Uncovering the role of macrophage/vascular crosstalk in tissue aging

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Billions of cells die every day in our bodies as a result of normal tissue turnover and environmental stress. Macrophages are an immune cell population that play a critical role in clearing this cellular debris and promoting tissue repair. Macrophages reside in distinct sub-tissue locations or niches, such as blood vessels or nerves, and are thought to support specialized tissue functions. There is a mounting evidence of a link between reduced blood vessel function and many age-associated diseases including diabetes, atherosclerosis, chronic obstructive pulmonary disease (COPD) and neurodegenerative conditions, such as Alzheimer’s disease. Similarly, macrophages show impaired phagocytic capacity in tissues with age-associated pathologies. However, the functional interplay between macrophages and blood vessels as well as how this crosstalk potentially breaks down during tissue aging remains unclear. Therefore, I aim to identify: 1) molecular signals that facilitate macrophage/vascular crosstalk, 2) the relative role of systemic and local factors in promoting aging of the vascular niche as well as 3) the molecular alterations that drive macrophage decline and vascular aging. Conventional methods have provided limited insights, due mainly to our inability to capture the spatial and temporal dynamics of this process under physiological conditions, such as in living animals. Therefore, I have set up intravital imaging approaches to visualize and manipulate macrophages and blood vessels in the skin of living mice. 1) First, by utilizing intravital microscopy, I will opto-genetically label and isolate cells from the skin vascular niche for single cell RNA-sequencing to identify signaling pathways with active ligand/receptor pair expression. From this, I will use a novel light-inducible Cre-recombinase to functionally interrogate the role of these signaling pathways in spatiotemporally-defined regions of the skin vasculature to track direct changes to macrophage behavior and vascular function. 2) Secondly, by performing intravital time-lapse imaging in both young and old mice, I will identify which cellular behaviors are altered in the vascular niche with age, including macrophage phagocytosis, vascular density, blood flow and vascular debris accumulation. Furthermore, I will perform heterochronic parabiosis to determine the relative role of the systemic milieu in modulating these functional changes in the vascular niche. 3) Finally, I will isolate cells from the vascular niche of young, middle- aged, and old mice and perform single cell RNA-sequencing to identify differential gene expression patterns of the vascular niche during aging. I will then use cell type-specific inducible Cre-recombinases to knock out genes in these pathways to test their potential roles in vascular aging. To date, the cellular and molecular mechanisms that alter macrophage and vascular function with age remain unknown. Work from this proposal will provide a basic understanding of the interplay between macrophages and the vascular niche as well as reveal cellular events that facilitate tissue aging. Identification of cellular and molecular processes responsible for macrophage functional decline with age may provide novel therapeutic targets to restore vascular health in the elderly.