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Alternatively activated brain-infiltrating macrophages facilitate recovery from collagenase-induced intracerebral hemorrhage.

Min H, Jang YH, Cho IH, Yu SW, Lee SJ - Mol Brain (2016)

Bottom Line: In the macrophage-depleted mice, ICH-induced brain lesion volume was larger and neurological deficits were more severe compared to those of control mice, indicating a protective role of these macrophages in ICH.Upon co-culture with glia, the number of mannose receptor-positive M2 macrophages was significantly increased.Furthermore, treatment with glia-conditioned media increased the number of BMDM of M2 phenotype.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience and Dental Research Institute, School of Dentistry, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul, 08826, Republic of Korea.

ABSTRACT

Background: Intracerebral hemorrhage (ICH) is one of the major causes of stroke. After onset of ICH, massive infiltration of macrophages is detected in the peri-hematoma regions. Still, the function of these macrophages in ICH has not been completely elucidated.

Results: In a collagenase-induced ICH model, CX3CR1(+) macrophages accumulated in the peri-hematoma region. Characterization of these macrophages revealed expression of alternatively activated (M2) macrophage markers. In the macrophage-depleted mice, ICH-induced brain lesion volume was larger and neurological deficits were more severe compared to those of control mice, indicating a protective role of these macrophages in ICH. In the ICH-injured brain, mannose receptor-expressing macrophages increased at a delayed time point after ICH, indicating M2 polarization of the brain-infiltrating macrophages in the brain microenvironment. To explore this possibility, bone marrow-derived macrophages (BMDM) were co-cultured with mouse brain glial cells and then tested for activation phenotype. Upon co-culture with glia, the number of mannose receptor-positive M2 macrophages was significantly increased. Furthermore, treatment with glia-conditioned media increased the number of BMDM of M2 phenotype.

Conclusions: In this study, our data suggest that brain-infiltrating macrophages after ICH are polarized to the M2 phenotype by brain glial cells and thereby contribute to recovery from ICH injury.

No MeSH data available.


Related in: MedlinePlus

Depletion of peripheral monocytes in ICH injury. a Schematic diagram of the myeloid cell depletion protocols using Cl2MDP. b Blood cells obtained from saline (black line)- or Cl2MDP-treated ICH mice (gray line) were stained with anti-CD11b-FITC antibody and analyzed with flow cytometry. c Cell suspensions obtained from saline- or Cl2MDP-treated mouse brain were stained with anti-CD11b-FITC and anti-CD45-PE antibody and analyzed with flow cytometry. Representative flow cytometry dot plots are shown. d ICH injured brain sections were collected and stained with anti-CD68 antibody. Representative images of three independent experiments are shown (Scale bar, 50 μm). e One day after Cl2MDP injection, mice were subjected to ICH, and the brains were sectioned and stained with cresyl violet at 3 dpi. Representative pictures are shown (Scale bar, 1 mm). f Hemorrhagic injured volumes were quantified and presented in a graph (n = 4 per group, *p < 0.05). g, h After Cl2MDP or saline i.p. injection, mice were subjected to ICH, and neurological outcomes were evaluated by focal deficit and sticky tape removal time at 1 and 3 days after ICH (n = 5 for saline and Cl2MDP groups in ART; n = 4 for saline group, n = 3 for Cl2MDP group in focal deficit, *p < 0.05). Data are expressed as mean ± SEM
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Fig3: Depletion of peripheral monocytes in ICH injury. a Schematic diagram of the myeloid cell depletion protocols using Cl2MDP. b Blood cells obtained from saline (black line)- or Cl2MDP-treated ICH mice (gray line) were stained with anti-CD11b-FITC antibody and analyzed with flow cytometry. c Cell suspensions obtained from saline- or Cl2MDP-treated mouse brain were stained with anti-CD11b-FITC and anti-CD45-PE antibody and analyzed with flow cytometry. Representative flow cytometry dot plots are shown. d ICH injured brain sections were collected and stained with anti-CD68 antibody. Representative images of three independent experiments are shown (Scale bar, 50 μm). e One day after Cl2MDP injection, mice were subjected to ICH, and the brains were sectioned and stained with cresyl violet at 3 dpi. Representative pictures are shown (Scale bar, 1 mm). f Hemorrhagic injured volumes were quantified and presented in a graph (n = 4 per group, *p < 0.05). g, h After Cl2MDP or saline i.p. injection, mice were subjected to ICH, and neurological outcomes were evaluated by focal deficit and sticky tape removal time at 1 and 3 days after ICH (n = 5 for saline and Cl2MDP groups in ART; n = 4 for saline group, n = 3 for Cl2MDP group in focal deficit, *p < 0.05). Data are expressed as mean ± SEM

