Epithelial-macrophage interactions determine pulmonary fibrosis susceptibility in Hermansky-Pudlak syndrome
Lisa R. Young, Peter M. Gulleman, Chelsi W. Short, Harikrishna Tanjore, Taylor Sherrill, Aidong Qi, Andrew P. McBride, Rinat Zaynagetdinov, John T. Benjamin, William E. Lawson, Sergey V. Novitskiy, Timothy S. Blackwell
Lisa R. Young, Peter M. Gulleman, Chelsi W. Short, Harikrishna Tanjore, Taylor Sherrill, Aidong Qi, Andrew P. McBride, Rinat Zaynagetdinov, John T. Benjamin, William E. Lawson, Sergey V. Novitskiy, Timothy S. Blackwell
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Research Article Pulmonology

Epithelial-macrophage interactions determine pulmonary fibrosis susceptibility in Hermansky-Pudlak syndrome

Abstract

Alveolar epithelial cell (AEC) dysfunction underlies the pathogenesis of pulmonary fibrosis in Hermansky-Pudlak syndrome (HPS) and other genetic syndromes associated with interstitial lung disease; however, mechanisms linking AEC dysfunction and fibrotic remodeling are incompletely understood. Since increased macrophage recruitment precedes pulmonary fibrosis in HPS, we investigated whether crosstalk between AECs and macrophages determines fibrotic susceptibility. We found that AECs from HPS mice produce excessive MCP-1, which was associated with increased macrophages in the lungs of unchallenged HPS mice. Blocking MCP-1/CCR2 signaling in HPS mice with genetic deficiency of CCR2 or targeted deletion of MCP-1 in AECs normalized macrophage recruitment, decreased AEC apoptosis, and reduced lung fibrosis in these mice following treatment with low-dose bleomycin. We observed increased TGF-β production by HPS macrophages, which was eliminated by CCR2 deletion. Selective deletion of TGF-β in myeloid cells or of TGF-β signaling in AECs through deletion of TGFBR2 protected HPS mice from AEC apoptosis and bleomycin-induced fibrosis. Together, these data reveal a feedback loop in which increased MCP-1 production by dysfunctional AECs results in recruitment and activation of lung macrophages that produce TGF-β, thus amplifying the fibrotic cascade through AEC apoptosis and stimulation of fibrotic remodeling.

Authors

Lisa R. Young, Peter M. Gulleman, Chelsi W. Short, Harikrishna Tanjore, Taylor Sherrill, Aidong Qi, Andrew P. McBride, Rinat Zaynagetdinov, John T. Benjamin, William E. Lawson, Sergey V. Novitskiy, Timothy S. Blackwell

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Figure 3

Epithelial MCP-1 drives fibrotic susceptibility in HPS2 mice via CCR2 signaling in myeloid cells.

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Epithelial MCP-1 drives fibrotic susceptibility in HPS2 mice via CCR2 si...
(A–D) MCP-1 was deleted in the lung epithelium of HPS mice by crossing HPS2 mice with SPC.Cre/MCP1 floxed mice (HPS2/MCP1ΔAEC mice). (A) MCP-1 production determined by ELISA from alveolar epithelial cells (AECs) isolated from unchallenged mice (mean ± SEM); n = 6 for WT groups and n = 12 for HPS groups. Comparisons between groups were conducted by ANOVA with Tukey post-test, *P < 0.05 vs. WT, **P < 0.001 vs. others. (B) Bronchoalveolar lavage MCP-1 levels 24 hours after bleomycin challenge (mean ± SEM); n = 9 WT and n = 5 each for WT/MCP1 ΔAEC, HPS2, and HPS2/MCP1ΔAEC. Comparisons between groups were conducted by Kruskal-Wallis test with Dunn’s multiple comparisons post-test, *P < 0.01 vs. other groups. (C) Lung collagen content of left lung quantitated in age-matched unchallenged mice or 7 days after bleomycin (mean ± SEM). For unchallenged groups, n = 4 WT and n = 6 WT/MCP1ΔAEC. For bleomycin-challenged groups, n = 6 WT/MCP1ΔAEC, n = 3 WT/SPC.Cre– littermate controls, n = 7 HPS2/MCP1ΔAEC, and n = 8 HPS2 SPC.Cre– littermate controls. Comparisons between groups were conducted by Kruskal-Wallis test with Dunn’s multiple comparisons post-test, *P < 0.05 vs. other bleomycin-challenged groups. (D) Fibrosis score. Data are presented as box-and-whisker Tukey plots; n = 4 for WT groups, n = 5 for HPS groups, and n = 3 SPC.Cre– controls. Comparisons between groups were conducted by Kruskal-Wallis test with Dunn’s multiple comparisons post-test, *P < 0.01 vs. WT, **P < 0.05 vs. HPS2/MCP1ΔAEC. (E and F) Mice underwent whole body irradiation followed by transplantation of whole marrow from WT, HPS2, or WT/CCR2–/– mice. After 60 days, transplanted mice were challenged with bleomycin. (E) Lung collagen content of the left lung quantitated in unchallenged transplant controls or 7 days after bleomycin treatment (mean ± SEM). For unchallenged groups, n = 4 WT and n = 10 HPS2. For bleomycin-challenged groups, n = 10 recipients of WT marrow and n = 11 recipients of CCR2–/– marrow. Comparisons between bleomycin-challenged groups were conducted using Mann-Whitney U analysis, *P < 0.01. (F) Fibrosis score. Data are presented as box-and-whisker Tukey plots; n = 6 per group. Comparisons were conducted using Mann-Whitney U analysis, *P < 0.05.