# Meningeal Lymphatics Drives Macrophage Clearance via CCL2-CCR2 Axis After Cerebral Ischemia

## TL;DR

This study shows that meningeal lymphatic vessels help clear immune cells after stroke through a CCL2-CCR2 signaling pathway, and enhancing this process could improve stroke recovery.

## Contribution

The study identifies a novel CCL2-CCR2 axis mechanism for macrophage clearance via meningeal lymphatics after cerebral ischemia.

## Key findings

- Ischemia expands meningeal macrophages that interact with lymphatic endothelial cells via the CCL2-CCR2 axis.
- CCR2 inhibition impairs macrophage trafficking and worsens stroke outcomes, while VEGF-C improves lymphatic drainage and recovery.
- Enhancing meningeal lymphatic function combined with CCL2-CCR2 modulation shows therapeutic potential for stroke.

## Abstract

The mechanisms underlying meningeal lymphatic vessel (mLV)-mediated immune cell clearance after stroke remain unclear. Using a mouse middle cerebral artery occlusion model, we performed single-cell RNA sequencing to analyze post-ischemic meningeal macrophages. In vitro co-culture and CCR2 inhibition (RS504393) validated the CCL2-CCR2 axis between lymphatic endothelial cells and macrophages. Macrophage trafficking to mLVs and cervical lymph nodes was assessed by Evans Blue tracing and F4/80 immunofluorescence. We utilized VEGF-C to enhance meningeal lymphatic vessel function and concomitantly evaluated neurological deficits, brain edema, and neuroinflammation. Ischemia expanded meningeal macrophages, whose crosstalk with lymphatic endothelial cells relied on CCL2-CCR2 axis. CCR2 inhibition impaired macrophage trafficking to mLVs and cervical lymph nodes, worsening edema, motor deficits, and inflammation, whereas VEGF-C enhanced mLV drainage and improved outcomes. We identify a novel mechanism where in mLVs recruit macrophages via CCL2 for perivascular clearance post-ischemia. Combining VEGF-C with modulation of the CCL2-CCR2 axis presents a promising synergistic therapeutic strategy for stroke.