Promotion of intestinal carcinogenesis by Streptococcus bovis.

The involvement of Streptococcus bovis, an member of the human gut flora, in colorectal neoplastic diseases is an object of controversy. The aim of this study was to determine the effects of S.bovis and of antigens extracted from the bacterial cell wall on early preneoplastic changes in the intestinal tract. Adult rats received i. p. injections of azoxymethane (15 mg/kg body weight) once per week for 2 weeks. Fifteen days (week 4) after the last injection of the carcinogen, the rats received, by gavage twice per week during 5 weeks, either S.bovis (10(10) bacteria) or wall-extracted antigens (100 microg). One week after the last gavage (week 10), we found that administration of either S.bovis or of antigens from this bacterium promoted the progression of preneoplastic lesions through the increased formation of hyperproliferative aberrant colonic crypts, enhanced the expression of proliferation markers and increased the production of IL-8 in the colonic mucosa. Our study suggests that S.bovis acts as a promoter of early preneoplastic lesions in the colon of rats. The fact that bacterial wall proteins are more potent inducers of neoplastic transformation than the intact bacteria may have important implications in colon cancer prevention.

Promotion of intestinal carcinogenesis by dietary methionine.

The metabolism of the polyamines spermidine and spermine is known to be enhanced in rapidly proliferating cells. Methionine is a precursor of the aminopropyl moieties of these amines. Therefore, it was of interest to study the effects of a methionine supplemented diet on polyamine metabolism and preneoplastic changes occurring in the intestinal tract of rats treated with the chemical carcinogen azoxymethane (AOM). Adult Wistar rats received 15 mg AOM/kg body wt (i.p.) once each week for 2 weeks. Thereafter, the rats were randomly divided into two groups and received controlled isoenergetic diets containing the same amount of folate, choline and vitamin B12 during 12 weeks: one group was kept on a standard diet; the other was fed the same diet, except that 1% L-methionine was added at the expense of carbohydrates. After 12 weeks, the administration of the methionine-supplemented diet stimulated the turnover rate of ileal epithelial cells, indicating enhanced crypt cell proliferation. Furthermore, in this group, a 2-fold increase in the number of aberrant hyperproliferative crypts and the appearance of tumors was observed in the colon. These effects were accompanied by the increased formation of spermidine and spermine due to the enhancement of S-adenosylmethionine decarboxylase activity and by the upregulation of Cdx-1, a homeobox gene with oncogenic potentials. The experimental data do not support the view of a chemopreventive effect of dietary methionine supplementation on intestinal carcinogenesis in rats, even at an early phase of preneoplastic development, but rather suggest that methionine promotes intestinal carcinogenesis.

Suppression of preneoplastic changes in the intestine of rats fed low levels of polyamines.

Administration for 7 days of an enteral diet that is naturally deficient in polyamines strikingly reduces the preneoplastic changes observed in the intestines of adult Wistar rats previously treated with the carcinogen 1,2-dimethylhydrazine. On the contrary, supplementing the enteral diet with spermidine favors preneoplastic development. The effects of the low-polyamine diet included a 40% decline in the putrescine content of the intestinal mucosa, a significant decrease in the turnover rate of the epithelial cells from the crypts to villus tip in the ileum, and a 2-fold reduction in the number of abnormal colonic crypts. The experimental data support the view that it might be of interest to control the dietary intake of polyamines in the clinical management of cancer patients.

Enteral supplementation with ornithine alpha ketoglutarate improves the early adaptive response to resection.

BACKGROUND: Polyamine synthesis or uptake, or both, might be an important event that initiates the adaptive hyperplasia seen in the intestinal remnant after partial small bowel resection. AIM: The ability of an enteral diet supplemented with the ornithine salt: ornithine alpha ketoglutarate (OKG), a precursor for polyamine synthesis, to modulate the adaptive response of the remnant ileum after jejunectomy was evaluated. METHODS: Adult Wistar rats underwent a resection of the proximal 50% of the small intestine. Controls underwent a single transection. The rats were fed intragastrically with a nutritive mixture supplemented either with casein hydrolysate or with OKG (1 g/kg). The isoenergetic and isonitrogeneous diets was given continuously for seven days. RESULTS: Villus and crypt hyperplasia was observed in the remnant ileum compared with transfected controls. OKG supplementation started after resection a further increase in villus height. After resection, OKG supplementation increased significantly the putrescine content and the amount of ornithine decarboxylase mRNA. A twofold to threefold increase of sucrase activity was measured in the resected animals compared with the transected rats. In contrast, the amount of sucrase mRNA was significantly lower in the ileum of the resected rats and OKG supplementation initiated a further drop in the amount of sucrase mRNA without pronounced changes in enzyme activity. CONCLUSIONS: The adaptive hypertrophy seen after resection can be accelerated by supplementing the diet with ornithine (OKG) a precursor of polyamine synthesis. In the remnant ileum, the reduced amount of sucrase mRNA, despite the increased level of sucrase activity, suggests a post-translational control of sucrase expression.

