Expressions of toll-like receptors and cytokine modulation of antiviral innate immune responses in feline infectious peritonitis virus infection
Feline infectious peritonitis (FIP) is a fatal immune-mediated disease of domestic cats caused by feline infectious peritonitis virus (FIPV), a member of coronavirus family. Previous studies have shown that the lack or absence of adaptive cellular immunity and aberrant antibody production may...
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| Format: | Thesis |
| Language: | English |
| Published: |
2020
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| Subjects: | |
| Online Access: | http://psasir.upm.edu.my/id/eprint/97878/1/FPV%202021%2019%20UPMIR.pdf |
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| Summary: | Feline infectious peritonitis (FIP) is a fatal immune-mediated disease of domestic cats
caused by feline infectious peritonitis virus (FIPV), a member of coronavirus family.
Previous studies have shown that the lack or absence of adaptive cellular immunity and
aberrant antibody production may lead to cats succumbing to FIP. FIPV viral protein
has been shown to antagonize the production of type I interferon (IFN), the key
cytokine crucial during early stage of viral infection and downregulate the production
of anti-inflammatory cytokine such as IL-10. Natural killer (NK) cells, another
important innate immune cell has been shown to be dysregulated in FIPV infection,
suggesting the importance of innate immunity in FIP pathogenesis. Despite these
findings, the information on pattern recognition receptors (PRRs) that play a role in the
detection of common molecules on pathogens, or also known as pathogen-associated
molecular patterns (PAMPs) is still lacking. Therefore, this study attempts to
investigate the role of the Toll-like receptors (TLRs), one of the PRRs in FIPV
infection in vitro and ex vivo. Stimulation of TLR pathway activates the NF-κB and
type I IFN-related pathways which in turn produce pro-inflammatory cytokines such as
TNF-α, and type-I IFN (IFN- α, IFN-β). To achieve these objectives, Crandell-Rees
Feline Kidney (CRFK) cells and feline CD14+ monocytes were infected with FIPV 79-
1146 and harvested at 4, 12, and 24 hours post-infection (hpi). The infection of FIPV
into these cells was confirmed by immunofluorescence (IF) assay. The mRNA
expression of several TLRs (TLR3, TLR7, and TLR9) and some of downstream
cytokines (TNF-α, IL-10, and IFN-β) were measured using real time PCR (qPCR). The
results were then correlated with the viral load copy number. Results from the in vitro
study revealed the involvement of TLR9 in TNF-α, and IFN-β cytokine modulation in
CRFK cells. However, TLR3 was expressed at low level and remained stable
throughout the in vitro infection while TLR7 was not detectable. In contrast, TLR7
expression was induced upon ex vivo infection of feline monocytes at earlier time point
indicating the activation of TLR7 by binding to its ligand, single-stranded RNA.
However, its expression was significantly reduced at later time points which could be
due to the immune evasion strategy posed by FIPV. A similar trend was also observed for TNF-α expression postulating the role of TLR7 in the regulation of TNF-α which
has been implicated as the major pro-inflammatory cytokine seen in FIP cats.
Furthermore, IFN-β gene was also expressed throughout the course of infection which
could be mediated by TLR7 activation and independent of TLR3 and TLR9 signaling
pathways as the level of expression of these two TLRs remained stable. In general, the
viral replication kinetics in CRFK cells and feline monocytes were consistent with
other studies where the virus increased from 4 hpi to 12 hpi and maintained or reduced
from 12 hpi to 24 hpi. Interestingly, the expression of TLR7 in monocytes of one
seronegative cat was sustained throughout the infection, and the expression of TNF-α,
IL-10, and IFN-β were markedly upregulated suggesting a control of viral protein
synthesis by the immune system which was confirmed by the absence of viral antigen
by IF, although the viral RNA was present. Taken together, this study provides a new
insight on the role of TLRs in modulating the immune responses in FIPV infection.
Although different cells express a different set of TLRs as observed in this study, this
report implicates the role of TLR7 and TLR9 in FIPV infection, therefore setting up an
avenue for further investigation into their signaling pathway and possible formulation
of new therapeutic strategies. |
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