The Temporal and Spatial Distribution of Macrophage Subpopulations During Arteriogenesis

Chronic arterial occlusion leads to growth of collaterals - a process termed arteriogenesis, in which macrophages play a prominent role in remodelling and growth. However, a detailed analysis which of distinct macrophage subpopulations involved in arteriogenesis has never been performed. In the present study the temporal and spatial distribution of macrophage subtypes during arteriogenesis in a rat model with chronically elevated fluid shear stress (FSS) is investigated. Local macrophage subpopulations were histologically immuno-phenotyped using CD68 (a ubiquitous macrophage marker) and CD163, a specific M2 macrophage marker. Without occlusion few M2-macrophages reside in the perivascular space. Early after occlusion (12h) the number of M2 macrophages increases strongly and M1 macrophages begin emerging into the collateral. After 3 days they appear in the perivascular space. Both macrophage subtypes increase until 28d after treatment, whereas M2 macrophages dominate at the site of collateral growth. The local distribution of the subpopulations changes during the arteriogenic process. Whereas M1 macrophages are detected directly adjacent to the media, M2 macrophages are present in the most outer perivascular region of the growing collateral vessel. Systemic alterations of blood leucocytes in mice after femoral artery ligature (FAL) were investigated by FACS analysis of serial blood samples. During collateral remodelling histological changes were not reflected in circulating monocytes in the peripheral blood. The activation state of macrophages in mice with FAL was modulated by injections of either dexamethasone or the interleukins IL10 or IL3/IL14. The arteriogenic response was assessed by hind limb perfusion with laser Doppler measurements after 3, 7 and 14d. Suppressing inflammatory monocyte subtypes (M1) with dexamethasone led to impaired perfusion recovery after FAL in mice, whereas IL10 or IL4/IL13 application significantly increased perfusion recovery. This investigation demonstrates that a forced shift towards M2 macrophages improves the arteriogenic response. The distinct early increase and spatial distribution of M2 macrophages support the idea that this subtype plays a predominant role during collateral remodelling. Keywords: Arteriogenesis, collateral artery, macrophages, activation type, arteriogenic response, dexamethasone