Mentions: To assess the role of these M2 macrophages, we depleted the monocyte cell population by treating with clodronate liposome (Cl2MDP) and then subjected mice to ICH injury. Upon Cl2MDP injection (i.p.), 97 % of blood monocytes were depleted from the blood (Fig. 3b). In the ICH-injured brain, the percentage of CD11b+/CD45high macrophages was increased to 8.1 % (saline-ICH) from 1.5 % (saline-sham) (Fig. 3c). However, in the Cl2MDP-treated mice brain, the macrophage population only increased to 1.6 % (Cl2MDP-ICH) from 0.9 % (Cl2MDP-sham). Meanwhile, the microglia (CD11b+/CD45low) populations were not significantly altered by Cl2MDP treatment (Fig. 3c). Decreased macrophage infiltration in the Cl2MDP-injected mouse brain was also confirmed by immunohistochemistry (Fig. 3d). The CD68+ macrophages in the ICH-injured brain were significantly reduced in Cl2MDP-treated mice (Fig. 3d). Cresyl violet staining of ipsilateral brain sections from Cl2MDP- or saline-treated mice showed that monocyte-depleted mice had larger infarct volumes after ICH compared to control mice (Fig. 3e and f). Consistent with the histological data, the Cl2MDP-treated mice displayed more severe motor function impairment as measured by the adhesive removal test (Fig. 3g) and focal deficits scoring (Fig. 3h) at 3 dpi. These findings indicate that brain-infiltrating M2 macrophages play a protective role by presumably facilitating recovery from ICH injury.Fig. 3


Alternatively activated brain-infiltrating macrophages facilitate recovery from collagenase-induced intracerebral hemorrhage.

Min H, Jang YH, Cho IH, Yu SW, Lee SJ - Mol Brain (2016)