Intestinal absorption of calcium in vivo is dependent on endogenous nitric oxide.

This study examines the role of nitric oxide (NO) in the regulation of calcium absorption in the small intestine. Calcium absorption was quantified by measuring 45Ca++ transport from lumen to blood in an intestinal segment (duodenum and 20 cm of the proximal jejunum) perfused by both intraluminal and vascular routes in anesthetized rats. When administered i.v. as bolus injections, NG-nitro-L-arginine methyl ester (L-NAME, 10 mg.kg-1), an inhibitor of NO biosynthesis, decreased calcium absorption with a concomitant increase in blood pressure and a decrease in mesenteric blood flow. Conversely, the nitrovasodilators 3-morpholinosydnonimine (2 mg.kg-1) and S-nitroso-N-acetylpenicillamine (10 micrograms.kg-1), which generate NO spontaneously, both increased calcium absorption with no change in mesenteric blood flow. When infused i.v., L-NAME (3 mg.hr-1.kg-1 for 40 min) induced a decrease in calcium absorption that was reversed by the NO donor sodium nitroprusside (1.5 mg.hr-1.kg-1 when infused for the last 20 min of the 40-min L-NAME infusion). Sodium nitroprusside infusion (1.5 mg.hr-1.kg-1) caused an increase in calcium absorption that was not reversed by L-NAME (3 and 30 mg.hr-1.kg-1). The present findings suggest that NO is involved in basal calcium absorption in rat small intestine in vivo.

Functional and metabolic changes in intestinal mucosa of rats after enteral administration of ornithine alpha-ketoglutarate salt.

BACKGROUND: Ornithine alpha-ketoglutarate salt efficiently improves the nutritional status of protein-depleted patients. Our aim was to explore the effects of ornithine alpha-ketoglutarate supplementation on intestinal physiology in healthy animals. METHODS: Rats were given a nutritive mixture supplemented with ornithine alpha-ketoglutarate (1 g.kg-1 per day) by enteral route for 7 days. Controls received the diet supplemented with casein acid hydrolysate under isoenergetic and isonitrogenous conditions. RESULTS: An adaptive hyperplasia of the villi and an increase in the brush-border hydrolase activities were observed in rats receiving ornithine alpha-ketoglutarate. Because of the high ornithine aminotransferase activity, ornithine alpha-ketoglutarate-derived ornithine was extensively transaminated with a concomitant enhancement of ornithine decarboxylation. Surprisingly, with glutamate and putrescine, the products of ornithine transamination and decarboxylation, gamma-aminobutyric acid accumulated (10-fold to 16-fold) dramatically in the intestinal mucosa of rats treated with ornithine alpha-ketoglutarate. Because gamma-aminobutyric acid formation was completely prevented by the diamine oxidase inhibitor aminoguanidine but was not modified after inactivation of ornithine aminotransferase by 5-fluoromethylornithine, it is evident that gamma-aminobutyric acid is formed in the mucosa from ornithine via putrescine as an intermediate. CONCLUSIONS: It is assumed that enhanced gamma-aminobutyric acid formation in the intestinal mucosa by ornithine alpha-ketoglutarate treatment might be of physiologic importance in the regulatory processes of cell growth and differentiation.

Beneficial effects of L-arginine on intestinal epithelial restitution after ischemic damage in rats.

