| Objectives:1. To investigate the effect of Ang II on expression of MMP-9 in THP-1 macrophages.2. To investigate the effect of Ang-(1-7) on expression of MMP-9 induced by Ang II in THP-1 macrophages, and explore the potential mechanism.Methods:1. Macrophages were converted from THP-1 monocytes induced by 0.1μmol/LPMA for 48 h.2. Part 1:①PMA group,also the control group②PMA+Ang II group(10-7 mol/L, 1 hour)③PMA+Ang II +PDTC group(10μmol/L, 30 minutes)④PDTC group. Part 2:①PMA group,the control group②Ang-(1-7) (10-6mol/L,0.5h)③AngII (10-7 mol/L,1h)④Ang-(1-7) in different concentrations (10-8~10-5mol/L,0.5 h)+AngII(10-7 mol/L,1h)⑤A-799 (10-5mol/L,10min)+Ang-(1-7)(10-6mol/L,0.5h)+AngII (10-7mol/L,1h).3. The expression of MMP-9 and phosphorylation of NF-κB p65 in THP-1 macrophages were determined by Western-Blotting, and the expression of MMP-9 mRNA was measured by RT-PCR.Results:1. Macrophages were converted from THP-1 monocytes induced by PMAMacrophages were converted from THP-1 monocytes induced by 0.1μmol/L PMA for 48h, the suspended, round THP-1 monocytes changede into macrophages which stretched out pseudopodia, out of shapes and adhered to the wall.2. The expression of NF-κB p65 and MMP-9 treated with PDTC and Ang IICompared with the control group, the expression of MMP-9(1.06±0.11,P<0.05) and phosphorylation of NF-κB p65 (1.02±0.10,P<0.05) in THP-1 macrophages were expressed significantly when treated with Ang II (10-7mol/L);and the expression of MMP-9mRNA was upregulated(1.22±0.08,P<0.05).PMA+AngII+PDTC group reduced MMP-9(1.04±0.14,)and NF-κB p65(0.99±0.12) expression (P<0.01),decreased the expression of MMP-9 mRNA(0.90±0.06,P<0.01); When compared with the control group, the expression of phosphorylation of NF-κB p65 has no significantly difference (P>0.05)but MMP-9 has(P<0.01).3. The expression of NF-κB p65 and MMP-9 treated with A-799,Ang-(1-7) and AngIICompared with the control group, the expression of phosphorylation of NF-κB p65(1.04±0.05,P<0.05) and MMP-9 (1.04±0.21, P<0.01) in THP-1 macrophages were expressed significantly when treated with Ang II (10-7mol/L,1h), the former control group 0.98±0.07 and the latter 0.97±0.07, MMP-9mRNA (0.86±0.81)expression was upregulated than the control group (0.70±0.06) (P<0.01).10-6mol/LAng-(1-7) inhibitted the phosphorylation of nuclear NF-κB p65 (1.00±0.09) and MMP-9 (0.97±0.18) expression, MMP-9mRNA downregulated(0.94±0.07) .However, when incubated with A-799, an receptor blocker of Ang-(1-7), the effects of Ang-(1-7) on inhibiting NF-κB p65 (1.04±0.04, P<0.01) and MMP-9(1.03±0.17,P<0.01) expression was significantly attenuated, MMP-9mRNA upregulated (1.29±0.12, P<0.01).Ang-(1-7) dose-dependently inhibited the expression of MMP-9 in THP-1 macrophages induced by AngII(P<0.05), MMP-9 expressed respectively was 1.01±0.17, 0.99±0.15, 0.97±0.18 and 0.96±0.30,and MMP-9mRNA was 1.22±0.10, 1.12±0.08, 0.94±0.07 and 0.91±0.05 in four groups of Ang-(1-7)10-8~10-5mol/L+AngII.Conclusion:1.AngII significantly induced the expression of the MMP-9 in THP-1 macrophages mediated by NF-κB activition2.NF-κB signaling pathway is one of the most important pathway, which mediated the expression of the MMP-9 induced by AngII in THP-1 macrophages.3.Combining with the specific receptor Mas ,Ang-(1-7)dose-dependently inhibits the expression of the MMP-9 induced by AngII in THP-1 macrophages; Ang-(1-7) rises the function by inhibiting NF-κB activation induced by AngII. |
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