Discussion
H. polygyrus infection reduces RSV infection peak viral load,
secondary immuno-pathology, and associated lung function
impairment6. We now describe how this helminth
infection induces enhanced monopoiesis in the bone marrow, elevated
circulatory monocytes and increased recruitment of monocyte derived
mononuclear phagocytes to the lung. Using monocyte ablation and transfer
we show the functional importance of circulatory monocyte derived cells
in the H. polygyrus -induced anti-viral effect against RSV.
Elevated numbers of circulatory leukocytes over the first two weeks ofH. polygyrus infection, including lymphocytes, neutrophils and
monocytes have previously been described but only in C57BL/6
mice29,30. The correlative observations of bone marrow
myelopoiesis to drive this expansion in circulatory monocytes, and the
subsequent elevation of lung mononuclear phagocytes are novel to our
study. We expect that the mononuclear phagocyte population in the lung
is directly derived from circulatory monocytes. This is supported by the
circulatory increase being observed from 7 dpi, whereas lung mononuclear
phagocytes remain unchanged until 10 dpi. The absence of the increase in
lung mononuclear phagocytes when circulatory monocytes are ablated
supports this interpretation. However, due to the potential for
persistence of CCR2 on tissue mononuclear phagocytes31we cannot discount the possibility that we directly ablated cells in the
lung as well.
A growing body of work has assessed the interplay between parasitic and
viral infections32. H. polygyrus infection has
been previously described to suppress influenza A (IAV) infections
although the mechanism was undetermined33.Trichinella spiralis infection improves disease outcome to IAV in
a gut damage dependent mechanism, but this was through suppression of
secondary inflammation, rather than reduction in viral titres and no
alteration in the mononuclear phagocyte compartment was
observed34. Schistosoma mansoni (S.
mansoni ) infection is protective against secondary infection with IAV
and in pneumonia virus of mice (PVM) infection it gave a small reduction
in peak PVM load35. S. mansoni larvae pass
through the lung in mice so their effect on respiratory viral infection
is likely due to local inflammatory responses rather than to the
systemic response observed with the strictly enteric H.
polygyrus . A mononuclear phagocyte response was not reported but others
have described S. mansoni to drive recruitment of alternatively
activated macrophages36. Their local immunosuppressive
effect is often detrimental to viral infection control, with bothH. polygyrus and S. mansoni infection reactivating latent
murine γ-herpesvirus infections by suppressing the antiviral IFN
response37.
The promotion of alternative activated macrophages in helminth infection
has also been described in tissues peripheral to the gut. H.
polygyrus induces heart macrophages with a strong type 2 polarisation
which appear at 28 dpi38. The absence of elevated
blood monocytes at this late time point, and the very small increase in
the number of monocytes and mononuclear phagocytes expressing the M2
marker CD301 (predominantly regulatory macrophages, data not shown) at
10 dpi in our study, further indicate that different mechanisms are
responsible for the early induction of lung mononuclear phagocytes afterH. polygyrus infection and a later type-2 immunity driven
expansion of alternatively activated macrophages.
An active factor driving the early H. polygyrus effects observed
in our study has not yet been identified but the fact that both the
antiviral effect and the expansion of both blood monocytes and lung
mononuclear phagocytes depend on IFNAR signalling likely implies a role
for IFN-Is either as direct effectors or stimuli for a secondary signal.
The monocytic response to IFN-I signalling is key to myeloid responses
in many other infections. IFNAR-deficient mice have been used widely in
SARS-CoV-2 models where this disruption impairs recruitment of
Ly6C+ monocytes to the lung39,40. In
other inflammatory conditions this may be due to decreased turnover to
Ly6C- monocytes, but the overall increase in total
monocyte and mononuclear phagocyte numbers in H. polygyrusinfection suggests other additional effects14. IFNAR
signalling has also been linked to mononuclear phagocyte function in IAV
infection41,42 and to inflammatory monocyte
accumulation in mucosal herpes simplex virus
infection43. IFN-I signalling is also key to the
accumulation of inflammatory Ly6C+ mononuclear
phagocytes in the lung associated with older age, with a phenotype
similar to that seen in viral infections such as
IAV44. In PVM infection IFNAR signalling has no effect
on monocytic cells but is required for the induction of inflammatory
conventional dendritic cells, indicating a likely infection specificity
of myeloid IFN-I signalling requirements45. However,
dendritic cells express CD11c and may contribute to the
CD11c+ macrophages we have found to expand afterH. polygyrus infection. Indeed the ongoing difficulty in
characterising the mononuclear phagocyte sub-compartments in murine
pulmonary inflammation (recently reviewed46,47) due to
shared expression of the key markers used for steady state
characterisation is also a limitation in this study. It is important to
note that similar to the work in our study the use of global IFNAR
knockouts in the studies above means that despite the usually high
expression of IFNAR on monocytes, their response may be dependent on
secondary cells.
