IFN-γ neutralization in AKR/J mice does not enhance basophil
Notch2 expression.
As AKR/J mice have a predominant IFN-γ response following T.
muris infection (Fig. 1C-E) (9-13, 15), we hypothesized that this Type
1 skew might suppress or prevent the upregulation of Notch2 on AKR/J
basophils. To test this idea, we treated AKR/J mice with a neutralizing
α-IFN-γ antibody throughout T. muris infection as described
previously (15, 31) and then examined the levels of Notch2 on cecum
basophils on day 14 p.i., the timepoint at which we see maximal basophil
Notch2 expression in the infected tissue site in C57BL/6 mice (7) (Fig.
4A). We first validated our neutralization protocol, observing an
infection-induced increase in IFN-γ protein levels in isotype
control-treated AKR/J mice but not α-IFN-γ-treated mice in tissue and
blood (Fig. 4B-C) and transcript levels in the colon at d14 p.i. (Fig.
4D). We did not observe an accompanying increase in transcription levels
of Il4 or Il13 in the colon in AKR/J mice at day 14 p.i.,
though this might be due to the early time point examined. As expected,
day 14 p.i. C57BL/6 mice had higher Il4 and Il13 levels in
the colon than naïve C57BL/6 mice, though these data were not
statistically significant (Fig. S3A-B).
At day 14 p.i., we did not observe an increase in cecum basophil Notch2
expression by frequency or gMFI in α-IFN-γ-treated AKR/J mice compared
to what was observed in infected isotype-treated or naïve AKR/J mice. As
previously reported, we observed that cecum basophils in C57BL/6 mice
had elevated Notch2 expression on d14 p.i with T. muris compared
to levels in naïve C57BL/6 mice (7) (Fig. 4E-G). These findings show
that AKR/J cecum basophils do not upregulate Notch2 expression at day 14
p.i. with T. muris when IFN-γ is neutralized. Collectively, these
studies suggest that IFN-γ levels do not control cecum basophil Notch2
expression in AKR/J mice in response to T. muris infection, at
least at the day 14 p.i. timepoint.