| Objectives The renin-angiotesin system is related to silicosis fibrosis.Use silicotic rat model made by Si O2 bronchial instillated and fibroblast cell model induced by Ang II to explore the anti-fibrosis oligopeptide,Ac-SDKP via regulate the ACE2-Ang-(1-7)-Mas axis inhibit rat silicosis fibrosis and analyze the mechanism.Methods Using non exposed bronchial perfusion of Si O2 produce silicosis fibrosis animal model in rats.According to the experimental design,rats devided into 6 experiental groups: control group for 4w(C4)and 8w(C8);silicotic model group for(S4)and(S8);Ac-SDKP pre-(Ac-pre)and post-(Ac-post)treatment groups.The vitro experiment use primary culture of rat lung fibroblasts and extend to the second generation cells use Ang II induce to establish cell model.According to the experimental design,the fibroblasts cells divided into 6 groups:control group;Ang II induced group;Ac-SDKP intervened group;Ang-(1-7)intervened group and inhibitor of Mas protein(A779)group.Using HE and Masson staining to observe the morphology of lung tissue.Detect the cellular localization and expression of ACE2 and alpha smooth muscle actin(SMA)use Immunohistochemical staining.Adopt ELISA experiment measures the Ang-(1-7)level in plasm of rat.Test the expression of ACE2,Ang(1-7),Mas,α-SMA and Col I in lung tissue and fibroblasts cells by westernblot.Adopt the method of immumofluorescence observe the location and measures the level of α-SMA and Ang(1-7).Results 1 From the results of HE staining,we can see that the pulmonary alveolis structure are completely normal in C4 and C8 groups.But in S4 group it have several isolate silicotic nodules and we can clear it is maded by proliferous cells,and the silicotic nodules become mixed together and change into fibrous nodules in S8 group.Great quantities of collagen type I deposition in Masson staining,but in Ac-SDKP before and after treatment group,the number and area of silicon nodule are significantly reduced and collagen type I deposition decreased.2 In the Silicosis rats model group,α-SMA expreses in smooth muscle tissue of blood vessels and bronches and strong positive expresses in silicon nodule area.While ACE2 positive expresses in normal alveolar epithelial cell and negative rxpresses in silicon nodules.In Ac-SDKP treatment groups,the α-SMA level is decreased and the ACE2 level is increased.3 Compared with the control groups,the level of Ang(1-7)in plasm of S4 and S8 groups decreased to 62.92% and 57.30%,but in the groups treatment by Ac-SDKP,the Ac-SDKP level increased as 1.60 and 1.68 times as the level of S8 groups(P<0.05).4 The result of immunofluorescence indicate that in the Ang II induced group α-SMA marker fluorescence is strengthened while the fluorescence of Ang(1-7)is weakened,and the Ac-SDKP or Ang(1-7)treatment can reverse that changes.Mas receptor inhibitors also can depress the function of Ac-SDKP and Ang(1-7).5 The protein of lung tissue examed by Westernblot display that the express of α-SMA and Col I are inhanced but the level of ACE2,Ang(1-7)and Mas are decreased;and AcSDKP can reverse it.6 The protein of fibroblast cell induced by Ang II,the level of α-SMA and Col I are increased and the express of ACE2,Ang(1-7)and Mas are decreased and Ac-SDKP and Ang(1-7)reverse the results.The A779 also inhibit the function of Ac-SDKP and Ang(1-7)in fibroblast cell cultivate.Conclusions 1 ACE2-Ang(1–7)-Mas take part in the process of silicosis in rat.2 AcSDKP Inhibit of silicosis fibrosis of rat via increases the expression of ACE2-Ang(1–7)-Mas axis.3 Ac-SDKP via Mas receptor regulated the ACE2-Ang(1–7)-Mas axis to inhibit fibroblasts transform to myofibroblast... |
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