Effects of IFN-Is on blood monocytes are less well studied including inIfnar1 deficiency. The historical characterisations of
IFNAR-deficient mice showed elevated circulatory monocytes, counter to
our model, but the increased risk of infection in IFNAR-deficient mice,
at a time before modern infection control measures were available, may
have contributed to this effect20. In human peripheral
blood mononuclear cells (PBMCs) classical monocytes are more responsive
to IFN-Is than non-classical monocytes due to differential abundance of
IFNAR48 and culture of PBMCs with IFN-Is will alter
their phenotype49-51. An expansion of circulatory
monocytes and lung CD64+ cells, similar to our
findings, was observed in idiopathic pulmonary fibrosis patients and
associated with increased circulatory IFN-Is52.
Exogenous treatment with IFNα for two to four weeks in asthma patients
also increased the numbers of circulating monocytes and increased their
antigen presenting phenotype53.
Clearly, from the array of responses described above the context of
IFN-I signalling is crucial. The outcome of IFN-I stimulation is
modulated by concomitant signalling/environmental factors that certainly
also play a role in H. polygyrus responses54.
One such environmental factor is the microbiome and it’s metabolic
products, the presence of which is essential for the H. polygyrusantiviral effect6. While we were unable to test the
impact of the microbiome in the present study, interactions between
IFN-I production and microbiome components have been well
described55,56 and the microbiome is known to be
modulated in helminth infections including H. polygyrusinfection57,58.
The apparent lack of dependency on IFNAR signalling for bone marrow
monopoiesis in this model suggests alternative signalling requirements.
Interferon treatments have been described to affect
lymphopoiesis59, IFN-I suppresses neutrophil
differentiation60,61, and IFNγ promotes
myelopoiesis62,63. Parasitic modulation of the bone
marrow has also been described, with Trichuris muris andTrichuris gondii infections modulating haematopoiesis in an IFNγ
dependent manner64,65. There is also substantial
evidence that interferon regulatory factor 5 (IRF5) is a key regulator
of hematopoietic development, particularly in myeloid
lineages66,67. H. polygyrus has been recently
described to induce IFN-γ (IFN-II) early in
infection68 which may be responsible for the initial
induction of monopoiesis. The deeply interlinked feedback loops between
IFN-I and IFN-II signalling may explain why we see a reduced effect on
monocytosis in the bone marrow69.
The mechanism of the anti-viral effect of H. polygyrus -induced
mononuclear phagocytes is as yet unclear. In RSV infection mononuclear
phagocytes are known to produce TNFα which is important for control of
viral expansion, providing a possible mechanism13 but
their direct effects on viral load were not assessed. Our findings
directly demonstrate, for the first time, that elevating mononuclear
phagocyte numbers in the lung prior to RSV infection can reduce viral
load. Monocytes/macrophages are also known to produce IFNβ, viperin and
OAS in response to viral exposure, thus the expansion of mononuclear
phagocytes in the lung may explain the previously observed increase in
these antiviral factors in whole lung samples during H. polygyrusinfection19,70-73.
In conclusion, we show that intestinal helminth infection can induce
transient systemic monocytosis and increased numbers of mononuclear
phagocytes within the lung that are essential to reducing the burden of
viral respiratory infection. Due to the IFNAR dependency of this
expansion in the blood and lung we hypothesise that helminth induced
IFN-Is or secondary signalling products play a key role in these
effects. Future work will be required to identify the specific factors
that act upon monocytes to drives these effects and to further
characterise the specific anti-viral mechanisms of these cells in RSV
infection.