Identification and isolation of splenic tissue-resident macrophage sub-populations by flow cytometry

Fujiyama, Satoshi; Nakahashi-Oda, Chigusa; Abe, Fumie; Wang, Yaqiu; Sato, Kenzo; Shibuya, Akira

doi:10.1093/intimm/dxy064

Cited by 48 publications

(39 citation statements)

References 20 publications

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“…The reciprocal contamination is also a significant issue, likely leading to false assignment of expression of macrophage-specific genes such as Csf1r to non-macrophages. Tissue digestion of spleen did not markedly change the splenic myeloid flow cytometry profile, compared to mechanical disruption (Fujiyama et al, 2019). Enzymatic tissue digestion does not prevent macrophage fragmentation, although it may facilitate isolation of subsets of intact macrophages from some tissues, including liver (Lynch et al, 2018) and lung (Becher et al, 2014;Misharin et al, 2013).…”

Section: Discussionmentioning

confidence: 80%

Fragmentation of macrophages during isolation confounds analysis of single cell preparations from mouse hematopoietic tissues

Millard

Heng

Opperman

et al. 2021

Preprint

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Mouse hematopoietic tissues contain abundant and heterogeneous populations of tissue-resident macrophages attributed trophic functions in control of immunity, hematopoiesis and bone homeostasis. A systematic strategy to characterise macrophage subsets in mouse bone marrow (BM), spleen and lymph node, unexpectedly revealed macrophage surface marker staining typically emanated from membrane-bound subcellular remnants associated with unrelated cell types. Remnant-restricted macrophage-specific membrane markers, cytoplasmic fluorescent reporters and mRNA were all detected in non-macrophage cell populations including isolated stem and progenitor cells. The profile of macrophage remnant association reflects adhesive interactions between macrophages and other cell types in vivo. Applying this knowledge, reduced macrophage remnant attachment to BM granulocytes in Siglec1 deficient mice was associated with compromised emergency granulocytosis, revealing a function for Siglec1-dependent granulocyte-macrophage interactions. Analysis of published RNA-seq data for purified macrophage and non-macrophage populations indicates that macrophage fragmentation is a general phenomenon that confounds bulk and single cell analysis of disaggregated tissues.

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Section: Discussionmentioning

confidence: 80%

Fragmentation of macrophages during isolation confounds analysis of single cell preparations from mouse hematopoietic tissues

Millard

Heng

Opperman

et al. 2021

Preprint

View full text Add to dashboard Cite

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“…The T-cells interact with B-cells, causing B-cell proliferation and differentiation into plasma cells that release immunoglobins. Monocytes, developed in the spleen, move to damaged tissues, and differentiate into macrophages that promote the healing process [1,4,[15][16][17][18][19][20][21][22]. There are at least two possibilities for their presence in the spleen.…”

Section: Discussionmentioning

confidence: 99%

Telomerase-Positive Stem Cells in Adult Porcine and Adult Rat Spleens I. Totipotent Stem Cells

Young¹,

Limnios²,

Lochner³

et al. 2020

JRMBR

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“…Mouse spleen cells were isolated following a previous description [ 17 ]. In detail, the spleen was first perfused in 10 mL phosphate buffered saline (PBS), which was then injected into left ventricular before dissection.…”

Section: Methodsmentioning

confidence: 99%

Acacetin ameliorates insulin resistance in obesity mice through regulating Treg/Th17 balance via MiR-23b-3p/NEU1 Axis

et al. 2021

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Background The role of miR-23b-3p in insulin resistance (IR) remained poorly understood. Methods After acacetin injection, obesity-induced IR model was constructed with or without miR-23b-3p upregulation and Neuraminidase 1 (NEU1) overexpression in mice. Body weight, serum metabolite and fat percent of the mice were measured. Tests on oral glucose and insulin tolerance were performed, and inflammatory cytokines C-reactive protein (CRP), Interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and monocyte chemoattractant protein 1 (MCP1) levels were quantified with enzyme-linked immunosorbent assay (ELISA). The binding sites between miR-23b-3p and NEU1 were predicted by TargetScan, and verified using dual-luciferase reporter assay. Relative expressions were detected with quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. Proportion of Treg and Th17 cells in total CD4+ T cells was detected with flow cytometry. Results MiR-23b-3p offset the effects of acacetin on body weight, fat percent, inflammatory cytokines levels and expressions of markers of regulatory T cells (Treg cells) and T helper 17 cells (Th17 cells), NEU1 and miR-23b-3p. NEU1 was a target of miR-23b-3p, and overexpressed NEU1 reversed the effects of upregulated miR-23b-3p on reducing Treg cells but increased body weight, fat percent and inflammatory cytokines levels, percentage of Th17 cells, and upregulated NEU1 expression. Conclusion Upregulation of miR-23b-3p offset the effects of acacetin on obesity-induced IR through regulating Treg/Th17 cell balance via targeting NEU1.The present findings provide a possible prevention strategy for obesity-induced IR.

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Identification and isolation of splenic tissue-resident macrophage sub-populations by flow cytometry

Abstract: Isolation of splenic macrophage subpopulations using an enzyme cocktail

Cited by 48 publications

References 20 publications

Fragmentation of macrophages during isolation confounds analysis of single cell preparations from mouse hematopoietic tissues

Fragmentation of macrophages during isolation confounds analysis of single cell preparations from mouse hematopoietic tissues

Telomerase-Positive Stem Cells in Adult Porcine and Adult Rat Spleens I. Totipotent Stem Cells

Acacetin ameliorates insulin resistance in obesity mice through regulating Treg/Th17 balance via MiR-23b-3p/NEU1 Axis

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