The trophic effect of dietary fibre is not associated with a change in total crypt number in the distal colon of rats

Cynthia S.M. Wong and Peter R. Gibson1,2

University of Melbourne Department of Medicine, The Royal Melbourne Hospital, Victoria, Australia and
1 Department of Gastroenterology, Box Hill Hospital, Box Hill, Victoria 3128, Australia


    Abstract
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Soluble fibres, such as guar gum, promote and wheat bran or methylcellulose protect from chemically induced colon carcinogenesis, relative to the effect of a fibre-free diet in rats. Mechanisms are poorly understood. Whereas all fibres are trophic to the colonic epithelium, the heterogeneity of effects on carcinogenesis may reflect different effects on the total number of crypts and, therefore, the size of the stem cell population. This study aimed to assess this hypothesis. Sprague–Dawley rats were fed one of fibre-free diets with or without 10% wheat bran, methylcellulose or guar gum for 4 weeks. The distal colons were stained with methylene blue and quantified for the number and density of crypts using an image analysis system. Epithelial proliferative kinetics was measured stathmokinetically. Methodology for quantifying crypts was valid and reproducible. Rats fed a fibre-free diet had atrophic distal colon, as shown by a decrease in crypt column height and a lower mitotic index. Fibre supplementation prevented the atrophy and was associated with crypt mouth areas that were 30–60% larger than those in the fibre-free group (P < 0.001, ANOVA), with the methylcellulose group being the largest (1.16 µm2). The crypt density of the fibre-free group was 16–19% greater than those in fibre fed groups (P + 0.006), due to the smaller size of the crypts. However, there was no difference in the total number of crypts across the four dietary groups (P > 0.1). Distal colons in all of the dietary groups contained ~105 crypts. In conclusion, although variation in the amount or type of dietary fibre exerts heterogeneous effects on the growth of the colonic epithelium and on colon carcinogenesis, the total number of crypts in the distal colon remains constant. It is, therefore, unlikely that fibres influence carcinogenic events by altering the size of the stem cell population.

Abbreviations: GG, guar gum; MC, methylcellulose; NF, no fibre; WB, wheat bran.


    Introduction
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
In rat models of colon cancer, dietary fibre influences the progression of carcinogenic process. Compared with the effects of diets containing no fibre, slowly fermented fibres, such as wheat bran (WB), or minimally fermented fibres, such as cellulose or methylcellulose, suppress carcinogenic events, whereas soluble fibres and resistant starch generally promote such events (1,2), although considerable variability has been reported [reviewed in ref. (3)]. In humans, the influence of fibres on colorectal carcinogenesis is more confusing as our understanding is largely dependent upon data from epidemiological studies. For example, some but not other studies suggest vegetable fibre to be more protective than cereal fibre (3). Multiple protective mechanisms have been postulated, including reduction of colonic transit, bulking of stool, binding to carcinogens and bile acids, and, via fermentation events, which reduce luminal pH and phenol production and increase epithelial exposure to short-chain fatty acids (3). In contrast, there is a paucity of explanations for the promotion of carcinogenesis observed in the rat model by soluble fibres. Changes to the metabolic activity of the expanded bacterial population induced by soluble fibres in association with substrate exhaustion have been suggested to lead to the synthesis of toxic metabolites (4,5). Evidence for this, however, is awaited. Little consideration has been given to the possibility that the growth-promoting effects of fibre might increase susceptibility of the colon to carcinogenesis. Promotion of growth leads to more cells undergoing mitosis and, therefore, an increase in the population of cells vulnerable to carcinogens.

Tumours arise in the colon due to mutations that occur in cells capable of establishing a clone within the epithelium. The majority of epithelial cells in the colon have a short life of ~6 days (6). They are born in the basal region of the crypt and the majority migrate to the surface compartment where death occurs. This short time span is unlikely to be long enough for a sufficient number of critical mutations to occur so that the clone can establish and spread. Stem cells, on the other hand, are self-renewing and capable of passing the mutations on to the next generations of cells through multiple rounds of division. The stem cell might, therefore, be the biologically most relevant target cell for carcinogens. If the colon has an expanded population of stem cells, the likelihood of mutations and subsequent clonal growth of abnormal cells would be greater.

