| Rhinovirus which belongs to genus Enterovirus,family Picornavirus enterovirus,is a single strand RNA virus.According to neutralising antibody studies,rhinovirus is divided into 100 types,mainly divided into rhinovirus A and rhinovirus B.In recent years,a new type of rhinovirus has been detected in patients with influenza like respiratory diseases,and their gene sequences suggest that they may be a proposed new species,rhinovirus C.The results showed that the three groups had different virological characteristics,and the replication efficiency of RV-B was low.Compared with that of RV-A or RV-C,the RV-B infection induced fewer cytokines and lower cytotoxicity.Clinical epidemiological study found that in patients with severe asthma,the detection rate of RV-C was significantly higher than RV-A and RV-B.Rhinovirus genome contrains about 7200 nucleotides,and had one open reading frame encoding a polyprotein.The protein of 2A protease and 3C enzyme cleaves the polyprotein precursor into 4 structural and 7 nonstructural proteins.The 3C protein is a cysteine protease,which plays an important role in viral replication and the antagonism with the host immune response.Previous studies have reported that RV-C causes more severe disease than RV-A and RV-B.Because of the difficulty to cultivate RV-C in vitro by conventional cell culture,so the mechanisms of RV-C leading to respiratory disease remains obscue.In order to study the likely pathogenesis of asthma exacerbation induced by rhinovirus,this research aimed to construct a chimeric infectious clone of RV1B-651 which the region of 3C gene of RV1 B was replaced by RV-C 3C gene.Then,the wide type RV1 B and the chimeric virus RV1B-651 infected the asthmatic mice sensitized by OVA.The index of respiratory allergy was detected to compare the severity of asthma induced by RV1 B and RV1B-651.Objective:1 To construct the infectious clone of the rhinovirus-A RV1 B and the infectious clone of chimeric RV1B-651,which layed the foundation for the study of the effects on asthma of 3C protein.2 To study the influence of RV1B-651 and RV1 B in asthmatic mice,so as to reveal the relationship between 3C protein and the severity of asthma exacerbation.Methods:1.At first,to construct the infectious clone of RV1 B full genome.Then,to construct the chimeric infectious clone of RV1B-651 with the 3C gene replaced with LZ651 3C gene.After infectious clone of RV1 B and RV1B-651 was transinfected the Hela cells,RV1 B and RV1B-651 virus was rescued.2.The rhinovirus RV1 B and chimeric virus infected ovalbumin sensitized asthmatic mice,and respiratory tract allergy index was detected to compare the severity of asthma induced by RV1 B and RV1B-651.Results:1.We have successfully construct the infectious clone of rhinovirus RV1 B and infectious clone of chimeric rhinovirus RV1B-651 to lay the foundation for a better study of the pathogenic mechanism of 3C protein.2.Cytokines in the BALF fluid of asthmatic mice induced by RV1 B and RV1B-651 was examined.In the OVA group,the levels of IFN-? and IFN-? were higher than PBS group(P < 0.05).To compare RV1B/OVA group and RV1B-651/OVA group with OVA group,the levels of IL-4,IFN-? IFN-?were significantly increased(P < 0.05).Compared with group RV1B/OVA,the levels of IFN-? and IL-4 were significantly increased in group RV1B-651/OVA(P < 0.05).In the lung pathological examination,RV1B/OVA group,RV1B651/OVA group and OVA group had obvious inflammatory cells infiltration to compare with PBS group.Conclusion:This study successfully constructed infectious clone of RV1 B and chimeric virus RV1B-651,and rescue the RV1 B and RV1B-651 virus to study the relationship between 3C protein and asthma.The ova induced asthma in mice;3C gene may be the cause of differences in cytokine levels in ovalbumin sensitized asthmatic mice after RV1 B virus and RV1B-651 chimeric virus infection,the specific mechanism needs to be further studied. |
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