Limits...
Macrophages retain hematopoietic stem cells in the spleen via VCAM-1.

Dutta P, Hoyer FF, Grigoryeva LS, Sager HB, Leuschner F, Courties G, Borodovsky A, Novobrantseva T, Ruda VM, Fitzgerald K, Iwamoto Y, Wojtkiewicz G, Sun Y, Da Silva N, Libby P, Anderson DG, Swirski FK, Weissleder R, Nahrendorf M - J. Exp. Med. (2015)

Bottom Line: Yet regulation of hematopoietic stem cell (HSC) activity in the spleen is incompletely understood.Nanoparticle-enabled in vivo RNAi silencing of the receptor for macrophage colony stimulation factor (M-CSFR) blocked splenic macrophage maturation, reduced splenic VCAM-1 expression and compromised splenic HSC retention.When we silenced either VCAM-1 or M-CSFR in mice with myocardial infarction or in ApoE(-/-) mice with atherosclerosis, nanoparticle-enabled in vivo RNAi mitigated blood leukocytosis, limited inflammation in the ischemic heart, and reduced myeloid cell numbers in atherosclerotic plaques.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Systems Biology, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114 mnahrendorf@mgh.harvard.edu dutta.partha@mgh.harvard.edu.

Show MeSH

Related in: MedlinePlus

Macrophages retain splenic HSCs via VCAM-1. (A) Flow cytometric plots showing VCAM-1 expression on splenic endothelial cells and macrophages after VCAM-1 knockdown. Flow cytometric plots show percentage of LSKs, HSCs, and GMPs in the spleen (B) and blood (C) of mice treated with siRNA against VCAM-1 (siVCAM-1) or control siRNA (siCON). Levels of LSKs, HSCs, and GMPs in the spleen (B), blood (C), and bone marrow (D; n = 8–10). Myeloid cells and monocytes in the blood (E) and spleen (F) after siVCAM-1 treatment in LPS-challenged mice (n = 4). (G) Reduced HSPC retention in the spleen after VLA-4 neutralization (n = 4–5). Two independent experiments were performed. Data are mean ± SEM. Significance was determined by Mann-Whitney test. *, P < 0.05; **, P < 0.01.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
getmorefigures.php?uid=PMC4387283&req=5

fig6: Macrophages retain splenic HSCs via VCAM-1. (A) Flow cytometric plots showing VCAM-1 expression on splenic endothelial cells and macrophages after VCAM-1 knockdown. Flow cytometric plots show percentage of LSKs, HSCs, and GMPs in the spleen (B) and blood (C) of mice treated with siRNA against VCAM-1 (siVCAM-1) or control siRNA (siCON). Levels of LSKs, HSCs, and GMPs in the spleen (B), blood (C), and bone marrow (D; n = 8–10). Myeloid cells and monocytes in the blood (E) and spleen (F) after siVCAM-1 treatment in LPS-challenged mice (n = 4). (G) Reduced HSPC retention in the spleen after VLA-4 neutralization (n = 4–5). Two independent experiments were performed. Data are mean ± SEM. Significance was determined by Mann-Whitney test. *, P < 0.05; **, P < 0.01.

Mentions: Our data suggest that splenic macrophages retain progenitors via expression of the adhesion molecule VCAM-1. To test this hypothesis directly, we formulated siRNA that targets VCAM-1 within macrophage-avid lipidoid nanoparticles (sense, AcuGGGuuGAcuuucAGGudTsdT; anti-sense, ACCUGAAAGUcAACCcAGUdTsdT). This treatment, which limited VCAM-1 protein concentrations in macrophages but not in endothelial cells (Fig. 6 A), reduced splenic progenitor retention (Fig. 6 B). In contrast, numbers of HSCs, LSKs, and GMPs increased significantly in the circulation (Fig. 6 C), indicating mobilization of these cells into the blood after VCAM-1 knockdown in macrophages. In the bone marrow, HSPC numbers did not change significantly after VCAM-1 knockdown in macrophages (Fig. 6 D), pointing to retention mechanisms that compensate for reduced VCAM-1 levels in the bone marrow. Reduced splenic HSC retention may attenuate LPS-induced myelopoiesis. To test this hypothesis, we enumerated myeloid cells and monocytes in the blood (Fig. 6 E) and spleen (Fig. 6 F) and found that VCAM-1 knockdown significantly reduced myeloid cell and monocyte numbers. Collectively, these data demonstrate a requirement for VCAM-1 expression by splenic macrophages for maintaining the organ’s hematopoietic niche. VLA-4 is an integrin expressed by leukocytes that binds with VCAM-1 on activated endothelial cells (Elices et al., 1990), resulting in recruitment of inflammatory cells to sites of inflammation. Similarly, VCAM-1 expressed by splenic macrophages may bind with VLA-4 expressed by HSCs (Williams et al., 1991). To investigate an interaction of VCAM-1 with VLA-4 in this setting, we transferred GFP+ HSPCs into LPS-treated mice after VLA-4 neutralization with an antibody. We found that this blocking antibody significantly decreased splenic HSPC retention (Fig. 6 G).


Macrophages retain hematopoietic stem cells in the spleen via VCAM-1.

