-specific (birch) rectal challenge the intestinal inflammation
was evaluated by a clinical score, that revealed a significantly
increased inflammatory reaction of animals serving as allergic positive
controls (PBMC + birch) compared to negative controls (PBMC), thereby
demonstrating the validity of the humanized model and the development of
an allergic immune reaction (Figure 5B, C). Intriguingly, injection of
iTregBet resulted in a significant abrogation of the
allergen-specific immune reaction as shown by a reduced intestinal
inflammation (Figure 5B, C).
For induction of allergic asthma, the animals were engrafted and boosted
as described above and were challenged intranasally with birch pollen
extract as published previously.40,51 Subsequently,
the airway resistance was assessed as outcome of the allergic immune
reaction.40,51 In these experiments, allergic positive
controls (PBMC + birch) showed a significantly enhanced airway
hyperreactivity in contrast to negative control groups (PBMC) (Figure
5D). Importantly, co-injection of iTregBet curtailed the
development of allergic asthma symptoms, as the airway resistance was
significantly decreased compared to allergic asthma positive control
animals (Figure 5D).
We also analyzed the serum concentrations of human birch-specific IgE
(Figure 5E) as typical immunological parameter of type I allergic
reactions prior to allergen challenge. Compared to enhanced amounts of
birch-specific IgE in allergic positive controls (PMBC + birch),
treatment with iTregBet significantly reduced
birch-specific IgE concentrations in vivo , confirming our data of
significantly reduced clinical symptoms of allergen-induced intestinal
and airway inflammation after iTregBet application.
In order to investigate the induction of cross-reactive tolerancein vivo , the immunodeficient mice were engrafted and boosted as
described above, but were challenged rectally with hazelnut extract
prior to assessment of the intestinal inflammation (Figure 6). In these
experiments, we found a less severe allergic immune response in the
hazelnut-challenged compared to the birch-challenged control group
(Figure 5C), likely due to lack of in vivo booster with hazelnut
extract and/or to the donors’ less severe sensitization towards the food
allergen. However, even after hazelnut challenge co-injection of
iTregBet resulted in a pronounced inhibition of
intestinal allergic symptoms compared to control animals, which was
shown with data of individual experiments (Figure 6A). In addition, we
observed reduced human hazelnut-specific IgE levels after
iTregBet application and hazelnut challenge compared to
the allergic positive control (Figure 6B). These data indicate the
induction of allergen-specific (birch) as well as cross-reactive
tolerance (hazelnut) in vivo through IL-10 DC-induced
iTregBet.