Retrograde movements determine effective stem cell numbers in the intestine

The morphology and functionality of the epithelial lining differ along the intestinal tract, but tissue renewal at all sites is driven by stem cells at the base of crypts(1-3). Whether stem cell numbers and behaviour vary at different sites is unknown. Here we show using intravital microscopy that, despite similarities in the number and distribution of proliferative cells with an Lgr5 signature in mice, small intestinal crypts contain twice as many effective stem cells as large intestinal crypts. We find that, although passively displaced by a conveyor-belt-like upward movement, small intestinal cells positioned away from the crypt base can function as long-term effective stem cells owing to Wnt-dependent retrograde cellular movement. By contrast, the near absence of retrograde movement in the large intestine restricts cell repositioning, leading to a reduction in effective stem cell number. Moreover, after suppression of the retrograde movement in the small intestine, the number of effective stem cells is reduced, and the rate of monoclonal conversion of crypts is accelerated. Together, these results show that the number of effective stem cells is determined by active retrograde movement, revealing a new channel of stem cell regulation that can be experimentally and pharmacologically manipulated.Peer reviewe

Plasticity of Lgr5-Negative Cancer Cells Drives Metastasis in Colorectal Cancer

Colorectal cancer stem cells (CSCs) express Lgr5 and display extensive stem cell-like multipotency and self-renewal and are thought to seed metastatic disease. Here, we used a mouse model of colorectal cancer (CRC) and human tumor xenografts to investigate the cell of origin of metastases. We found that most disseminated CRC cells in circulation were Lgr5- and formed distant metastases in which Lgr5+ CSCs appeared. This p

In vivo visualization of intestinal stem cell and crypt dynamics

To cope with the hazardous environment in the lumen, the intestinal epithelium is renewed every 3-5 days. This rapid tissue turnover is fueled by stem and progenitor cells at the bottom of crypts – small invaginations into the intestinal wall – that proliferate approximately once a day. The proliferation pushes postmitotic cells upward towards the lumen where they get shed. In this thesis, we use intravital microscopy to visualize this highly dynamic intestinal epithelium in real time. We aim to understand the dynamics that underlie stem cell competition at the base of small and large intestinal crypts and how stem cell and crypt dynamics in the healthy epithelium can affect mutation accumulation and ultimately tumor initiation. In Chapter 1 we highlight how over the recent years live cell and intravital microscopy have contributed to the understanding of stem cell dynamics and plasticity during development, homeostasis, regeneration and tumor formation. In Chapter 2 we give our perspective on how intestinal stem cell and crypt dynamics minimize the retention of mutations in the intestinal epithelium by ensuring that most mutated cells get lost. In Chapter 3 we describe the differences in stem cell dynamics in small and large intestinal crypts. We find that the amount of retrograde movement of the cells within the crypt determines which cells can act as long-term stem cells, as this allows gaining a favorable position in the niche center. While in the small intestine we do observe retrograde movement resulting in functional stemness of cells further away from the crypt base, this retrograde movement is (near) absent in the large intestine. Therefore, only cells at the center of the crypt can function as long-term stem cells in the large intestine, while cells further away from the crypt base are destined to be lost. In Chapter 4 we investigate how Wnt ligands influence tumor initiation by controlling the number of intestinal stem cells. We show that when Wnt secretion is reduced using a Porcupine inhibitor, stem cells located further away from the crypt base are lost, resulting in a smaller pool of competing stem cells. When APC is deleted in this scenario, APC mutated stem cells can take over the crypt more rapidly, leading to accelerated tumorigenesis. In Chapter 5 we study the effects of enlarging the stem cell pool by a calorie restricted diet. We demonstrate that a larger stem cell pool induced by this diet results in a lower retention of mutated cells, since there are more wild-type stem cell competitors that can outcompete mutated cells. In Chapter 6 we uncover the previously unobserved phenomenon of crypt fusion. We show that in addition to crypt duplication through crypt fission, two crypts can fuse together to form one daughter crypt. We propose that crypt fission and crypt fusion can function as counterbalancing processes. In Chapter 7 I summarize the results described in this thesis, discuss the findings in the light of the current literature, and propose future research directions