Unfortunately, no markers are available by which stem cells can be recognized in the colon and, thus, their numbers cannot be quantified directly. It has been proposed that there is a threshold value for the number of stem cells each crypt is able to support (7,8). If this number exceeds its threshold, crypt fission will occur, resulting in expansion of the total number of stem cells. If the number of stem cells per crypt is indeed the controlling mechanism by which crypts divide, the number of crypts in the colon may act as a useful surrogate marker for the number of colonic epithelial stem cells.

There is limited understanding of factors that control crypt fission. It is clearly a regenerative response to epithelial injury, such as that occurring in association with mucosal inflammation (9) or following exposure to 1,2-dimethylhydrazine in the experimental situation (10). It has been postulated that the rate of crypt fission is also stimulated by increases in crypt size (11,12). As dietary fibre increases crypt size and cellularity, it may induce crypt fission, and indeed, McCullough et al. (13) demonstrated that soluble dietary fibre increased both the number of crypts per circumference and the number of branched crypts in the proximal colon of rats. However, the hypertrophic effect of epidermal growth factor is not associated with an increase in the rate of crypt fission in the rat colon (10).

This study aimed to examine the hypotheses that dietary fibre increases the total number of crypts in the distal colon and that fibres that promote colon carcinogenesis (soluble fibre) do so by differentially increasing the number of colonic crypts in the distal colon. This was addressed by comparing the effect of different types and amounts of fibre, which have well documented effects on colon carcinogenesis, on the total number of crypts in the distal colon of healthy rats. In order to achieve this, a method for quantifying the number of crypts in a whole specimen was developed.


    Materials and methods
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Animals
Forty male Sprague–Dawley rats at 150 ± 10 g were randomly divided into four groups of 10 rats and fed a no fibre (NF) diet with or without supplements of 10% dietary fibre as WB (Purina Health Foods Company, Wahgunyah, Victoria, Australia), methylcellulose (MC) (Dow Chemical Company, Midland, MI) or guar gum (GG) (Procol®, Polypro International, Minneapolis, MN) for 4 weeks (Table IGo). These diets were identical to those used in carcinogenic experiments previously (1). The rats were placed on wire-bottom cages for the duration of the study to prevent coprophagy and eating of sawdust. After 4 weeks, the rats were killed by CO2 asphyxiation and cervical dislocation. An i.p. injection of 1 mg/kg vincristine sulphate was given 3 h before death to arrest the colonic epithelial cells at metaphase. The experiment was approved by The Royal Melbourne Hospital Research Foundation Animal Ethics Committee and carried out in a registered animal facility in accordance to the guidelines set out in The Australian Code of Practice for the Care and Use of Animals for Scientific Purposes.


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Table I. Composition of the no fibre and fibre-supplemented diets
 
Upon death, the distal colon was removed, opened longitudinally, washed briefly with saline and laid flat on Hybond paper (Highland-C Super, 0.45 micron, Amersham, Castle Hill, NSW, Australia). The tissue was then fixed in 4% buffered formalin for at least 24 h.

Quantification of crypts in the distal colon
The colon was washed briefly in distilled water and stained with 0.1% methylene blue for 5 min. The tissue was then pinned on a board and immersed in distilled water. The number of crypts was quantified using a stereomicroscope (Leica MZ6, Heerbrugg, Switzerland) coupled to an image analysis system (Leica Q500MC, Leica Cambridge, Cambridge, UK) that was calibrated and set at a magnification of 120 times. The proximal end of the distal colon was taken at the site where the longitudinal mucosa began (14). The distal end of the colon was considered to be 4 cm from the anus. The distal colon was examined from a distal-to-proximal direction.

Before counting commenced, the length and width of the colon were measured and recorded. The tissue was marked at 1 cm intervals and for each centimetre of colon the number of crypts in three areas of 1.5x1.5 mm was counted (Figure 1Go) by tracing the outline of the crypts on a computer. To do this, a grid was designed to enable three random sections of 0.5x0.5 mm to be examined. The crypt counts were totalled up to produce a count for one 1.5x1.5 mm area. This procedure was repeated for three 1.5x1.5 mm areas per 1 cm of colon throughout the length of the colon. The total number of crypts per colon and crypt density were calculated as follows:


where: c = number of crypts in three 1.5x1.5 mm areas; n = number of counts made; l = length of distal colon; w = width of distal colon.



