Regulation of toll-like receptor-mediated innate immune response in chikungunya virus-infected and Fisetin-treated human hepatocellular carcinoma Huh7 cells / Rafidah Lani
Excessive inflammation caused by chikungunya virus (CHIKV) infection is manifested by incapacitating joint pain and prolonged arthritis in the chronic phase of CHIKV infection. Arthritis resulted from a large influx of infiltrating immune cells driven by pro-inflammatory cytokines and chemokines....
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| Format: | Thesis |
| Published: |
2022
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| Online Access: | http://studentsrepo.um.edu.my/14814/4/Rafidah_Lani.pdf http://studentsrepo.um.edu.my/14814/ |
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| Summary: | Excessive inflammation caused by chikungunya virus (CHIKV) infection is manifested
by incapacitating joint pain and prolonged arthritis in the chronic phase of CHIKV
infection. Arthritis resulted from a large influx of infiltrating immune cells driven by
pro-inflammatory cytokines and chemokines. Fisetin demonstrated anti-CHIKV activity
and has long been known to possess anti-inflammatory and immunomodulatory
properties. Modulating toll-like receptor (TLR)-mediated innate immune responses
might be one of the mechanisms utilized by fisetin against CHIKV infection. This study
was designed to assess fisetin capacity in modulating the TLR-mediated antiviral
responses against CHIKV in Huh7 cells. The hallmarks of inflammation and
cytopathological effects (CPE) were evaluated to test the hypothesis that fisetin
promotes TLR-mediated antiviral responses and limit CPE in CHIKV-infected Huh7
cells. CHIKV-induced CPE was observed using bright-field microscopy. Viral
infectivity was determined using a plaque-forming unit assay. CHIKV-E1 RNA copy
number and antiviral genes mRNA level were quantified using qRT-PCR. The
expression of TLR and CHIKV-E2 proteins was measured by performing an
immunofluorescence assay. The level of the inflammatory cytokines was determined
using a human cytokine array. The expression of the key transcription factors was
determined using immunoblot assay. The results showed that fisetin induced the
expression of the antiviral genes at an early time-point by promoting the
phosphorylation of IRF3 and IRF7 in CHIKV-infected Huh7 cells. Fisetin hindered
excessive inflammation in CHIKV-infected Huh7 cells by impeding
over-phosphorylation of NF-κB and reducing pro-inflammatory cytokines levels. Fisetin
also recovered Huh7 cells from CHIKV-induced CPE. The results also suggested that fisetin does not affect the level of the pro-and anti-inflammatory cytokines such as
IL-1α, IL-1β, IL-8, IL-10, IL-13, GM-CSF, GRO, MCP-1, MIP-1β, and RANTES; in
uninfected Huh7 cells. These suggested that fisetin is pharmacologically active only in
CHIKV-infected Huh7 cells and not uninfected cells. Findings from the study supported
the hypothesis that fisetin promotes antiviral responses and limit CPE in
CHIKV-infected Huh7 cells. Fisetin's balancing act in CHIKV-infected Huh7 cells was
subjected to fisetin modulating the key transcription factors manipulated by CHIKV
infection. On this basis, further research into fisetin in targeting host inflammation as a
therapeutic strategy is being considered. |
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