In vivo visualization of intestinal stem cell and crypt dynamics

To cope with the hazardous environment in the lumen, the intestinal epithelium is renewed every 3-5 days. This rapid tissue turnover is fueled by stem and progenitor cells at the bottom of crypts – small invaginations into the intestinal wall – that proliferate approximately once a day. The proliferation pushes postmitotic cells upward towards the lumen where they get shed. In this thesis, we use intravital microscopy to visualize this highly dynamic intestinal epithelium in real time. We aim to understand the dynamics that underlie stem cell competition at the base of small and large intestinal crypts and how stem cell and crypt dynamics in the healthy epithelium can affect mutation accumulation and ultimately tumor initiation. In Chapter 1 we highlight how over the recent years live cell and intravital microscopy have contributed to the understanding of stem cell dynamics and plasticity during development, homeostasis, regeneration and tumor formation. In Chapter 2 we give our perspective on how intestinal stem cell and crypt dynamics minimize the retention of mutations in the intestinal epithelium by ensuring that most mutated cells get lost. In Chapter 3 we describe the differences in stem cell dynamics in small and large intestinal crypts. We find that the amount of retrograde movement of the cells within the crypt determines which cells can act as long-term stem cells, as this allows gaining a favorable position in the niche center. While in the small intestine we do observe retrograde movement resulting in functional stemness of cells further away from the crypt base, this retrograde movement is (near) absent in the large intestine. Therefore, only cells at the center of the crypt can function as long-term stem cells in the large intestine, while cells further away from the crypt base are destined to be lost. In Chapter 4 we investigate how Wnt ligands influence tumor initiation by controlling the number of intestinal stem cells. We show that when Wnt secretion is reduced using a Porcupine inhibitor, stem cells located further away from the crypt base are lost, resulting in a smaller pool of competing stem cells. When APC is deleted in this scenario, APC mutated stem cells can take over the crypt more rapidly, leading to accelerated tumorigenesis. In Chapter 5 we study the effects of enlarging the stem cell pool by a calorie restricted diet. We demonstrate that a larger stem cell pool induced by this diet results in a lower retention of mutated cells, since there are more wild-type stem cell competitors that can outcompete mutated cells. In Chapter 6 we uncover the previously unobserved phenomenon of crypt fusion. We show that in addition to crypt duplication through crypt fission, two crypts can fuse together to form one daughter crypt. We propose that crypt fission and crypt fusion can function as counterbalancing processes. In Chapter 7 I summarize the results described in this thesis, discuss the findings in the light of the current literature, and propose future research directions.

Expanding the Tissue Toolbox : Deriving Colon Tissue from Human Pluripotent Stem Cells

Organoid technology holds great potential for disease modeling and regenerative medicine. In this issue of Cell Stem Cell, Múnera et al. (2017) establish the generation of pluripotent stem cell-derived colon organoids that upon transplantation in mice, resembling human colon to a large extent, opening up avenues to study disease pathogenesis in human colon tissue. Organoid technology holds great potential for disease modeling and regenerative medicine. In this issue of Cell Stem Cell, Múnera et al. (2017) establish the generation of pluripotent stem cell-derived colon organoids that upon transplantation in mice, strikingly resemble human colon, opening up avenues to study disease pathogenesis in human colon tissue

Retrograde movements determine effective stem cell numbers in the intestine

The morphology and functionality of the epithelial lining differ along the intestinal tract, but tissue renewal at all sites is driven by stem cells at the base of crypts1–3. Whether stem cell numbers and behaviour vary at different sites is unknown. Here we show using intravital microscopy that, despite similarities in the number and distribution of proliferative cells with an Lgr5 signature in mice, small intestinal crypts contain twice as many effective stem cells as large intestinal crypts. We find that, although passively displaced by a conveyor-belt-like upward movement, small intestinal cells positioned away from the crypt base can function as long-term effective stem cells owing to Wnt-dependent retrograde cellular movement. By contrast, the near absence of retrograde movement in the large intestine restricts cell repositioning, leading to a reduction in effective stem cell number. Moreover, after suppression of the retrograde movement in the small intestine, the number of effective stem cells is reduced, and the rate of monoclonal conversion of crypts is accelerated. Together, these results show that the number of effective stem cells is determined by active retrograde movement, revealing a new channel of stem cell regulation that can be experimentally and pharmacologically manipulated

Retrograde movements determine effective stem cell numbers in the intestine

The morphology and functionality of the epithelial lining differ along the intestinal tract, but tissue renewal at all sites is driven by stem cells at the base of crypts(1-3). Whether stem cell numbers and behaviour vary at different sites is unknown. Here we show using intravital microscopy that, despite similarities in the number and distribution of proliferative cells with an Lgr5 signature in mice, small intestinal crypts contain twice as many effective stem cells as large intestinal crypts. We find that, although passively displaced by a conveyor-belt-like upward movement, small intestinal cells positioned away from the crypt base can function as long-term effective stem cells owing to Wnt-dependent retrograde cellular movement. By contrast, the near absence of retrograde movement in the large intestine restricts cell repositioning, leading to a reduction in effective stem cell number. Moreover, after suppression of the retrograde movement in the small intestine, the number of effective stem cells is reduced, and the rate of monoclonal conversion of crypts is accelerated. Together, these results show that the number of effective stem cells is determined by active retrograde movement, revealing a new channel of stem cell regulation that can be experimentally and pharmacologically manipulated.Peer reviewe