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Fig. 1. Quantification of crypts in the distal colon. Crypts were counted from the distal end of the distal colon in 1 cm intervals. Three random areas of 1.5 x 1.5 mm were counted to obtain a mean number of crypts per cm of colon. An enlargement of one 1.5 x 1.5 mm area is shown to illustrate how the number of crypts within the 1.5 x 1.5 mm area were quantified.

 
The area of the crypt mouth was estimated by tracing its outline using the image analyser, as shown in Figure 2Go. Crypts from four pieces of colon from each group were chosen at random and 60 cross-sectional areas of the crypt mouth were measured per whole distal colon.



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Fig. 2. Measuring the area of crypt mouth. Using a stereomicroscope coupled to an image analyser, the crypt mouth was outlined and the area of that crypt mouth measured and expressed as µm2 at a magnification of 120x.

 
In preliminary experiments, the number of crypts counted in three 1.5x1.5 mm areas did not differ from those counted in four or more areas. For example, the mean difference (95% confidence interval) of the values obtained from counting in three areas from those obtained from five areas was 0.71 (–1.05 – 2.47; Bland–Altman analysis). The method of counting was also reproducible. The median variance (interquartile range) between crypt counts in eight pieces of colon across four dietary groups assessed on two occasions on separate days (to eliminate possible bias introduced by memory) was 7.6% (4.2–13.5%).

Measurement of epithelial proliferative kinetics
After the crypt numbers were quantified, a 1.5 cm length of tissue was taken 1 cm from the distal end of colon for histological examination. The tissue was cut into 2 mm vertical strips, processed and embedded in paraffin wax. H&E sections were examined, and the crypt column height and proliferative indices were evaluated. Only longitudinal crypts that were cut completely from the lumen to the base were counted and 20 crypt columns were quantified (15). The total cell population was measured by the crypt column height (CCH), defined as the mean number of cells per crypt column. Cells arrested in metaphase, recognized by a dark staining nuclei displaced towards the lumen, were also counted and expressed as the mean number per crypt column. This provided an index of the rate of cell proliferation. The cell turnover was measured by mitotic index, which was defined by the proportion of crypt epithelial cells arrested at metaphase.

In order to ensure the reliability of the results obtained, proliferative indices were also assessed by another technician highly experienced in this technique (M.Fielding) in a sample of sections (n = 10). The results from two independent readers correlated well (r = 0.94, P = 0.001; linear regression) with a coefficient of variance (CV) of 5.5%.

Statistical analysis
Data were analysed using Minitab Release 10 (Minitab, State College, PA) and MedCalc version 6.16 (MedCalc Software, Mariakerke, Belgium). Comparison of the effects of diet on proliferative kinetics and cross-sectional area of crypt mouth across multiple (>2 groups) were analysed by one-way analysis of variance (ANOVA), choosing the Tukey’s pairwise comparison option. The Tukey’s test performed intergroup comparisons by analysing the interval between means of the two groups. Intervals of groups that showed significance were then analysed by two-tailed Student’s t-test to obtain a P value. A two-way ANOVA was used to analyse the effects of diet and length on crypt density across the dietary groups. When validating the method for quantifying crypts, the relationship between two parameters was analysed by the Bland–Altman analysis and a difference of measurements that was within 2 standard deviations were considered to be interchangeable (16). A P value of <=0.05 was considered statistically significant.


    Results
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Effects of diets on epithelial proliferative kinetics in the distal colon
The crypt column height differed across the dietary groups (P < 0.001, ANOVA) (Figure 3Go). Atrophy of colonic crypts was observed in the NF diet group, indicated by shorter crypts than those in the three fibre-fed groups. The mitotic index differed across the groups (P = 0.03), with that of the WB group being significantly higher than that in the NF group (P = 0.006, t-test). The number of metaphase arrests per crypt column was numerically greater in all of the fibre-fed groups when compared with the NF group and a significant difference was found across the groups (P = 0.004, ANOVA). An intergroup analysis showed that the WB group with a mean (95% CI) of 1.8 [1.4–2.2] metaphase arrests/CCH was the only fibre-fed group that had significantly more metaphase arrests per crypt column than the NF group (0.8 [0.6–1]) (P < 0.001, t-test). The MC- and GG-fed groups had 1.5 [1–2] and 1.1 [0.7–1.5] metaphase arrests/CCH, respectively.



