Macrophage exosomes transfer angiotensin II type 1 receptor to lung fibroblasts mediating bleomycin-induced pulmonary fibrosis

Background: Macrophages are involved in the pathogenesis of idiopathic pulmonary fibrosis, partially by activating lung fibroblasts. However, how macrophages communicate with lung fibroblasts is largely unexplored. Exosomes can mediate intercellular communication, whereas its role in lung fibrogenesis is unclear. Here we aim to investigate whether exosomes can mediate the crosstalk between macrophages and lung fibroblasts and subsequently induce fibrosis. Methods: In vivo, bleomycin (BLM)-induced lung fibrosis model was established and macrophages infiltration was examined. The effects of GW4869, an exosomes inhibitor, on lung fibrosis were assessed. Moreover, macrophage exosomes were injected into mice to observe its pro-fibrotic effects. In vitro, exosomes derived from angiotensin II (Ang II)-stimulated macrophages were collected. Then, lung fibroblasts were treated with the exosomes. Twenty-four hours later, protein levels of alpha-collagen I, angiotensin II type 1 receptor (AT1R), transforming growth factor-beta (TGF-beta), and phospho-Smad2/3 (p-Smad2/3) in lung fibroblasts were examined. The Student's t test or analysis of variance were used for statistical analysis. Results: In vivo, BLM-treated mice showed enhanced infiltration of macrophages, increased fibrotic alterations, and higher levels of Ang II and AT1R. GW4869 attenuated BLM-induced pulmonary fibrosis. Mice with exosomes injection showed fibrotic features with higher levels of Ang II and AT1R, which was reversed by irbesartan. In vitro, we found that macrophages secreted a great number of exosomes. The exosomes were taken by fibroblasts and resulted in higher levels of AT1R (0.22 +/- 0.02 vs. 0.07 +/- 0.02, t = 8.66, P = 0.001), TGF-beta (0.54 +/- 0.05 vs. 0.09 +/- 0.06, t = 10.00, P < 0.001), p-Smad2/3 (0.58 +/- 0.06 vs. 0.07 +/- 0.03, t = 12.86, P < 0.001) and alpha-collagen I (0.27 +/- 0.02 vs. 0.16 +/- 0.01, t = 7.01, P = 0.002), and increased Ang II secretion (62.27 +/- 7.32 vs. 9.56 +/- 1.68, t = 12.16, P < 0.001). Interestingly, Ang II increased the number of macrophage exosomes, and the protein levels of Alix (1.45 +/- 0.15 vs. 1.00 +/- 0.10, t = 4.32, P = 0.012), AT1R (4.05 +/- 0.64 vs. 1.00 +/- 0.09, t = 8.17, P = 0.001), and glyceraldehyde-3-phosphate dehydrogenase (2.13 +/- 0.36 vs. 1.00 +/- 0.10, t = 5.28, P = 0.006) were increased in exosomes secreted by the same number of macrophages, indicating a positive loop between Ang II and exosomes production. Conclusions: Exosomes mediate intercellular communication between macrophages and fibroblasts plays an important role in BLM-induced pulmonary fibrosis.

Automated summary

The study reveals the crucial role of exosomes in mediating communication between macrophages and lung fibroblasts, leading to pulmonary fibrosis. The presence of Ang II in exosomes from macrophages creates a positive feedback loop.