Depletion of peripheral monocytes in ICH injury. a Schematic diagram of the myeloid cell depletion protocols using Cl2MDP. b Blood cells obtained from saline (black line)- or Cl2MDP-treated ICH mice (gray line) were stained with anti-CD11b-FITC antibody and analyzed with flow cytometry. c Cell suspensions obtained from saline- or Cl2MDP-treated mouse brain were stained with anti-CD11b-FITC and anti-CD45-PE antibody and analyzed with flow cytometry. Representative flow cytometry dot plots are shown. d ICH injured brain sections were collected and stained with anti-CD68 antibody. Representative images of three independent experiments are shown (Scale bar, 50 μm). e One day after Cl2MDP injection, mice were subjected to ICH, and the brains were sectioned and stained with cresyl violet at 3 dpi. Representative pictures are shown (Scale bar, 1 mm). f Hemorrhagic injured volumes were quantified and presented in a graph (n = 4 per group, *p < 0.05). g, h After Cl2MDP or saline i.p. injection, mice were subjected to ICH, and neurological outcomes were evaluated by focal deficit and sticky tape removal time at 1 and 3 days after ICH (n = 5 for saline and Cl2MDP groups in ART; n = 4 for saline group, n = 3 for Cl2MDP group in focal deficit, *p < 0.05). Data are expressed as mean ± SEM
© Copyright Policy - OpenAccess
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Fig3: Depletion of peripheral monocytes in ICH injury. a Schematic diagram of the myeloid cell depletion protocols using Cl2MDP. b Blood cells obtained from saline (black line)- or Cl2MDP-treated ICH mice (gray line) were stained with anti-CD11b-FITC antibody and analyzed with flow cytometry. c Cell suspensions obtained from saline- or Cl2MDP-treated mouse brain were stained with anti-CD11b-FITC and anti-CD45-PE antibody and analyzed with flow cytometry. Representative flow cytometry dot plots are shown. d ICH injured brain sections were collected and stained with anti-CD68 antibody. Representative images of three independent experiments are shown (Scale bar, 50 μm). e One day after Cl2MDP injection, mice were subjected to ICH, and the brains were sectioned and stained with cresyl violet at 3 dpi. Representative pictures are shown (Scale bar, 1 mm). f Hemorrhagic injured volumes were quantified and presented in a graph (n = 4 per group, *p < 0.05). g, h After Cl2MDP or saline i.p. injection, mice were subjected to ICH, and neurological outcomes were evaluated by focal deficit and sticky tape removal time at 1 and 3 days after ICH (n = 5 for saline and Cl2MDP groups in ART; n = 4 for saline group, n = 3 for Cl2MDP group in focal deficit, *p < 0.05). Data are expressed as mean ± SEM
Mentions: To assess the role of these M2 macrophages, we depleted the monocyte cell population by treating with clodronate liposome (Cl2MDP) and then subjected mice to ICH injury. Upon Cl2MDP injection (i.p.), 97 % of blood monocytes were depleted from the blood (Fig. 3b). In the ICH-injured brain, the percentage of CD11b+/CD45high macrophages was increased to 8.1 % (saline-ICH) from 1.5 % (saline-sham) (Fig. 3c). However, in the Cl2MDP-treated mice brain, the macrophage population only increased to 1.6 % (Cl2MDP-ICH) from 0.9 % (Cl2MDP-sham). Meanwhile, the microglia (CD11b+/CD45low) populations were not significantly altered by Cl2MDP treatment (Fig. 3c). Decreased macrophage infiltration in the Cl2MDP-injected mouse brain was also confirmed by immunohistochemistry (Fig. 3d). The CD68+ macrophages in the ICH-injured brain were significantly reduced in Cl2MDP-treated mice (Fig. 3d). Cresyl violet staining of ipsilateral brain sections from Cl2MDP- or saline-treated mice showed that monocyte-depleted mice had larger infarct volumes after ICH compared to control mice (Fig. 3e and f). Consistent with the histological data, the Cl2MDP-treated mice displayed more severe motor function impairment as measured by the adhesive removal test (Fig. 3g) and focal deficits scoring (Fig. 3h) at 3 dpi. These findings indicate that brain-infiltrating M2 macrophages play a protective role by presumably facilitating recovery from ICH injury.Fig. 3

Bottom Line: In the macrophage-depleted mice, ICH-induced brain lesion volume was larger and neurological deficits were more severe compared to those of control mice, indicating a protective role of these macrophages in ICH.Upon co-culture with glia, the number of mannose receptor-positive M2 macrophages was significantly increased.Furthermore, treatment with glia-conditioned media increased the number of BMDM of M2 phenotype.

View Article: PubMed Central - PubMed

Affiliation: Department of Neuroscience and Dental Research Institute, School of Dentistry, Seoul National University, 1 Gwanak-ro, Gwanak-gu, Seoul, 08826, Republic of Korea.

ABSTRACT

Background: Intracerebral hemorrhage (ICH) is one of the major causes of stroke. After onset of ICH, massive infiltration of macrophages is detected in the peri-hematoma regions. Still, the function of these macrophages in ICH has not been completely elucidated.

Results: In a collagenase-induced ICH model, CX3CR1(+) macrophages accumulated in the peri-hematoma region. Characterization of these macrophages revealed expression of alternatively activated (M2) macrophage markers. In the macrophage-depleted mice, ICH-induced brain lesion volume was larger and neurological deficits were more severe compared to those of control mice, indicating a protective role of these macrophages in ICH. In the ICH-injured brain, mannose receptor-expressing macrophages increased at a delayed time point after ICH, indicating M2 polarization of the brain-infiltrating macrophages in the brain microenvironment. To explore this possibility, bone marrow-derived macrophages (BMDM) were co-cultured with mouse brain glial cells and then tested for activation phenotype. Upon co-culture with glia, the number of mannose receptor-positive M2 macrophages was significantly increased. Furthermore, treatment with glia-conditioned media increased the number of BMDM of M2 phenotype.

Conclusions: In this study, our data suggest that brain-infiltrating macrophages after ICH are polarized to the M2 phenotype by brain glial cells and thereby contribute to recovery from ICH injury.

No MeSH data available.


Related in: MedlinePlus