Circadian variation in migration velocity in small intestinal epithelium

The variation in migration rates of cells within the small intestinal epithelium was studied over a 24-hr period at 3-hr intervals (migration of cells was studied independently for the crypts and the villi using the changing distributions of [3H]TdR labelled cells as an indicator of cell migration). Clear changes in the rates of cell movement were observed during a 24-hr period for both crypt and villus epithelium. The rates of cell migration in these two compartments did not correlate well with the exception of samples taken at 18.00 hours. At this time of day there appeared to be no cell movement at all in either crypts or villi. There was not a good correlation between the migration velocity throughout the day and the changes in the number of mitoses. It is proposed that mitotic rates do not directly govern migration rates but that the converse may be true. Further, the lack of correlation between crypt and villus migration rates at any time of day suggest that the mechanisms controlling all movement in these two regions of small intestinal epithelium may be different.     

Cell migration velocities in the crypts of the small intestine after cytotoxic insult are not dependent on mitotic activity

The role of mitotic activity in the normal process of intestinal epithelial cell migration was investigated. The movement of [3H]TdR-labelled cells in the crypt-villus column was used to study migration both in the crypts and on the villi. Radiation alone or in conjunction with other cytotoxic agents (hydroxyurea, cyclophosphamide and isopropyl-methane sulphonate) was used to eliminate cell division activity and to decrease crypt cellularity. This was done in order to determine the role of 'mitotic pressure' in driving cell migration. It has been clearly demonstrated in this study that cell migration, both within the crypts and on the villi, can take place in the complete absence of mitotic activity and after a drastic decrease in crypt cellularity. These results add to the continually mounting evidence against the idea that the 'pressure' generated by mitoses within the crypt or indeed in other epithelial regions is responsible for propelling epithelial cells. The data also demonstrate that the migration mechanisms are resistant to cytotoxic exposure.     

Cell Migration Velocities In the Crypts of the Small Intestine After Cytotoxic Insult Are Not Dependent On Mitotic Activity

Abstract. The role of mitotic activity in the normal process of intestinal epithelial cell migration was investigated. the movement of [³H]TdR-labelled cells in the crypt-villus column was used to study migration both in the crypts and on the villi. Radiation alone or in conjunction with other cytotoxic agents (hydroxyurea, cyclophosphamide and isopropyl-methane sulphonate) was used to eliminate cell division activity and to decrease crypt cellularity. This was done in order to determine the role of 'mitotic pressure' in driving cell migration.
It has been clearly demonstrated in this study that cell migration, both within the crypts and on the villi, can take place in the complete absence of mitotic activity and after a drastic decrease in crypt cellularity. These results add to the continually mounting evidence against the idea that the 'pressure' generated by mitoses within the crypt or indeed in other epithelial regions is responsible for propelling epithelial cells. the data also demonstrate that the migration mechanisms are resistant to cytotoxic exposure.     

Effects of puromycin, cycloheximide and noradrenaline on cell migration within the crypts and on the villi of the small intestine. A model to explain cell movement in both regions

The normal process of cell migration, occurring as part of the replacement scheme within the small intestinal epithelium, was investigated extensively. The effects of puromycin, cycloheximide and noradrenaline on the movement of tritiated thymidine [( 3H]TdR) prelabelled crypt or villus cells have been studied. These studies have led to the formulation of a model for the mechanism of cell migration, postulating that the crypts and villi behave as separate units, with regard to cell migration, in addition to their distinct structural and functional properties. It is proposed that crypt cell migration is an active process requiring protein synthesis and protein glycosylation, whilst movement of villus epithelial cells is passive, depending on the continued contraction of smooth muscle cells in the lamina propria.     

Effects of Puromycin, Cycloheximide and Noradrenaline On Cell Migration Within the Crypts and On the Villi of the Small Intestine

Abstract. The normal process of cell migration, occurring as part of the replacement scheme within the small intestinal epithelium, was investigated extensively. the effects of puromycin, cycloheximide and noradrenaline on the movement of tritiated thymidine ([³H]TdR) prelabelled crypt or villus cells have been studied. These studies have led to the formulation of a model for the mechanism of cell migration, postulating that the crypts and villi behave as separate units, with regard to cell migration, in addition to their distinct structural and functional properties. It is proposed that crypt cell migration is an active process requiring protein synthesis and protein glycosylation, whilst movement of villus epithelial cells is passive, depending on the continued contraction of smooth muscle cells in the lamina propria.     