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Fig. 3. Epithelial proliferative kinetics in the distal colon of the dietary groups of rats. Rats were fed one of the four diets: no fibre (NF), or NF supplemented with 10% (weight:weight) wheat bran (WB), methylcellulose (MC) or guar gum (GG) for 4 weeks. The crypt column height (shown as the bars) was defined as the number of colonic epithelial cells per crypt column, and the mitotic index ({blacklozenge}) calculated as the proportion of cells at metaphase to the total number of cells in 20 crypt columns. Both indices significantly differed across the dietary groups (P < 0.001 and P = 0.03, respectively, ANOVA). Data are expressed as mean ±95% CI.

 
Effects of diets on the cross-sectional area of the crypt mouth
Cross-sectional area of the crypt mouth differed across the dietary groups, as shown in Figure 4Go (P < 0.001, ANOVA). The smallest areas were observed in the NF diet group and the largest were observed in rats fed MC. All of the fibre-fed groups had significantly larger crypt mouth area, ranging from 30–60%, when compared with the NF group (P < 0.001 for all groups, t-test), but were not significantly different from each other.



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Fig. 4. The effect of diets on the area of the crypt mouth. Crypt mouth area differed across the four dietary groups—no fibre (NF), or NF supplemented with 10% (weight:weight) wheat bran (WB), methylcellulose (MC) or guar gum (GG) (P < 0.001, ANOVA). Crypt mouth areas were significantly greater in fibre-fed groups than those in the NF group (P < 0.001 for all three fibres, t-test). The results are expressed as mean ±95% CI of 60 cross-sectional areas per crypt from four pieces of colon from each group.

 
Effects of diets on the length and surface area of the distal colon
As shown in Table IIGo, the length and area of the distal colon differed across the dietary groups (P = 0.005 and P < 0.001, respectively, ANOVA). The mean length of the distal colons of rats fed NF diet was significantly shorter in comparison to the fibre-fed groups. The NF diet was also associated with smaller colonic surface area than those associated with diets supplemented with fibre and MC supplementation produced the largest area.


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Table II. The effects of diets on length and surface area of the distal colon, shown as mean (95% confidence intervals)
 
Effects of diets on crypt number and density in the distal colon
As shown in Figure 5AGo, crypt density significantly differed across the dietary groups (P = 0.006). The crypt density in the distal colon of the NF group was 16–19% greater than those of all of the fibre groups (P <= 0.01 for all three groups, t-test). There were no significant distal-proximal gradients of crypt density observed for any of the dietary groups (P > 0.1 for all four dietary groups, Figure 5BGo), but the site-specific crypt density did differ along the length of the distal colon when compared across the dietary groups (length: P = 0.003 and diet: P < 0.001, two-way ANOVA). This difference was due to a greater crypt density in the NF group than those of the three fibre groups (P < 0.001 for all groups, t-test).



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Fig. 5. The effect of diets on crypt density in the distal colon. The diets tested comprised no fibre (NF), or NF supplemented with 10% (weight:weight) wheat bran (WB), methylcellulose (MC) or guar gum (GG). (A) Crypt densities of the entire distal colon are shown for individual rats. Differences were observed across four dietary groups (P = 0.006, ANOVA) with the NF group containing more crypts per area than the other three groups (P >= 0.01, t-test). (B) Crypt densities shown according to the dietary group ({square} = NF, {blacktriangleup} = WB, {circ} = MC and {blacklozenge} = GG) along the length of the distal colon in 1 cm segments starting from the distal end. The crypt density did not differ significantly along the length of the colon for each dietary group (P < 0.1, ANOVA) but differences in crypt density along the length across the dietary groups were observed (P < 0.003 [length] and P < 0.001 [diet], two-way ANOVA). The results are expressed as mean ±95% CI.

 
The total number of crypts in the distal colon was calculated by correcting for the density for differences in surface area. As shown in Figure 6Go, means of 1.1–1.4x105 crypts were found. No differences in the total number of crypts were found across the dietary groups (P = 0.15).



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Fig. 6. The effect of diets on the total number of crypts in the distal colon. No differences were found in the total number of crypts in the distal colon across the four dietary groups of rats – no fibre (NF), or NF supplemented with 10% (weight:weight) wheat bran (WB), methylcellulose (MC) or guar gum (GG) (P = 0.15, ANOVA). The data are expressed as mean ±95% CI.

