Microbial infection of the cornea is one of the leading causes of preventable blindness world-wide. The infection results in corneal opacity, inflammation, and intense
pain that could lead to permanent blindness if left untreated. Treatment involves
antimicrobial agents followed by corticosteroids to suppress inflammation. However,
there is a need for a more targeted treatment to regulate inflammation as steroids are
non-specific.
In both human patients and our murine model of bacterial and fungal keratitis,
neutrophils are the predominant infiltrating cell-type in the cornea comprising up to 90%
of recruited cells. Neutrophils are essential for bacterial killing and acts as an important
source of proinflammatory cytokines during microbial keratitis. Depletion of neutrophils
leads to tremendously impaired bacterial clearance and decreased cytokine levels in the
cornea. We previously reported that the proinflammatory cytokines IL-1α and IL-1β are
highly upregulated in both human and mice during microbial keratitis. IL-1β is necessary
for neutrophil recruitment to the corneas and subsequently protection from the infection.
However, whether there is a role for IL-1α and what role that may be is unclear. This
thesis explores the production and regulation of IL-1α, and what role it is playing in the
cornea during infection.
We found that while neutrophils are the predominant source of IL-1β in the
cornea, both neutrophils and monocytes contribute to IL-1α production which peaked at
24hpi. To examine whether there is a role for IL-1α in the response to bacterial infection
of the cornea, we compared WT, Il1a-/-, Il1b-/-, and Il1a-/-Il1b-/- mice infected with P.
aeruginosa. In accordance with our previous studies, Il1b-/- and DKO mice were unable
to control the infection due to delayed neutrophil and monocyte recruitment to the
cornea. On the contrary, Il1a-/- mice had the opposite effect—showing enhanced
bacteria killing compared to WT mice despite no differences in neutrophil recruitment.
We found no differences in neutrophil effector functions between WT and Il1a-/-
neutrophils. However, RNA sequencing revealed Il1a-/- neutrophils had a more
proinflammatory transcriptomic profile than WT neutrophils with elevations in C1q
expression. These findings reveal a novel role for IL-1α, that is distinct from IL-1β, in
regulating the response to P. aeruginosa infection in the cornea.
Further, we examined the regulation of IL-1α secretion compared to IL-1β in
macrophages and neutrophils. The mechanism for IL-1β secretion is well characterized
in macrophages where it is tightly regulated in a two-step process mediated by
inflammasomes and GSDMD. We found that IL-1α secretion by macrophages is also
mediated by GSDMD, but not NLRP3 inflammasome. However, IL-1α secretion by
neutrophils is independent of both NLRP3 and GSDMD in response to stimulation with
β-glucan. Secretion of IL-1α by neutrophils is also independent of cell death. Instead,
we found that IL-1α secretion is mediated by extracellular vesicle release.
In conclusion, we described a novel role for IL-1α that is distinct from IL-1β
during bacterial infection of the corneas—where IL-1α is detrimental and IL-1β is
protective. We also found that IL-1α is important for monocyte recruitment to the cornea
whereas IL-1β is essential for neutrophil recruitment. Our finding adds to the increasing
evidence showing non-redundant roles for the two cytokines despite sharing the same
receptor, IL-1R1. Further, we found differential regulation for IL-1α and IL-1β secretion
by neutrophils. While IL-1β is dependent on NLRP3 and GSDMD for secretion, IL-1α
release is mediated by extracellular vesicles. Together, these studies improve our
understanding of the inflammatory response during microbial keratitis and identify
potential targets for therapeutics.