Macrophages regulate colonic crypt renewal

Colonic epithelial cells are constantly insulted by luminal contents. The self-renewing, single epithelial layer conserves its barrier function through Lgr5+ epithelial stem cells located at the base of epithelial invaginations called crypts. Crypt stem cells first proliferate, then differentiate (giving rise to enterocytes, goblet cells; enteroendocrine cells and Tuft cells) and migrate along the crypt-axis where they are shed into the crypt lumen. Skoczek et al. 2014, previously showed that monocytes, (a macrophage precursor) induce colonic crypt proliferation and regulate the number of Lgr5+ stem cells both in vivo and in vitro. During health and inflammation, differential macrophage activity and functions have previously been characterised in vivo and in vitro, however, the effect of macrophages on crypt renewal and the cellular interactions at the crypt stem cell niche during homeostasis is yet to be fully understood. The aim of this study was to determine the effect of macrophages on colonic crypt renewal, in particular, its influence on epithelial proliferation and differentiation using a 3D macrophage-crypt co-culture model. This study demonstrates that colonic crypt proliferation significantly increases in the presence of non-activated (Naive), pro-inflammatory (M1) and anti-inflammatory macrophages (M2). Here, Naive and M2 macrophages require crypt-macrophage contact to trigger epithelial proliferation and pro-inflammatory macrophages (M1) can induce crypt proliferation via a physical contact and secretory factor-dependent pathway. Differentiated colonic crypt cell numbers of UEA-1+ goblet cells, DCAMKL1+ tuft cells, Cro-A+ enteroendocrine and Lgr5+ stem cell numbers were maintained in the presence of anti-inflammatory (M2), however a decrease in UEA-1+ goblet cells and DCAMKL1+ tuft cells and an increase in Lgr5+ stem cell numbers was observed in crypts cultured with pro-inflammatory (M1) macrophages that was dependent on M1-crypt contact. Taken together, this thesis demonstrates that differentially polarised macrophages (Naive, M1 and M2) can regulate colonic crypt growth, differentiation and in particular suggest that the epithelial stem cell fate is influenced by pro-inflammatory (M1) macrophages when in contact with the colonic epithelium in vitro