Abstract Body: Embryonic tissue resident macrophages (TRMs) are a distinct innate immune cell population that contribute to cardiac vascular development, mitochondrial homeostasis, and electrical conductance. While cardiac TRM functions in the developing heart and in the setting of disease have been characterized, their roles in early postnatal cardiac maturation and physiological remodeling are not fully elucidated. Understanding substates and functions of TRMs during postnatal angiogenesis, myocyte cell cycling, and adaptive hypertrophy may open up avenues for enhancing these processes by targeting the innate immune system.
A major obstacle in manipulating TRMs is marker profile heterogeneity. To overcome this, we leverage an orthogonal genetic approach targeting two genes highly expressed in TRMs, Cd169 and Cx3cr1. We hypothesize this will allow us to more selectively interrogate the TRMs of the heart. We performed both lineage tracing and cell ablation experiments. For lineage tracing, Siglec1^Cre mice (Cre expression driven by CD169) were crossed with mice expressing both CX3CR1^GFP and Cre-responsive Rosa26^TdT alleles to characterize TRM localization and morphology during the first week of postnatal life (p1-p7).
Preliminary analysis of the heart by confocal microscopy and by flow cytometry demonstrated robust GFP and TdTomato expression, with co-expression representing over 70% of labeled CD45⁺ cells across postnatal week one. In contrast, splenic CD45⁺ cells were primarily single CX3CR1-GFP⁺, suggesting that a dual CD169⁺ and CX3CR1⁺ targeted approach may more readily affect cardiac immune cell populations. To that end, we crossed Siglec1^Cre Rosa26^TdT mice with the CX3CR1^DTR mouse line to express the human diphtheria toxin receptor only in CD169⁺ CX3CR1⁺ cells. Mice were given IP injections of diphtheria toxin or vehicle at p1, p2, and p3 and then analyzed at p7. Preliminary results suggest a reduction in normalized heart weight (p=0.03, n=9) and slightly decreased vascular density (1.78 vs. 2.04 CD31⁺ vessels/myocyte; p=0.26, n=6) at p7. These genetic models suggest feasibility and a possible adverse effect of ablating CD169⁺ CX3CR1⁺ TRMs during postnatal heart maturation, and open new avenues for mapping TRM states and functions in this window.