Mfge8 diminishes the severity of tissue fibrosis in mice by binding and targeting collagen for uptake by macrophages
Kamran Atabai, Sina Jame, Nabil Azhar, Alex Kuo, Michael Lam, William McKleroy, Greg DeHart, Salman Rahman, Dee Dee Xia, Andrew C. Melton, Paul Wolters, Claire L. Emson, Scott M. Turner, Zena Werb, Dean Sheppard
Kamran Atabai, Sina Jame, Nabil Azhar, Alex Kuo, Michael Lam, William McKleroy, Greg DeHart, Salman Rahman, Dee Dee Xia, Andrew C. Melton, Paul Wolters, Claire L. Emson, Scott M. Turner, Zena Werb, Dean Sheppard
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Research Article Pulmonology

Mfge8 diminishes the severity of tissue fibrosis in mice by binding and targeting collagen for uptake by macrophages

Abstract

Milk fat globule epidermal growth factor 8 (Mfge8) is a soluble glycoprotein known to regulate inflammation and immunity by mediating apoptotic cell clearance. Since fibrosis can occur as a result of exaggerated apoptosis and inflammation, we set out to investigate the hypothesis that Mfge8 might negatively regulate tissue fibrosis. We report here that Mfge8 does decrease the severity of tissue fibrosis in a mouse model of pulmonary fibrosis; however, it does so not through effects on inflammation and apoptotic cell clearance, but by binding and targeting collagen for cellular uptake through its discoidin domains. Initial analysis revealed that Mfge8–/– mice exhibited enhanced pulmonary fibrosis after bleomycin-induced lung injury. However, they did not have increased inflammation or impaired apoptotic cell clearance after lung injury compared with Mfge8+/+ mice; rather, they had a defect in collagen turnover. Further experiments indicated that Mfge8 directly bound collagen and that Mfge8–/– macrophages exhibited defective collagen uptake that could be rescued by recombinant Mfge8 containing at least one discoidin domain. These data demonstrate a critical role for Mfge8 in decreasing the severity of murine tissue fibrosis by facilitating the removal of accumulated collagen.

Authors

Kamran Atabai, Sina Jame, Nabil Azhar, Alex Kuo, Michael Lam, William McKleroy, Greg DeHart, Salman Rahman, Dee Dee Xia, Andrew C. Melton, Paul Wolters, Claire L. Emson, Scott M. Turner, Zena Werb, Dean Sheppard

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Figure 4

Mfge8 mediates collagen uptake in vitro.

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Mfge8 mediates collagen uptake in vitro. (A) Primary alveolar macrophage...
(A) Primary alveolar macrophages were cultured for 30 minutes with FITC-conjugated type I collagen, and uptake was evaluated by fluorescence microscopy (red arrows). Scale bar: 10 μm. (B) Ingestion of collagen was quantified as the collagen uptake index (CUI: number of macrophages with ingestions divided by the total number of macrophages counted). The addition of unlabeled type I collagen (25, 50, 150 μg/ml) inhibited uptake of FITC-conjugated collagen in a dose-dependent fashion (*P < 0.001, 1-way ANOVA with Bonferroni t test for multiple comparisons; n = 3–4; data are expressed as percent control relative to wild-type uptake). (C) Alveolar macrophages from Mfge8–/– mice had significantly impaired collagen uptake index as compared with Mfge8+/+ alveolar macrophages. (*P = 0.001, 1-way ANOVA with Bonferroni t test for multiple comparisons; n = 3–5). Addition of rMfge8 (μg/ml) rescued collagen uptake in Mfge8–/– alveolar macrophages (**P = 0.007, ***P = 0.023). (D) Addition of rMfge8 (μg/ml) increased collagen uptake in Mfge8+/+ alveolar macrophages under serum-starved conditions (*P = 0.009, Student’s t test to compare indicated columns; n = 5–6). Data are presented as mean ± SEM.