 

    Discussion
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 
Previous studies on the effects of dietary fibre on the size of the population of epithelial cells in the colon have focused on individual crypt cellularity and have ignored possible effects on the total number of crypts. The observation that dietary fibre could alter both rates of crypt fission and crypt density in the colon (13) suggested that fibre might increase the total population of crypts. If so, this would be important information in terms of risk of carcinogenesis, since an expanded number of crypts would increase the size of the cell population vulnerable to carcinogens.

In order to define the relationship between diet and total crypt number, a method to accurately quantify the number of crypts and to enable valid comparison across dietary groups required development. The application of image analysis to methylene blue-stained mucosa proved to be reproducible over time. Since diets may influence colonic length and surface area, unequivocal anatomical landmarks were needed to ensure accurate definition of the regions of the large bowel being studied. The distal colon was chosen for evaluation for three reasons. First, anatomical landmarks clearly define the distal colon. Secondly, the distal colon is the most relevant part of the colon in the rat model being studied since it is the site of most tumours induced by carcinogens. Thirdly, variations in the type and amount of fibre appeared to exert greater influence over the distal colon as opposed to the proximal colon and caecum (15,17,18).

The choice of dietary fibres for this experiment was based on their fermentability and on effects on colon carcinogenesis well documented previously in the same rat species in the same laboratory. Thus, GG is a fast fermenting fibre that promotes carcinogenesis whereas WB is more slowly fermented in the colon and suppresses carcinogenic events (1,19). MC is a non-fermentable fibre (20) that decreases tumour load (G.P.Young, unpublished observation).

This study showed as expected, that fibre deprivation led to atrophy of the distal colon as indicated by the colons in the NF group being smaller in both length and area. Crypts in the NF group were shorter and had fewer proliferating cells per crypt column when compared to the other three fibre-fed groups. Furthermore, the cross-sectional area of the crypt mouth in the NF group was smaller than that of the fibre-fed groups. This would reflect smaller epithelial cells, but reduced luminal volume may have contributed. As also anticipated from previous experience (15,21), all three fibres reversed the atrophy induced by the NF diet. The patterns of the responses were, however, different across the dietary groups. MC was associated with the greatest surface area and numerically with the most cellular crypts and largest crypt mouth area. WB was associated with numerically greatest proliferative activity, whereas GG had the least hypertrophic effect.

These differences may have related to the differing mechanisms by which the different fibres are likely to act. Fermentation of fibres delivers butyrate and other short-chain fatty acids to the colonic epithelium and, as well documented previously (2225), this delivery alone can reverse atrophy. In the rat model used in the present study, dietary WB leads to higher butyrate concentrations in the distal colon than do soluble fibres (1). This difference alone might explain the greater effect on epithelial proliferation and crypt hypertrophy than GG. The mechanisms by which methylcellulose induced its effects are clearly independent of short-chain fatty acid production as luminal levels are similar to or less than those associated with a NF diet (15,18). By virtue of its bulking effect, dietary MC changes the physical properties of the lumen, dilutes and possibly binds potentially injurious substances in the soluble compartment of the luminal contents, and may exert a cleansing effect that can be likened to the beneficial effects of saline enemas or distal colonic infusions on the epithelium and/or carcinogenesis (26,27). The greater colonic surface area induced by the MC diet was likely to reflect an expanded surface epithelial compartment, which might be considered a manifestation of a reduced rate of cell death and/or shedding due to a less harsh luminal environment.

Whether atrophy of crypts due to the lack of fibre is also associated with changes in the total number of crypts has not been examined previously. Crypt density in the distal colon of the NF groups was greater than that in the fibre-fed groups. This was not unexpected because the crypts in the NF group were smaller than the fibre-fed groups as indicated by the smaller cross-sectional area of the crypt mouths (Figure 5Go), allowing more crypts to be ‘packed’ into an area of tissue. However, the total number of crypts in the distal colon was similar across all of the dietary groups, indicating that variations in the type or quantity of fibre have no effect on the total crypt population. Although this result would seem at odds with those reported previously (13), the total number of crypts was not quantified in the earlier report and the study was limited to the proximal not distal colon.