Cell migration pathway in the intestinal epithelium: an in situ marker system using mouse aggregation chimeras

The cell migration pathway in the intestinal epithelium of DDK in equilibrium C57BL/6JLac mouse chimeras is demonstrated using Dolichos biflorus agglutinin-peroxidase as strain-specific marker. Cell sheets of one genotype extend in relatively straight lines from crypt to villus apex. Narrow sheets are mostly interrupted in the distal two-thirds of duodenal but not ileal villi, suggesting that in the duodenum cell loss occurs below the apical extrusion zone. These differences between duodenum and ileum correspond to differences in villus shape. The pattern of cell migration in Peyer's patch epithelium is consistent with that of the duodenum. In chimeric colon, sharply demarcated territories of crypts with a narrow cuff of surface epithelium represent the counterpart of the villus/crypt unit of the small intestine.     

Cell migration and organization in the intestinal crypt using a lattice-free model

We present a novel class of spatial models of cell movement and arrangement applied to the two-dimensional cellular organization of the intestinal crypt. The model differs from earlier approaches in using a dynamic movement on a lattice-free cylindrical surface. Cell movement is a consequence of mitotic activity. Cells interact by viscoelastic forces. Voronoi tessellation permits simulations of individual cell boundaries. Simulations can be compared with experimental data obtained from cell scoring in sections. Simulation studies show that the model is consistent with the experimental results for the spatial distribution of labelling indices, mitotic indices and other observed phenomena using a fixed number of stem cells and a fixed number of transit cell divisions.     

Circadian Variation In Migration Velocity In Small Intestinal Epithelium

Abstract. The variation in migration rates of cells within the small intestinal epithelium was studied over a 24-hr period at 3-hr intervals (migration of cells was studied independently for the crypts and the villi using the changing distributions of [³H]TdR labelled cells as an indicator of cell migration).
Clear changes in the rates of cell movement were observed during a 24-hr period for both crypt and villus epithelium. the rates of cell migration in these two compartments did not correlate well with the exception of samples taken at 18.00 hours. At this time of day there appeared to be no cell movement at all in either crypts or villi. There was not a good correlation between the migration velocity throughout the day and the changes in the number of mitoses.
It is proposed that mitotic rates do not directly govern migration rates but that the converse may be true. Further, the lack of correlation between crypt and villus migration rates at any time of day suggest that the mechanisms controlling all movement in these two regions of small intestinal epithelium may be different.     

Coordination of cell differentiation and migration in mathematical models of caudal embryonic axis extension

Vertebrate embryos display a predominant head-to-tail body axis whose formation is associated with the progressive development of post-cranial structures from a pool of caudal undifferentiated cells. This involves the maintenance of active FGF signaling in this caudal region as a consequence of the restricted production of the secreted factor FGF8. FGF8 is transcribed specifically in the caudal precursor region and is down-regulated as cells differentiate and the embryo extends caudally. We are interested in understanding the progressive down-regulation of FGF8 and its coordination with the caudal movement of cells which is also known to be FGF-signaling dependent. Our study is performed using mathematical modeling and computer simulations. We use an individual-based hybrid model as well as a caricature continuous model for the simulation of experimental observations (ours and those known from the literature) in order to examine possible mechanisms that drive differentiation and cell movement during the axis elongation. Using these models we have identified a possible gene regulatory network involving self-repression of a caudal morphogen coupled to directional domain movement that may account for progressive down-regulation of FGF8 and conservation of the FGF8 domain of expression. Furthermore, we have shown that chemotaxis driven by molecules, such as FGF8 secreted in the stem zone, could underlie the migration of the caudal precursor zone and, therefore, embryonic axis extension. These mechanisms may also be at play in other developmental processes displaying a similar mode of axis extension coupled to cell differentiation.     

