Résumé
The apoptosis of RK13 cells induced by RHDV was investigated with DAPI staining, DNA ladder, Caspase 3 activity and flow cytometry, etc. The results showed that nuclear staining of infected cells with DAPI showed gradually morphological changes of the nuclei. As shown in the paper, a canonic oligonucleosome-sized DNA ladder was observed in cells harvested at 24h, 48h and 72h post-infection, confirming that DNA fragmentation was induced by RHDV infection. The results of flow cytometry showed that about 63% of cells were in apoptosis at 48h post-infection. Besides, we also demonstrated that the activation of Caspase 3 occurred during the infection process. In conclusion, our results showed that apoptosis in RHD might be determinant in the development of the pathogenesis of RHD.
Sujets)
Animaux , Lapins , Apoptose , Infections à Caliciviridae , Génétique , Virologie , Caspase-3 , Métabolisme , Lignée cellulaire , Noyau de la cellule , Génétique , Virologie , Fragmentation de l'ADN , Virus de la maladie hémorragique du lapin , PhysiologieRésumé
To provide an efficient and safe technology platform for studying the replication and pathogenesis mechanisms of RHDV, the interaction between the RHDV and its host cells, a replicon system of RHDV, was constructed based on the infectious cDNA clone of RHDV, in which VP60 gene encoding the capsid protein was deleted, but all the necessary protease coding regions and non-coding regions were retained. Results from RT-PCR, IFA and qRT-PCR confirmed that the replicon RNA could efficiently replicate in RK-13 cells. Besides, the results also suggested that the capsid protein which is the structural protein of RHDV is necessary for maintaining the viral infectivity.