Resident Macrophages in Tendon Growth and Development

Abstract

The tendon is a dense connective tissue with a highly organized extracellular matrix (ECM) that withstands regular tensile loads. Assembly and maintenance of the ECM—and therefore tendon function overall—is regulated by the cells that inhabit the tendon. The heterogeneity of tendon cells and their interactions with each other and the ECM is just beginning to be understood. In this dissertation, we focus on the relationship between two tendon cell types—the fibroblast and the resident macrophage—and their impact on the ECM. Resident macrophages populate nearly every tissue during normal physiology and have niche-specific functions during development and homeostasis. Their role in tendon, however, is poorly understood. To address this gap in knowledge, we begin by characterizing the spatiotemporal distribution and phenotypic profile of resident macrophages and their crosstalk with fibroblasts and the ECM during murine tendon development and maturation. We find that resident macrophages are regularly spaced within the fascicle and reside adjacent to fibroblasts from embryonic development into adulthood. We report an increase in the proportion of macrophages during early postnatal development that is driven by local proliferation. We define surface and transcriptional markers for tendon resident macrophages and find differences in both pro-inflammatory and anti-inflammatory cytokine expression with age. Next, we demonstrate a spatial relationship between Csf1-expressing fibroblasts and Csf1r-expressing resident macrophages. Macrophage colony stimulating factor (M-CSF), also known as CSF1, is an essential growth factor for the differentiation and survival of most resident macrophage populations. To test the dependence of tendon resident macrophages on fibroblast-derived CSF1 , we generate Scx^Cre;Csf1^flox/flox conditional knockout mice. We find that the presence of resident macrophages in the tendon requires fibroblast CSF1 expression. Unexpectedly, we find that cell organization, tissue morphology, and biomechanical properties appear normal in the absence of resident macrophages in young adult mice. Yet, we also find an altered collagen fibril distribution, with larger diameter fibrils in the knockout tendons. Together, this work establishes the tendon-specific profile of resident macrophages and the requirement of tendon fibroblast signaling in maintaining resident macrophages during development and growth.