Epithelial cell kinetics in the descending colon of the rat

The influence of experimental bypass on the epithelial cell kinetics in the rat descending colon was studied. It was found that the number of cells per crypt was markedly reduced at 6 weeks after bypass. The percentage of labelled crypt cells, 1 h after 3HTdR, and the distribution of labelled cells in the crypt was normal. Also the life span of the epithelial cells was the same in control and bypassed colon. The response of crypt cell proliferation to ischaemia-induced cell loss in the bypassed descending colon was similar to the one previously described for normal descending colon. This indicates that the absence of the normal luminal contents does not result in a different response of colonic crypts to induced cell loss. Furthermore, it was found that the number of cells per crypt and the proliferative activity did not change in the transverse colon after temporary ischaemia of the bypassed descending colon. This indicates that the increase in crypt cell proliferation after ischaemia-induced cell loss is a local response.     

A model of the control of cellular regeneration in the intestinal crypt after perturbation based solely on local stem cell regulation

Abstract. The control mechanisms involved in regeneration of murine intestinal crypts after perturbations are presently not well understood. The existence of some feedback signals from the cells on the villus to the cells in the crypt has been suggested. However, some recent experimental data point to the fact that regeneration in the crypt starts very early after perturbation, at a time when the villus cell population has hardly changed. In particular, this early cell proliferative activity is seen specifically at the bottom of the crypt, i.e. in the presumed stem cell zone and furthest from the villus.
The objective of this study was to investigate whether a new concept of regulation operating solely at the stem cell level could explain the present mass of accumulated data on the post-irradiation recovery, which is an extensively studied perturbation from the experimental point of view. In order to check its validity, the new concept was formalized as a mathematical simulation model thus enabling comparison with experimental data. The model describes the cellular development from stem cells to the mature villus cells. As a basic feature it is assumed that the self-maintenance and the cell cycle activity of the stem cells are controlled by the number of these cells in an autoregulatory fashion. The essential features of the experimental data (i.e. the recovery with time and the consistency between different types of measurements) can be very well reproduced by simulations using a range of model parameters. Thus, we conclude that stem cell autoregulation is a valid concept which could replace the villus crypt feedback concept in explaining the early changes after irradiation when the damage primarily affects the crypt. The question of the detailed nature of the control process requires further investigation.     

Cytoarchitectural reorganization of rabbit colonic goblet cells during baseline secretion

Light and electron microscopy were coupled with point counting methods to quantitate shape and volume changes of goblet cells during their migration and maturation from the base of the crypt to the colonic surface epithelium in the rabbit. After differentiation, goblet cells attain a broad pyramidal configuration in the basal third of the crypt. The cells elongate and dramatically decrease in volume as they move into the surface epithelium. The distributions and volume fractions of organelles were found to vary considerably, depending on the location of the goblet cell in the epithelium. Mucin granules are initially synthesized throughout the cytoplasm, but become increasingly concentrated as the cell matures. Organelles involved in synthesis such as the Golgi apparatus and rough endoplasmic reticulum (RER) similarly attain a more concentrated arrangement as the cell moves up in the crypt. The mean cell volume decreases from 1,228.8 microns3 for cells in the basal third of the crypt to 541.3 microns3 for goblet cells on the surface. Most organelles decrease in proportion to this decrease, although a disproportionately large decrease in the RER was measured. When actual subcellular volumes are calculated, a net decrease in several subcellular compartments is detected. This loss of granules and organelles is accomplished by the continual synthesis and secretion of mucin granules. Cytoplasm and organelles become entrapped in the upward movement of granules towards the cell apex, become irretrievably isolated, and are sloughed into the crypt lumen. This process accounts for the decrease in cell volume and contributes to the altered cytoarchitecture of the cell.     

