Galectin-3 mediates lysosome-related inflammation within monocyte-derived macrophages in a mouse model of ischemic brain injury
Miao Wang, Zhentai Huang, Zhihong Du, Jiajing Shan, Qing Ye, Lingxiao Lu, Ming Jiang, Fei Xu, Ziyang Liu, David J.R. Fulton, Rehana K. Leak, Babak Razani, Jun Chen, Xiaoming Hu
Miao Wang, Zhentai Huang, Zhihong Du, Jiajing Shan, Qing Ye, Lingxiao Lu, Ming Jiang, Fei Xu, Ziyang Liu, David J.R. Fulton, Rehana K. Leak, Babak Razani, Jun Chen, Xiaoming Hu
View: Text | PDF
Research Article Inflammation Neuroscience

Galectin-3 mediates lysosome-related inflammation within monocyte-derived macrophages in a mouse model of ischemic brain injury

Abstract

Circulating monocyte-derived macrophages (MDMø) rapidly invade the brain after stroke, exerting both detrimental and beneficial effects. Elucidating mechanisms that mediate detrimental properties of MDMø may identify therapeutic strategies to divert MDMø from destructive phenotypes, while preserving their favorable effects. Toward this goal, the current study explores the function of Galectin-3 (GAL3) in MDMø and elucidates mechanisms whereby MDMø-derived GAL3 exacerbates stroke injury. In the acutely injured brain, GAL3 expression was upregulated primarily within MDMø. Global KO of GAL3 reduced brain infarcts in the short term but did not sustain long-term positive outcomes. Using BM chimera mice, macrophage transplantation, and myeloid cell–specific GAL3-KO (LysMCre+/–Lgals3fl/fl) mice, we demonstrated that GAL3 in MDMø mediated acute infarct expansion after stroke. Coculturing brain lysate–treated, BM-derived macrophages (BMDMs) with oxygen glucose deprivation–challenged neurons induced neurotoxicity that was mitigated by the cell-permeable, selective GAL3 inhibitor TD139. GAL3 triggered cathepsin induction and lysosomal leakage in BMDMs, leading to inflammasome activation. Systemic and transient TD139 treatment in the acute injury phase reduced infarcts, tempered neuroinflammation, and improved long-term neurological outcomes. Therefore, MDMø-derived GAL3 represents a drug target that could be accessed in peripheral blood to potentially mitigate post-stroke brain injury.

Authors

Miao Wang, Zhentai Huang, Zhihong Du, Jiajing Shan, Qing Ye, Lingxiao Lu, Ming Jiang, Fei Xu, Ziyang Liu, David J.R. Fulton, Rehana K. Leak, Babak Razani, Jun Chen, Xiaoming Hu

×

Figure 6

High GAL3 expression in MDMø amplifies cathepsin induction and leakage after stroke, thereby enlarging brain lesions.

Options: View larger image (or click on image) Download as PowerPoint
High GAL3 expression in MDMø amplifies cathepsin induction and leakage a...
(A) Representative immunostaining of cathepsin B/GAL3/CD68 in WT/WT and WT/GAL3–/– chimera mice 3 d after tMCAO. Scale bar: 50 μm. (B) Quantification of cathepsin B+ staining areas within CD68+ areas of macrophages. (C) Quantification of CD68+ areas. N = 7/group. (D) MAP2 staining of brain in vehicle- or CA-074–treated (10 mg/kg, i.v., 2 h after tMCAO and then daily for 2 d) WT/WT or WT/GAL3–/– chimeric mice 3 d after tMCAO. (E) Quantification of brain infarct volume. N = 5–6/group. (F) Cathepsin B activity was analyzed using the Magic Red cathepsin kit in GAL3-KO or WT BMDMs with vehicle or TD139 (10 μM) treatment. Scale bar: 50 μm. (G) Quantification of the percentages of Magic Red+ cells among total cells. N = 6/condition. (H) Representative Magic Red staining in WT and GAL3-KO BMDMs treated with or without WT or GAL3-KO lysate for 6 h. Scale bars: 50 μm. (I) Quantification of the percentages of Magic Red+ cells among total cells. N = 6/condition. Gray blocks show SD. ***P < 0.001 versus (WT BMDM + WT lysate); #P < 0.05, ##P < 0.01 versus (KO BMDM + WT lysate); $P < 0.05 versus (WT BMDM + KO lysate). (J) Immunoblot and quantification of cathepsin B expression in cytosol of BMDMs under the indicated conditions for 6 h. N = 3/condition. (K) Coimmunostaining of LAMP2 and cathepsin B in BMDMs under specified conditions for 6 h. Scale bars: 5 μm, 2 μm (zoom). (L) Fluorescence intensity profiles of cathepsin B and LAMP2. (M) Pearson’s correlation analyses of LAMP2 and cathepsin B. N = 7/condition. (N) BMDMs in inserts treated with brain lysates and CA-074 (10 μM) or vehicle were cocultured with OGD neurons. Coverage areas of MAP2-stained neurons were quantified. N = 6/condition. Scale bar: 40 μm. *P < 0.05, **P < 0.01, ***P < 0.001. Two-tailed, unpaired Student’s t test (B, C, and E) or 1-way (G, J, M, and N) or 2-way (I) ANOVA and Bonferroni’s test.