Intact function of Lgr5 receptor-expressing intestinal stem cells in the absence of Paneth cells

Abstract

Lifelong self-renewal of the adult intestinal epithelium requires the activity of stem cells located in mucosal crypts. Lgr5 and Bmi1 are two molecular markers of crypt-cell populations that replenish all lineages over time and hence function as stem cells. Intestinal stem cells require Wnt signaling, but the understanding of their cellular niche is incomplete. Lgr5-expressing crypt base columnar cells (CBCs) reside deep in the crypt, mingled among mature Paneth cells that are well positioned for short-range signaling. Partial lineage ablation previously had implied that Paneth cells are nonessential constituents of the stem-cell niche, but recently their absence was reported to interfere with Lgr5 + CBCs, resurrecting an appealing idea. However, previous mouse models failed to remove Paneth cells completely or permanently; defining the intestinal stem-cell niche requires clarity with respect to the Paneth cell role. We find that Lgr5 + cells with stem-cell activity cluster in future crypts early in life, before Paneth cells develop. We also crossed conditional Atoh1 −/− mice, which lack Paneth cells entirely, with Lgr5 GFP mice to visualize Lgr5 + CBCs and to track their stemcell function. In the sustained absence of Paneth cells, Lgr5 + CBCs occupied the full crypt base, proliferated briskly, and generated differentiated progeny over many months. Gene expression in fluorescence-sorted Lgr5 + CBCs reflected intact Wnt signaling despite the loss of Paneth cells. Thus, Paneth cells are dispensable for survival, proliferation, and stem-cell activity of CBCs, and direct contact with Lgr5-nonexpressing cells is not essential for CBC function. S tem cells in selected adult tissues, such as the bone marrow, skin, and digestive tract, play a vital role in replenishing multiple cell types throughout life, and their unique and potent capacity for self-renewal is replicated in cancer (1). These stem cells occupy specialized niches and respond to the local environment (2). The functions of such niches range from delivering trophic signals that control cell proliferation and prevent stemcell depletion to preventing unrestrained cell replication (3). Defining the cellular and molecular constituents of adult stem-cell niches therefore is an important challenge in biology and medicine.Intestinal stem cells reside in mucosal crypts and generate four distinct cell types. Enterocytes, goblet cells, and enteroendocrine cells line deep crypts in the colon and villi that project into the small bowel lumen; Paneth cells lie at the crypt base in the small intestine, increasing in number from duodenum to ileum, but are absent from the colon (4). Two small intestine crypt-cell populations are able to generate all four cell types over extended periods: Lgr5-expressing crypt base columnar cells (CBCs), which lie deep in the crypt, interspersed among Paneth cells (5), and Bmi1-expressing cells that occupy several crypt tiers, most notably the +4 position (6). Although recent evidence suggests that each of these cell populations can engende...

Shp2/MAPK signaling controls goblet/paneth cell fate decisions in the intestine

“…Thus, the absence of paneth cells did not completely disrupt the stem cell niches. This confirms recent work that noted a decrease but not complete loss of Lgr5+ stem cells after genetic ablation of paneth cells (5,6). By contrast, in organoid cultures the interaction between the two cell types is crucial: Lgr5+ stem cells are only able to generate organoids in the presence of paneth cells (4,26).…”

“…By contrast, in organoid cultures the interaction between the two cell types is crucial: Lgr5+ stem cells are only able to generate organoids in the presence of paneth cells (4,26). This difference suggests that in vivo, additional cell types such as stromal components might produce factors that allow stem cell maintenance in the absence of paneth cells (5,6). Ablation of Shp2 could increase the numbers of Lgr5+ cells indirectly, i.e., by increasing the numbers of paneth cells that in turn produce factors that stimulate stem cells.…”

“…Lgr5 is a receptor for R-spondins and participates in canonical Wnt signaling (4). Lgr5+ stem cells are reduced but not depleted when paneth cells are lacking (5,6), which indicates that paneth cells are not the sole source of the signals that maintain stem cells.…”

ER stress transcription factor Xbp1 suppresses intestinal tumorigenesis and directs intestinal stem cells

“…Stable Xbp1 silencing resulted in profound Stat3 Tyr-705 phosphorylation compared with (Sato et al, 2011;Durand et al, 2012;Kim et al, 2012;Yilmaz et al, 2012), are morphologically condensed to Paneth cell remnants that lack their characteristic secretory apparatus when Xbp1 is deleted (Kaser et al, 2008). Among the genes that had been reported as most highly enriched in Paneth cells and that could support a niche function for ISCs are Wnt3, Wnt11, Egf, Tgfa, and Dll4 (Sato et al, 2011).…”

Untitled

“…This method enabled us to culture intestinal stem cells without a mesenchymal cellular niche, the essential mechanism for the differentiation and proliferation of intestinal stem cell incorporating Paneth cells [12,38,39]. The form of cultured stem cells became a round-shape called "spheroid'' and differed from the original shape.…”