As there are no markers enabling the quantification of stem cells, the significance of the constancy of the total number of crypts in the distal colon under several dietary conditions to the size of the stem cell population cannot be made directly. However, if the current belief that an increase in the number of stem cells in a crypt is the stimulus for crypt fission is correct, then the findings of this study suggest that fibres do not lead to an expansion of the stem cell population, despite stimulating growth within the crypt. These observations lend further support to the notion that crypt fission and epithelial kinetics in the crypt itself are independently controlled, as originally proposed by Park et al. (10) from their key observations on the effects of epidermal growth factor and 1,2-dimethylhydrazine on crypt size and the frequency of crypt fission in rats.

While the number of stem cells in the distal colon may not be overtly influenced by dietary fibre in a normal colonic environment, effects may be different during exposure to a carcinogen. Azoxymethane increases crypt density in the distal colon of rats (28) and, as discussed above, 1,2-dimethylhydrazine induces crypt fission. The effects of alterations in the amount and type of dietary fibre consumed under these conditions might be different and warrants further investigation.

In conclusion, total crypt numbers in the distal colon are not influenced by altering the type or amount of dietary fibre, despite the growth-promoting effects of fibres on the crypt epithelium. These observations supported the notion that cell proliferation and crypt fission are regulated independently. These results suggest that fibre is unlikely to promote or suppress carcinogenesis in the colon by changing the size of the stem cell population in the distal colon.


    Notes
 
2 To whom correspondence should be addressed Email: peter.gibson{at}med.monash.edu.au Back


    Acknowledgments
 
The authors would like to thank Marisa Fielding for her technical assistance in validating the method for crypt quantification. The Cancer Council of Victoria and the National Health and Research Council of Australia supported this work.


    References
 Top
 Abstract
 Introduction
 Materials and methods
 Results
 Discussion
 References
 