Increased apoptosis and accelerated epithelial migration following inhibition of hedgehog signaling in adaptive small bowel postresection

The intestinal epithelium undergoes a marked adaptive response following loss of functional small bowel surface area characterized by increased crypt cell proliferation and increased enterocyte migration from crypt to villus tip, resulting in villus hyperplasia and enhanced nutrient absorption. Hedgehog (Hh) signaling plays a critical role in regulating epithelial-mesenchymal interactions during morphogenesis of the embryonic intestine. Our previous studies showed that blocking Hh signaling in neonatal mice results in increased small intestinal epithelial crypt cell proliferation and altered enterocyte fat absorption and morphology. Hh family members are also expressed in the adult intestine, but their role in the mature small bowel is unclear. With the use of a model of intestinal adaptation following partial small bowel resection, the role of Hh signaling in the adult gut was examined by determining the effects of blocking Hh signaling on the regenerative response following loss of functional surface area. Hh-inactivating monoclonal antibodies or control antibodies were administered to mice that sustained a 50% intestinal resection. mRNA analyses of the preoperative ileum by quantitative real-time PCR revealed that Indian hedgehog was the most abundant Hh family member. The Hh receptor Patched was more abundant than Patched 2. Analyses of downstream targets of Hh signaling demonstrated that Gli3 was twofold more abundant than Gli1 and Gli2 and that bone morphogenetic protein (BMP)2 was most highly expressed compared with BMP1, -4, and -7. Following intestinal resection, the expression of Hh, Patched, Gli, and most BMP genes was markedly downregulated in the remnant ileum, and, in anti-Hh antibody-treated mice, expression of Patched 2 and Gli 1 was further suppressed. In Hh antibody-treated mice following resection, the enterocyte migration rate from crypt to villus tip was increased, and by 2 wk postoperation, apoptosis was increased in the adaptive gut. However, crypt cell proliferation, villus height, and crypt depth were not augmented. These data indicate that Hh signaling plays a role in adult gut epithelial homeostasis by regulating epithelial cell migration from crypt to villus tip and by enhancing apoptosis.     

Intestinal villus structure contributes to even shedding of epithelial cells

In the intestinal epithelium, proliferated epithelial cells ascend the crypts and villi and shed at the villus tips into the gut lumen. In this study, we theoretically investigate the roles of the villi on cell turnover. We present a stochastic model that focuses on the duration over which cells migrate the shortest paths between the crypt orifices and the villus tips, where shedding cells are randomly chosen from among those older than the shortest-path cell migration times. By extending the length of the shortest path to delay cell shedding, the finger-like shape of the villus would tightly regulate shedding-cell ages compared with flat surfaces and shorter projections; the villus allows epithelial cells to shed at around the same age, which limits them from shedding early or staying in the epithelium for long periods. Computational simulations of cell dynamics agreed well with the predictions. We also examine various mechanical conditions of cells and confirm that coordinated collective cell migration supports the predictions. These results suggest the important roles of the villi in homeostatic maintenance of the small intestine, and we discuss the applicability of our approach to other tissues with collective cell movement.     

Emerging role for nuclear rotation and orientation in cell migration

Nucleus movement, positioning, and orientation is precisely specified and actively regulated within cells, and it plays a critical role in many cellular and developmental processes. Mutation of proteins that regulate the nucleus anchoring and movement lead to diverse pathologies, laminopathies in particular, suggesting that the nucleus correct positioning and movement is essential for proper cellular function. In motile cells that polarize toward the direction of migration, the nucleus undergoes controlled rotation promoting the alignment of the nucleus with the axis of migration. Such spatial organization of the cell appears to be optimal for the cell migration. Nuclear reorientation requires the cytoskeleton to be anchored to the nuclear envelope, which exerts pulling or pushing torque on the nucleus. Here we discuss the possible molecular mechanisms regulating the nuclear rotation and reorientation and the significance of this type of nuclear movement for cell migration.     