The polyamines are involved in repair processes after intestinal ischemia. Arginine and ornithine, both precursors of polyamines were therefore expected to exert beneficial effects on mucosal barrier dysfunction. Arginine may also generate NO and there is support for the view that NO may be beneficial after an ischemic insult. Male Wistar rats were given, by gavage, isonitrogenous solutions of L-arginine (0.5 g/kg) or L-ornithine (0.7 g/kg) 17 and 2 h before ischemia. Controls received an isonitrogenous solution of casein hydrolysate (1 g/kg). Transient intestinal ischemia was produced in anesthetized rats by occluding the superior mesenteric artery for 90 min. Intestinal morphology, hydrolase activities, polyamine and cGMP contents, and cell proliferation rates were determined 4 h after reperfusion. Administration of arginine or ornithine did not prevent ischemic damage but accelerated morphological repair, enhanced cell proliferation, and polyamine content was observed. Arginine was significantly more effective than ornithine. Formation of cGMP was enhanced after arginine administration. NG-nitroarginine methylester, an inhibitor of NO synthase, prevented the arginine effects on mucosal repair. We conclude that arginine-derived NO is an important mediator in the restitution of intestinal mucosa by minimizing cell injury during reperfusion.

Effect of fasting and thyroidectomy on cysteine proteinase activities in liver and muscle.

Prolonged starvation mimics chronic negative nitrogen balance observed in many physiopathological situations. During starvation, an initial decrease in protein utilization (phase I) is followed by a long period of protein sparing (phase II) that ends with a marked rise in nitrogen excretion (phase III). Variations in protein metabolism during starvation are determined by changes in protein synthesis and degradation rates (Cherel, Y., Attaix, D. Rosolowska-Huszcz, D., Belkhou, R., Robin, J.P., Arnal, M. and Le Maho, Y. (1991) Clin. Sci. 81, 611-619), but little information is available on expression of proteolytic systems. In this study, cathepsin B, H and L activities were compared in hindlimb muscles and liver at various phases of starvation in thyroidectomized and sham-operated rats. In muscle, cathepsin activities fell from the fed state to phase II, which suggests that cathepsins may play a role in the curtailment of muscle proteolysis during protein sparing phase. This decrease of muscle cathepsin activities was reproduced by thyroidectomy alone. In contrast, liver cathepsin B and H activities fell during starvation, but were not affected by thyroidectomy alone. Liver cathepsin L decreased only during starvation in thyroidectomized animals. These observations emphasize that different mechanisms modulate cathepsin expression in skeletal muscle and liver.

Dietary calcium supplementation, blood pressure, and intestinal calcium absorption.

This study investigated the potential intestinal mechanism in the blood pressure-lowering effect of dietary calcium supplement. Adult Wistar rats received, through a gastrotomy cannula, either 5 or 30 mg elemental calcium.d-1 x 100 g BW-1 for 10 days. At day 11, mean arterial pressure was measured directly in anesthetized animals and calcium absorption determined using an in situ measurement technique of calcium absorption. An intestinal loop (duodenum and proximal jejunum) was perfused both by the intraluminal and the vascular routes. A solution containing 45Ca was perfused intraluminally and the 45Ca appearing in the venous effluent was determined to estimate calcium absorption. Oral calcium supplementation caused intestinal calcium uptake by the isolated loop to increase almost twofold. It also resulted in an 18% increase in mesenteric blood flow. The mean arterial pressure was decreased in calcium supplemented rats compared with control rats (87 +/- 5 vs. 78 +/- 5 mmHg; p < 0.05). These findings could indicate that the passive part of intestinal calcium transport increases in response to dietary calcium supplement. This improvement may participate in the blood pressure-lowering effect of a high calcium diet.

Polyamines and the recovery of intestinal morphology and function after ischemic damage in rats.

We have followed the time-course of the morphological and functional recovery of intestinal mucosa after 90 min of mesenteric vascular occlusion. At the end of the ischemic period the villi were smashed, but crypts were preserved. Microvillous hydrolase activities showed a dramatic drop when compared with sham-operated controls. Reperfusion was followed by an immediate upsurge of ornithine decarboxylase activity and a significant (p < 0.01) enhancement of putrescine and N1-acetyl-spermidine concentrations, while spermidine and spermine concentrations in mucosal cells decreased. This indicated that, both, de novo synthesis and degradation rates of the polyamines were increased. Treatment with alpha-difluoromethyl-ornithine, a selective inactivator of ornithine decarboxylase prevented the accumulation of active enzyme, but did not prevent morphological healing. It delayed however the recovery of sucrase and aminopeptidase-specific activities. Our results suggest that in addition to de novo synthesis, other sources of polyamines are mobilized to an extent that growth at a normal rate is supported. This indicates that the presence of active ornithine decarboxylase enzyme is not a prerequisite for the restitution of intestinal integrity after ischemia. We suggest that in a situation of inadequate polyamine supply the restoration of vital processes (mucosal regeneration) has priority over the restoration of specific functions.

