| Summary: | 博士 === 國防醫學院 === 生命科學研究所 === 106 === Enterovirus 71 (EV71) is a member belonged to Picornaviridae family. The infection of EV71 causes outbreaks of hand-foot-mouth-disease(HFMD) and meningitis, paralysis and even death. To study the pathogenesis of EV71, we had established the transgenic mice expressing EV71 receptor for infection (hSCARB2 Tg mice). In our past study, we had unexpectedly discovered an important phenomenon; using rhabdomyosarcoma cell line (RD) to produce EV71, the progeny EV71 will be rapidly mutated and reduce its toxicity.
In this study, we prepare EV71 in Vero (named as EV-V) or sub-passaged in RD (named as EV-R) cells and then used them as pathogens. Interestingly, EV-R exhibited differential virulence: challenging of hSCARB2 Tg mice with EV71 revealed that EV-V was more virulent than EV-R: 100% of mice that received lethal amounts of EV-V died, while all the mice that received EV-R were survived. Severe pathogenesis correlated with viral burdens and proinflammatory cytokine levels were observed in EV-V-challenged mice, but controversy in EV-R-challenged mice. Histopathological sections of spinal cord and the surrounding muscle show EV-V cause more severe myositis, muscle fiber breakage and lymphocyte infiltration. In immunohistochemical (IHC) staining sections confirmed that EV-V leads a higher amount of virus in the tissue. Quantification reverse transcriptase PCR (Q-RTPCR) results evealed that the EV-R virus replicates less and induces less proinflammatory responses as well.
Consensus sequence analysis revealed EV-R rapidly acquired complete mutations at E145G and S241L and partial mutations at V146I of VP1, and acquired a T to C substitution at nucleotide 494 of the 5’-UTR. EV-R exhibited higher binding affinity for another EV71 receptor, human P-selectin glycoprotein ligand-1(hPSGL-1), than EV-V. Both EV71s exhibited no significant difference in binding to hSCARB2. In virus entry assay and replication rate assays showed that EV-R infect cells through PSGL-1 increased and replication faster.
In summary, we pointed to the neglected details of the past EV71 study that the culture system of virus isolated enterovirus and appropriate animal model are critical key points.
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