| Objective:To explore the dynamic of complement mRNA from mouse macrophage ANA-1 infected with DENV-2 virus and its polarization. Methods: DENV-2 replicated in C6/36 cells was identified by RT-PCR and detected by TCID50. The polarized models of macrophage ANA-1 were established with LPS or IL-4 After treated 3h,8h,12h and 24h, macrophage polarization were estimated by detecting the expression of IL-1?, CCL2 and Arg-1 mRNA with Real-time fluorescent quantitative PCR. The intracellular complement C1q, C3, CfB and CRIg mRNA were quantitatively analyzed. Identify the infection of DENV-2 by PCR and the expression of NS1, Mavs and complement components mRNA were detected by Real-time PCR in M0/M2 polarized macrophage were infected by DENV-2. Results:1) The expression of complement mRNA in M1/M2 polarized ANA-1 cells:The intracellular complement C1q, C3, CfB and CRIg mRNAs all were up-regulated in M1/M2 polarized macrophage. ?The intracellular Clq and C3 mRNA in polarized M2 were significantly higher, in which the peak value of C1q and C3 were to 94.9±12.9 and 11.3±2.4 at 12h, respectively (P<0.05). ?The CfB mRNA in polarized M1 increased obviously, in which its 2-??ct was to 61.4±6.2 at 12h. ? The CRIg mRNA in both groups was only up-regulated at 24h, in which the 2-??ct value was 6.5±1.8 in M1 and 10.8±3.2 in M2 (P<0.05).2) The expression of complement mRNA in M0/M2 polarized ANA-1 cells infected by DENV-2:Comparing with M2 uninfected group, the expression of C1q and C3 were reduced in M2 infected cells except for CRIg: ?The peak expression of C1q was 3.84±1.9 at 12h in M2 infected group; ?The highest expression of C3 was 5.08±2.25 in 12h. ? The expression of CRIg in M2 infected group was induced. However in contrast with MO infected group:? The expression of C1q and CRIg mRNA was higher in M2 infected cell.? The expression of C3 mRNA was higher than MO infected group at 12h, thereafter it was opposite at 24h. ? The mRNA of CfB was inhibited in ANA-1 infected by DENV-2, however there was no significant difference between MO and M2 polarization ANA-1.3) The dynamic characteristic of polarization in ANA-1 infected by DENV-2:The most expression of IL-1?, CCL2 and Arg-1 mRNA were inhibited in MO infected ANA-1 cells. The M2 polarization was delayed or reversed in ANA-1 cell infected by DENV-2.4) The expression of NS1, Mavs and complement components mRNA:There was no significant difference of the expression of NS1 and Mavs mRNA between M0 and M2 polarized ANA-1 cells infected by DENV-2. Conclusion:1) There are differences in the expression of complement mRNA between M1 and M2 polarized ANA-1 cells:The mRNA of CfB was higher in M1 polarization. However the mRNA of C1q, C3 and CRIg were higher in M2 polarization ANA-1.2) There are differences in the expression of complement mRNA in M0/M2 polarized ANA-1 infected by DENV-2:The expression of C1q and C3 were down-regulated in M2 infected ANA-1, while the mRNA of CRIg was in the regular or induced levels. The expression of C1q and CRIg were increased relatively comparing with M0 infected ANA-1.3) The polarization state was delayed or reversed by the infection of DENV.4) There was no significant relationship among the expression of complement, the multiplication of DENV-2 and anti-virus of ANA-1. |
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