Comparison of different lipid substrates on intestinal adaptation in the rat.

The relative effects of medium chain (MCT) and long chain triglycerides (LCT) on intestinal morphology and functions were compared. Adult rats received intragastrically for 10 days an isoenergetic mixture containing either 50% MCT/50% LCT or 100% LCT. The other constituents of the diets were identical, and animals fed a standard diet orally were used as a reference group. Animals who were given the MCT/LCT diet showed a higher mucosal mass and protein content and increased villus length and crypt depth in the proximal part of the small intestine compared with the LCT and control diet groups. Administration of [3H] thymidine 12 hours before death resulted in a significant increase in the incorporation of the precursor into cellular DNA in the jejunum of rats given MCT. In rats given LCT as the only fat, the free fatty acid content of the microvillus membrane showed a 20 fold increase and at the same time there was a significant drop in the cholesterol content and in the cholesterol/protein ratio. Differences in the lipid composition of enterol diet or in the microvillus membrane did not effect adversely membrane bound hydrolase activities. These findings suggest that MCT in the diet confers advantages in addition to the provision of rapidly available energy.

Development of a technique for in situ studies of calcium absorption in the intestine of rats.

The aim of the present study was to develop (feasibility, reliability, reproducibility) a technique for the in situ measurement of intestinal calcium absorption in the rat. An intestinal loop (duodenum+proximal jejunum) was perfused both by the intraluminal and vascular routes. A solution of NaCl 155 mM and CaCl2 1.25 mM containing 45Ca was perfused intraluminally at a flow rate of 0.2 ml/min and the 45Ca appearing in the venous effluent was determined to estimate calcium absorption. This technique was used to study the effect of a 10-day period of calcium supplementation on calcium absorption. The animals received enterally either 5 or 30 mg/day per kg BW. The results showed that intestinal calcium transport was enhanced when rats were given the calcium supplement for 10 days prior to the experiment (99.3 +/- 2.5 (n = 5) versus 36.6 +/- 3.6 (n = 5) nmol/min per kg BW, P < 0.001). This study indicates that dietary calcium supplement enhances calcium absorption probably by increasing the passive transport of calcium in the small intestine.

Dietary control of the lactase mRNA distribution along the rat small intestine.

At weaning, the lactase-phlorizin hydrolase (LPH) mRNA was shown to disappear specifically from the distal part of ileum while remaining abundant in the more proximal segments of the small intestine. The purpose of this study was to analyze the longitudinal distribution of this transcript in rats whose intestinal lumen content was modified before and after weaning. Preweaned animals force-fed with an artificial diet retained a high amount of LPH mRNA in the jejunum, whereas this transcript precociously decreased in the distal ileum. Conversely, prolonged nursing delayed the specific decay of the LPH mRNA in the latter segment. Food deprivation in preweaned animals did not alter the longitudinal distribution of this transcript in that it remained abundant in the distal ileum. In adult rats, rearranging the order of the small intestinal segments with regard to the intraluminal flow of nutrients did not modify the typical distribution of the LPH mRNA. These results suggest that switching over from milk to the adult-type diet at weaning contributes to the modification of the longitudinal distribution of the LPH mRNA that normally occurs at this stage. However, once the adult pattern of expression of this transcript is established, it cannot be significantly altered by changing the position of each intestinal segment as well as its luminal content.

Effects of amino acids in mixtures given by enteral or parenteral route on intestinal morphology and hydrolases in rats.

This study compares the effects of amino acid addition to an elemental liquid diet containing carbohydrates and triglycerides given either intragastrically or intravenously on the morphology and on hydrolase activities in the jejunum and ileum of adult rats. The isocaloric mixtures were administered for 4 days and control rats received an isocaloric laboratory diet orally. Independent of their content in amino acid, all mixtures given intravenously caused a drop in mucosal weight and a shortening of the height of the villi in both the jejunum and ileum. By enteral route, the addition of amino acids to a carbohydrate-triglyceride liquid diet led to the maintenance of normal villus height (this effect being prominent in the ileum) and to a significant increase of jejunal sucrase and aminopeptidase activities when compared with the carbohydrate-triglyceride mixture. Feeding the mixtures by parenteral route caused a significant drop of both enzyme activities. In contrast, lactase activity was generally not modified by the route of nutrient administration or by the composition of the diets. However, the absence of amino acid in the mixture given intravenously caused a specific drop of lactase activity in the ileum. Ileal sucrase activity was lowered dramatically by intragastric or intravenous feeding of the elemental diets. This effect was not modulated by the presence of amino acids. The presence of amino acids caused a significant drop of aminopeptidase activity in the ileum independently of the route of administration when compared with animals receiving the carbohydrate-triglyceride liquid diet.(ABSTRACT TRUNCATED AT 250 WORDS)

Rat lactase activity and mRNA expression in relation to the thyroid and corticoid status.

