Detection Of Eosinophil Proliferation And Apoptosis Ex Vivo Using A Single ShRNA-expressing Lentiviral Vector For Mouse CCR3Gene

Objective:This study was performed to investigate the effects of a lentiviral vectorexpressing shRNA for RNA interference of murine CCR3gene on the level of mRNAand protein inhibition and to evaluate its effect on proliferation and apoptosis ofeosinophils, laying a solid foundation for gene therapy of allergic rhinitis in vivo.Methods:Primary cultured mouse bone marrow-derived eosinophils were infected withshRNA-mCCR3interfering virions for48h, and the optimal dose was chosenaccording to preliminary experiments. The interference effects on the level of mRNAand protein inhibition were identified by QPCR and western blot, respectively.Additionally, eosinophil viability was assessed by MTS assay and apoptosis wasdetermined by TUNEL method.Results:(1)The QPCR and western blot results indicate that recombinant shRNA-expressing lentiviral vector specific to mouse CCR3gene has high efficientinterference effect on the level of mRNA and protein inhibition compared withcontrol groups (P<0.05).(2) Compared with control groups, interfering virions inhibit the viability ofeosinophils by MTS assay (P<0.05) and induce apoptosis by TUNEL method(P<0.05).Conclusion:(1) Lentiviral interference vector for mouse CCR3gene has been constructedsuccessfully.(2) We have demonstrated that shRNA-mCCR3interfering virions inhibiteosinophil growth and induce apoptosis.