Interleukin-34-Dependent Perivascular Macrophages Promote Vascular Function in the Brain

Neurology Life Sciences Medicine Immunology
macrophages brain vasculature interleukin-34 perivascular macrophages vascular function

Academic Background

Macrophages in the central nervous system (CNS) include microglia and border-associated macrophages (BAMs). BAMs are distributed in the meninges, choroid plexus, and perivascular spaces, with perivascular macrophages (PVMs) being closely associated with cerebrovascular function. However, the mechanisms underlying BAM maintenance and their regulatory role in cerebrovascular function remain unclear.

Previous studies have shown that microglial development depends on colony-stimulating factor 1 (CSF-1), while their homeostasis in adulthood requires interleukin-34 (IL-34). Yet, whether IL-34 similarly regulates the survival and function of BAMs remains unknown. Additionally, how PVMs interact with vascular cells (e.g., vascular smooth muscle cells, pericytes) to modulate cerebral blood flow (CBF) and vasomotion lacks in-depth investigation.

This study, led by Melanie Greter’s team, aimed to elucidate:
1. The dependency of BAMs on CSF-1 and IL-34 at different developmental stages;
2. The cellular sources of IL-34 and its regulatory mechanisms for PVM homeostasis;
3. The impact of PVM depletion on cerebrovascular function.

Source of the Paper

  • Authors: Hannah Van Hove, Chaim Glück, Wiebke Mildenberger et al., from the Institute of Experimental Immunology, University of Zurich, among other institutions.
  • Corresponding Author: Melanie Greter (greter@immunology.uzh.ch).
  • Journal: Immunity (May 13, 2025, Volume 58).
  • DOI: 10.1016/j.immuni.2025.04.003.

Research Process and Results

1. BAM Development and Homeostasis Depend on CSF-1R Signaling

Experimental Design:
- Used conditional knockout mouse models (Cx3cr1CreER;Csf1rfl/fl) to delete the CSF-1 receptor (CSF-1R) during embryogenesis (E14.5 and E16.5) or adulthood via tamoxifen induction.
- Analyzed BAMs (marked as CX3CR1+CD206+ or CD11b+F4/80hi) and microglia (CX3CR1+CD206−) in embryonic (E18.5) and adult mouse brains using flow cytometry.

Results:
- Embryonic deletion of CSF-1R led to a significant reduction in BAMs and microglia (Figure 1A).
- Adult deletion of CSF-1R or treatment with the inhibitor PLX5622 similarly reduced both cell types (Figure 1B, S1D), indicating that CSF-1R signaling is essential for BAM development and maintenance.

2. IL-34 Is Critical for Adult BAM Homeostasis

Experimental Design:
- Analyzed BAM numbers in Il34LacZ/LacZ mice at different developmental stages.
- Subdivided BAM subsets via high-dimensional flow cytometry and immunofluorescence staining: PVMs (CD163+CD206+), MHC II+ BAMs, etc.

Results:
- IL-34 deficiency did not affect embryonic BAM development but significantly reduced BAM numbers in adulthood (Figure 1D).
- All BAM subsets (except dural macrophages) depended on IL-34 (Figures 2A-B).
- Immunofluorescence revealed near-complete loss of PVMs around arterioles in IL-34-deficient mice (Figures 2E-F).

3. Cellular Sources of IL-34: Perivascular Cells and Fibroblasts

Experimental Design:
- Analyzed IL34 expression in murine and human cerebrovascular cells using single-cell RNA sequencing (scRNA-seq).
- Specifically deleted IL-34 in mural cells and fibroblasts using Tbx18CreER;Il34fl/fl mice.

Results:
- IL-34 was primarily expressed by vascular smooth muscle cells (VSMCs), pericytes, and fibroblasts (Figures 3A, D).
- Deletion of perivascular IL-34 reduced PVMs but did not affect microglia (Figures 3F-G).
- IL-34 was similarly localized around blood vessels in human brain samples (Figure 3E).

4. IL-34 Deficiency Leads to Cerebrovascular Dysfunction

Experimental Design:
- Measured cerebral blood flow velocity using laser speckle contrast imaging (LSCI) and widefield localization microscopy (pia-flow).
- Observed vasomotion in awake mice via two-photon microscopy.

Results:
- IL-34-deficient mice exhibited a 15% increase in CBF and elevated flow velocity in penetrating arterioles (Figures 6B-D).
- Vasomotion amplitude and frequency were significantly enhanced (Figure 6I).
- Electron microscopy revealed structural abnormalities in arteriolar basement membranes and swollen astrocyte endfeet (Figures 5D, S4C).

5. Functional Validation via PVM-Specific Deletion

Experimental Design:
- Generated Mrc1Cre;Csf1rfl/fl mice to specifically delete CD206+ PVMs.

Results:
- PVM depletion similarly caused cerebrovascular hemodynamic abnormalities (Figures 7C-G), confirming that PVMs regulate vascular function independently of microglia.

Conclusions and Significance

  1. Developmental and Homeostatic Cytokine Division: BAM embryonic development relies on CSF-1, while adult homeostasis requires IL-34.
  2. Novel Cell-Cell Interaction Mechanism: Perivascular cells and fibroblasts maintain PVM survival via IL-34 secretion, forming a “vascular niche-macrophage” regulatory axis.
  3. Cerebrovascular Function Regulation: PVMs stabilize cerebrovascular homeostasis by suppressing vasomotion and modulating blood flow velocity.

Scientific Value:
- First demonstration of IL-34’s role in non-neuronal cells, expanding understanding of CNS macrophage niches.
- Provides new therapeutic targets for cerebrovascular diseases (e.g., hypertension, amyloid angiopathy).

Clinical Implications:
- Targeting the IL-34/PVM pathway may ameliorate cerebral blood flow disorders.
- Human IL-34 levels correlate with vascular dementia, highlighting clinical potential (Figure 3E).

Research Highlights

  1. Multidisciplinary Integration: Combined conditional gene knockout, high-dimensional flow cytometry, scRNA-seq, and live imaging to systematically dissect PVM regulation.
  2. Cross-Species Validation: Strengthened generalizability from mouse models to human brain samples.
  3. Innovative Discovery: Identified PVMs as direct regulators of cerebrovascular dynamics, challenging the traditional “neurovascular unit” framework.

Additional Information

  • Data Availability: scRNA-seq data uploaded to GEO (GSE292245, GSE292306, GSE292466).
  • Limitations: Further research is needed to determine whether IL-34 affects neurons or glia via other receptors (e.g., PTPRZ1).