Emerging role for nuclear rotation and orientation in cell migration

Nucleus movement, positioning, and orientation is precisely specified and actively regulated within cells, and it plays a critical role in many cellular and developmental processes. Mutation of proteins that regulate the nucleus anchoring and movement lead to diverse pathologies, laminopathies in particular, suggesting that the nucleus correct positioning and movement is essential for proper cellular function. In motile cells that polarize toward the direction of migration, the nucleus undergoes controlled rotation promoting the alignment of the nucleus with the axis of migration. Such spatial organization of the cell appears to be optimal for the cell migration. Nuclear reorientation requires the cytoskeleton to be anchored to the nuclear envelope, which exerts pulling or pushing torque on the nucleus. Here we discuss the possible molecular mechanisms regulating the nuclear rotation and reorientation and the significance of this type of nuclear movement for cell migration.     

Renewal of the epithelium in the descending colon of the mouse. IV. Cell population kinetics of vacuolated-columnar and mucous cells.

Proliferation and migration of cells in the vacuolated-columnar and mucous cell lines were studied in the descending colon of adult female mice given a single injection or a continuous infusion of 3H-thymidine and killed at various intervals from one hour to 12 days. This investigation was carried out using one mum-thick Epon sections which were radioautographed after staining with the periodic acid-Schiff technique and iron-hematoxylin. In the normalized crypts with ten equal segments, labeled vacuolated cells at one hour after injection of 3H-thymidine were encountered in the lower four segments and in decreasing numbers in segments 5 through 7. From the percent labeled cells in segments of the crypt, the birth rate and fluxes of cells were computed. Moreover, it was found that a cell in the vacuolated-columnar cell line would undergo three mitotic cycles on the average from its birth at the cryptal base to its extrusion from the surface; of these three cycles, the last one which took place from segment 3 to segment 7 appeared to be a changeover from dividing cells to non-dividing cells, in accordance with the "slow cut-off" model of Cairnie et al. ('65b). Mucous cells located in segments 1 through 6 of the crypt were capable of incorporating 3H-thymidine and thus capable of undergoing mitosis. However, the rate of turnover of mucous cells based on proliferative rate was found to be much lower than the rate of turnover of mucous cells based on the transit time in the non-dividing segments of the crypt. Since there was a concomitant overproduction of cells in the vacuolated cells and newly formed mucous cells in the lower portion of the crypt, it was concluded that some vacuolated cells would give rise to mucous cells. This putative transformation occurred in the lower four segments of the crypt. Mucous cells which were formed by transformation would migrate upward along the cryptal wall and accumulate more mucus in the theca; in doing so, they would undergo two divisions, on the average, before they became non-dividing mucous cells. In ascending the cryptal walls, both vacuolated-columnar cells and mucous cells appeared to migrate at a similar speed; they moved much slower at the base of the crypt and accelerated toward the upper portion of the crypt, but they migrated at a constant speed in the non-dividing segments of the crypt.     

Vitamin A deficiency inhibits intestinal adaptation by modulating apoptosis,proliferation, and enterocyte migration

In a prior study, vitamin A-deficient rats subjected to submassive small bowel resections did not mount a normal intestinal adaptive response by 10 days postoperatively, although adaptive increases in crypt cell proliferation were not attenuated and there were no differences in apoptotic indexes. The present study was designed to address the mechanisms by which vitamin A status effects adaptation by analyzing proliferation, apoptosis, and enterocyte migration in the early postoperative period (16 and 48 h) in vitamin A-sufficient, -deficient, and partially replenished sham-resected and resected rats. At 16 h postresection, apoptosis was significantly greater in the remnant ileum of resected vitamin A-deficient rats compared with the sufficient controls. Crypt cell proliferation was increased by resection in all dietary groups at both timepoints. However, at 48 h postresection, proliferation was significantly decreased in the vitamin A-deficient and partially replenished rats. By 48 h after resection, vitamin A deficiency also reduced enterocyte migration rates by 44%. This occurred in conjunction with decreased immunoreactive collagen IV at 48 h and 10 days postoperation. Laminin expression was also reduced by deficiency at 10 days postresection, whereas fibronectin and pancadherin were unchanged at 48 h and 10 days. These studies indicate that vitamin A deficiency inhibits intestinal adaptation following partial small bowel resection by reducing crypt cell proliferation, by enhancing early crypt cell apoptosis, and by markedly reducing enterocyte migration rates, which may be related to changes in the expression of collagen IV and other extracellular matrix components.