Aesculetin Attenuates Pulmonary Fibrosis Induced by Close Contact of Macrophages with Alveolar Epithelial Cells

Abstract

Abstract Objectives Pulmonary fibrosis is a disease in which lung tissues become fibrous and causes severe respiratory disturbances. Various stimuli induce infiltration of macrophages to the respiratory tract. These macrophages secrete various inflammatory cytokines leading to development of pulmonary fibrosis via epithelial–mesenchymal transition (EMT) process. Aesculetin, a major component of Sancho tree and Chicory, is known to have antioxidant and anti-inflammatory effects in the vascular and immune system. Methods Human alveolar basal epithelial A549 cells were cultured in conditioned media of THP-1 monocyte-derived macrophages for 24 h. Aesculetin at the concentrations of 1–20 μM did not show cytotoxicity of A549 cells. Alveolar epithelial cells were incubated with interleukin (IL)-8. Western blotting examined EMT-associated fibrotic proteins from A549 cell lysates. Matrix metalloproteinase (MMP) activity was measured with gelatin zymography. In addition, inflammation- and fibrosis-related cytokines were measured by using ELISA kits. Results The epithelial markers of E-cadherin and ZO-1 were reduced in cells exposed to macrophage-conditioned media containing IL-8 and TNF-α. Macrophage-conditioned media enhanced expression of the mesenchymal fibrotic markers of α-smooth muscle actin (α-SMA), vimentin and fibronectin, and the fibrotic proteins of collagen I and collagen IV were enhanced. However, ≥10 μM aesculetin reciprocally manipulated the expression levels of these proteins of A549 cells. In addition, macrophage-conditioned media enhanced the expression and activity of MT1-MMP, MMP-2 and MMP-9. In contrast, the expression of tissue inhibitor of metalloproteinases (TIMP)-1 and TIMP-2 were reduced by exposure of alveolar cells to conditioned media. Proinflammatory and chemotactic IL-8 reduced E-cadherin and conversely enhanced N-cadherin and α-SMA in A549 cells, which was reciprocally modulated by ≥ 10 μM aesculetin. These results demonstrate that aesculetin may ameliorate EMT-associated pulmonary fibrosis caused by contact of blood-derived macrophages and alveolar cells. Conclusions Aesculetin maybe a promising agent treating progressive pulmonary disorders owing to macrophage-mediated inflammation. Funding Sources No funding sources to report.