  1. McIntyre,A., Gibson,P.R. and Young,G.P. (1993) Butyrate production from dietary fibre and protection against large bowel cancer in a rat model. Gut, 34, 386–391.[Abstract]
  2. Young,G.P., McIntyre,A., Albert,V., Folino,M., Muir,J.G. and Gibson,P.R. (1996) Wheat bran suppresses potato starch—potentiated colorectal tumorigenesis at the aberrant crypt stage in a rat model. Gastroenterology, 110, 508–514.[ISI][Medline]
  3. Sengupta,S., Tjandra,J.J. and Gibson,P.R. (2001) Dietary fiber and colorectal neoplasia. Dis. Colon Rectum, 44, 1016–1033.[ISI][Medline]
  4. McBurney,M.I., Van Soest,P.J. and Jeraci,J.L. (1987) Colonic carcinogenesis: the microbial feast or famine mechanism. Nutr. Cancer, 10, 23–28.[Medline]
  5. Goodlad,R.A. (2001) Dietary fibre and the risk of colorectal cancer. Gut, 48, 587–589.[Free Full Text]
  6. Junqueira,L.C., Carneiro,J. and Kelley,R.O. (1992) Digestive tract. In Evans,C.F. (ed.), Basic Histology, 7th Edn. Appleton and Lange, Norwalk, CT, pp. 306–308.
  7. Loeffler,M., Bratke,T., Paulus,U., Li,Y.Q. and Potten,C.S. (1997) Clonality and life cycles of intestinal crypts explained by a state dependent stochastic model of epithelial stem cell organization. J. Theor. Biol., 186, 41–54.[CrossRef][ISI][Medline]
  8. Loeffler,M. and Grossmann,B. (1991) A stochastic branching model with formation of subunits applied to the growth of intestinal crypts. J. Theor. Biol., 150, 175–191.[ISI][Medline]
  9. Cheng,H., Bjerknes,M., Amar,J. and Gardiner,G. (1986) Crypt production in normal and diseased human colonic epithelium. Anat. Rec., 216, 44–48.[ISI][Medline]
  10. Park,H.S., Goodlad,R.A., Ahnen,D.J., Winnett,A., Sasieni,P., Lee,C.Y. and Wright,N.A. (1997) Effects of epidermal growth factor and dimethylhydrazine on crypt size, cell proliferation and crypt fission in the rat colon. Cell proliferation and crypt fission are controlled independently. Am. J. Pathol., 151, 843–852.[Abstract]
  11. Totafurno,J., Bjerknes,M. and Cheng,H. (1987) The crypt cycle. Crypt and villus production in the adult intestinal epithelium. Biophys. J., 52, 279–294.[Abstract]
  12. Bjerknes,M. (1986) A test of the stochastic theory of stem cell differentiation. Biophys. J., 49, 1223–1227.[Abstract]
  13. McCullough,J.S., Ratcliffe,B., Mandir,N., Carr,K.E. and Goodlad,R.A. (1998) Dietary fibre and intestinal microflora: effects on intestinal morphometry and crypt branching. Gut, 42, 799–806.[Abstract/Free Full Text]
  14. Ward,J.M. (1974) Morphogenesis of chemically induced neoplasms of the colon and small intestine in rats. Lab. Invest., 30, 505–513.[ISI][Medline]
  15. Folino,M., McIntyre,A. and Young,G.P. (1995) Dietary fibers differ in their effects on large bowel epithelial proliferation and fecal fermentation-dependent events in rats. J. Nutr., 125, 1521–1528.[ISI][Medline]
  16. Bland,J.M. and Altman,D.G. (1986) Statistical methods for assessing agreement between two methods of clinical measurement. Lancet, 1, 307–310.[ISI][Medline]
  17. Liberman,V., Nyska,A., Kashtan,H., Zajicek,G., Lubin,F. and Rozen,P. (1996) Differing proliferative responses in proximal and distal colons of growing rats fed food eaten by adenoma patients. Dig. Dis. Sci., 41, 1057–1064.[ISI][Medline]
  18. Mariadason,J.M., Catto-Smith,A. and Gibson,P.R. (1999) Modulation of distal colonic epithelial barrier function by dietary fibre in normal rats. Gut, 44, 394–399.[Abstract/Free Full Text]
  19. McIntyre,A., Young,G.P., Taranto,T., Gibson,P.R. and Ward,P.B. (1991) Different fibers have different regional effects on luminal contents of rat colon. Gastroenterology, 101, 1274–1281.[ISI][Medline]
  20. Campbell,J.M. and Fahey,G.C. Jr (1997) Psyllium and methylcellulose fermentation properties in relation to insoluble and soluble fiber standards. Nutr. Res., 17, 619–629.[CrossRef][ISI]
  21. Lupton,J.R. and Kurtz,P.P. (1993) Relationship of colonic luminal short-chain fatty acids and pH to in vivo cell proliferation in rats. J. Nutr., 123, 1522–1530.[ISI][Medline]
  22. Kissmeyer-Nielsen,P., Mortensen,F.V., Laurberg,S. and Hessov,I. (1995) Transmural trophic effect of short chain fatty acid infusions on atrophic, defunctioned rat colon. Dis. Colon Rectum, 38, 946–951.[ISI][Medline]
  23. Sakata,T. (1987) Stimulatory effect of short-chain fatty acids on epithelial cell proliferation in the rat intestine a possible explanation for trophic effects of fermentable fibre, gut microbes and luminal trophic factors. Br. J. Nutr., 58, 95–103.[ISI][Medline]
  24. Ichikawa,H. and Sakata,T. (1998) Stimulation of epithelial cell proliferation of isolated distal colon of rats by continuous colonic infusion of ammonia or short-chain fatty acids is nonadditive. J. Nutr., 128, 843–847.[Abstract/Free Full Text]
  25. Kripke,S.A., Fox,A.D., Berman,J.M., Settle,R.G. and Rombeau,J.L. (1989) Stimulation of intestinal mucosal growth with intracolonic infusion of short-chain fatty acids. J. Parenter. Enteral Nutr., 13, 109–116.[Abstract]
  26. Gibson,P.R., Kilias,D., Rosella,O., Day,J.M., Abbott,M., Finch,C.F. and Young,G.P. (1998) Effect of topical butyrate on rectal epithelial kinetics and mucosal enzyme activities. Clin. Sci., 94, 671–676.[ISI][Medline]
  27. Sengupta,S., Wong,C.S.M., Tjandra,J.J. and Gibson,P.R. (2001) Proof that butyrate delivery to the distal colon suppresses colorectal tumorigenesis in rats. J. Gastroenterol. Hepatol., 16 (suppl.), A21–22.[CrossRef]
  28. Hong,M.Y., Chang,W.C., Chapkin,R.S. and Lupton,J.R. (1997) Relationship among colonocyte proliferation, differentiation and apoptosis as a function of diet and carcinogen. Nutr. Cancer, 28, 20–29[ISI][Medline]
Received July 25, 2002; revised November 14, 2002; accepted November 15, 2002.





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Articles by Wong, C. S.M.
Articles by Gibson, P. R.