Dutta P, Hoyer FF, Grigoryeva LS, Sager HB, Leuschner F, Courties G, Borodovsky A, Novobrantseva T, Ruda VM, Fitzgerald K, Iwamoto Y, Wojtkiewicz G, Sun Y, Da Silva N, Libby P, Anderson DG, Swirski FK, Weissleder R, Nahrendorf M - J. Exp. Med. (2015)

Macrophages retain splenic HSCs via VCAM-1. (A) Flow cytometric plots showing VCAM-1 expression on splenic endothelial cells and macrophages after VCAM-1 knockdown. Flow cytometric plots show percentage of LSKs, HSCs, and GMPs in the spleen (B) and blood (C) of mice treated with siRNA against VCAM-1 (siVCAM-1) or control siRNA (siCON). Levels of LSKs, HSCs, and GMPs in the spleen (B), blood (C), and bone marrow (D; n = 8–10). Myeloid cells and monocytes in the blood (E) and spleen (F) after siVCAM-1 treatment in LPS-challenged mice (n = 4). (G) Reduced HSPC retention in the spleen after VLA-4 neutralization (n = 4–5). Two independent experiments were performed. Data are mean ± SEM. Significance was determined by Mann-Whitney test. *, P < 0.05; **, P < 0.01.
© Copyright Policy - openaccess
Related In: Results  -  Collection

License 1 - License 2
Show All Figures
getmorefigures.php?uid=PMC4387283&req=5

fig6: Macrophages retain splenic HSCs via VCAM-1. (A) Flow cytometric plots showing VCAM-1 expression on splenic endothelial cells and macrophages after VCAM-1 knockdown. Flow cytometric plots show percentage of LSKs, HSCs, and GMPs in the spleen (B) and blood (C) of mice treated with siRNA against VCAM-1 (siVCAM-1) or control siRNA (siCON). Levels of LSKs, HSCs, and GMPs in the spleen (B), blood (C), and bone marrow (D; n = 8–10). Myeloid cells and monocytes in the blood (E) and spleen (F) after siVCAM-1 treatment in LPS-challenged mice (n = 4). (G) Reduced HSPC retention in the spleen after VLA-4 neutralization (n = 4–5). Two independent experiments were performed. Data are mean ± SEM. Significance was determined by Mann-Whitney test. *, P < 0.05; **, P < 0.01.
Mentions: Our data suggest that splenic macrophages retain progenitors via expression of the adhesion molecule VCAM-1. To test this hypothesis directly, we formulated siRNA that targets VCAM-1 within macrophage-avid lipidoid nanoparticles (sense, AcuGGGuuGAcuuucAGGudTsdT; anti-sense, ACCUGAAAGUcAACCcAGUdTsdT). This treatment, which limited VCAM-1 protein concentrations in macrophages but not in endothelial cells (Fig. 6 A), reduced splenic progenitor retention (Fig. 6 B). In contrast, numbers of HSCs, LSKs, and GMPs increased significantly in the circulation (Fig. 6 C), indicating mobilization of these cells into the blood after VCAM-1 knockdown in macrophages. In the bone marrow, HSPC numbers did not change significantly after VCAM-1 knockdown in macrophages (Fig. 6 D), pointing to retention mechanisms that compensate for reduced VCAM-1 levels in the bone marrow. Reduced splenic HSC retention may attenuate LPS-induced myelopoiesis. To test this hypothesis, we enumerated myeloid cells and monocytes in the blood (Fig. 6 E) and spleen (Fig. 6 F) and found that VCAM-1 knockdown significantly reduced myeloid cell and monocyte numbers. Collectively, these data demonstrate a requirement for VCAM-1 expression by splenic macrophages for maintaining the organ’s hematopoietic niche. VLA-4 is an integrin expressed by leukocytes that binds with VCAM-1 on activated endothelial cells (Elices et al., 1990), resulting in recruitment of inflammatory cells to sites of inflammation. Similarly, VCAM-1 expressed by splenic macrophages may bind with VLA-4 expressed by HSCs (Williams et al., 1991). To investigate an interaction of VCAM-1 with VLA-4 in this setting, we transferred GFP+ HSPCs into LPS-treated mice after VLA-4 neutralization with an antibody. We found that this blocking antibody significantly decreased splenic HSPC retention (Fig. 6 G).

Bottom Line: Yet regulation of hematopoietic stem cell (HSC) activity in the spleen is incompletely understood.Nanoparticle-enabled in vivo RNAi silencing of the receptor for macrophage colony stimulation factor (M-CSFR) blocked splenic macrophage maturation, reduced splenic VCAM-1 expression and compromised splenic HSC retention.When we silenced either VCAM-1 or M-CSFR in mice with myocardial infarction or in ApoE(-/-) mice with atherosclerosis, nanoparticle-enabled in vivo RNAi mitigated blood leukocytosis, limited inflammation in the ischemic heart, and reduced myeloid cell numbers in atherosclerotic plaques.

View Article: PubMed Central - HTML - PubMed

Affiliation: Center for Systems Biology, Massachusetts General Hospital and Harvard Medical School, Boston, MA 02114 mnahrendorf@mgh.harvard.edu dutta.partha@mgh.harvard.edu.

Show MeSH
Related in: MedlinePlus