The effect of glucocorticoids and thyroid hormones on lactase expression was investigated along the small intestine of rats. In sucklings thyroxine injections promoted a precocious drop of enzyme activity but not of mRNA. Hydrocortisone did neither modify lactase activity nor its mRNA expression. In adults decreasing the amount of thyroid hormones led to a slight and reversible increase of the lactase mRNA content whereas lactase activity rised more dramatically. Thus, thyroid hormones in contrast to corticoids, are involved in the posttranscriptional control of lactase during the suckling period. Yet, none of these hormones might induced the modification of the longitudinal distribution of the lactase mRNA that occurred at weaning.

Adaptation of intestinal hydrolases to starvation in rats: effect of thyroid function.

The effects of long-term starvation on the activities of sucrase, lactase, and aminopeptidase, and on their respective mRNA were determined in the small intestine of thyroidectomized and sham-operated adult rats. Thyroidectomy reduced the protein loss at the level of the intestinal brush border membranes during starvation. Prolonged fasting caused a significant decrease in sucrase activity, but thyroidectomy partly prevented this effect. However, the amount of the corresponding mRNA dropped during long term starvation without incidence of thyroidectomy. Lactase activity in the brush border membranes was increased by starvation, and thyroidectomy caused a further elevation of the enzyme activity. Simultaneously, lactase mRNA content rose only slightly compared to the enzyme activity. Aminopeptidase activity and mRNA content decreased during starvation and thyroidectomy did not prevent this process. These results indicate that intestinal hydrolases respond non-coordinately to long-term food deprivation. In addition, the thyroid status of the animals has a direct influence on the adaptation of several brush border hydrolases to starvation. This suggests that the drop in plasma thyroid hormones during fasting allows a better maintenance of protein content and of hydrolase activities in the brush border membranes of the small intestine. These adaptive processes seemed to be partly controlled at a post-transcriptional level.

Imbalance between jejunum and ileum in the response of brush border hydrolases to oral feeding after intravenous alimentation in rats.

The effect of oral refeeding after total parenteral nutrition (TPN) on brush border hydrolases was measured in the proximal jejunum and ileum of adult rats. The animals received intravenously for 4 days a mixture of Intralipid 10% and Vamine-Glucose. At the end of TPN, oral feeding was reinstituted and the rats were fed with an isocaloric standard diet (60% carbohydrate, 17% protein, 3% lipid). Sucrase, isomaltase, lactase, and aminopeptidase N activities were measured at the end of TPN and at 1, 3, and 5 days after TPN. Sham-operated rats nourished orally with the standard diet were used as controls. In both intestinal segments, lactase activity showed no significant changes at the end of TPN or during oral realimentation. Isomaltase, and especially sucrase activities, exhibited an important drop at the end of TPN. After TPN, a complete restoration of isomaltase and sucrase activities was obtained in the jejunum only. During oral refeeding a 40% deficit in sucrase activity persisted in the ileum throughout the experimental period, whereas normal isomaltase activity was restored in this segment. Aminopeptidase N activity was lowered by TPN and recovered normal values within a few hours after oral realimentation. Thus, reinstitution of oral feeding after TPN should take into account that the intestine is capable of digesting normal amounts of dietary protein but has a reduced tolerance for carbohydrates.

Adaptation of intestinal enzymes to seasonal and dietary changes in a hibernator: the European hamster (Cricetus cricetus).

Effects of diet, hibernation and seasonal variations on hydrolase activities were determined in mucosa and purified brush border membranes of the small intestine of European hamsters. Wild hamsters captured in April and fed for several weeks with an equilibrated laboratory chow (20% protein, 50% carbohydrates) exhibited a rise in disaccharidase activities (sucrase, isomaltase, lactase) but no changes in aminopeptidase N activity. During deep hibernation, in contrast to sucrase and isomaltase activities which showed only minor changes, lactase activity was significantly enhanced along the jejunoileum, and aminopeptidase N activity was maximum in the ileum. After a short period (48 h) of wakefulness and feeding following 10 days of starvation during the hibernation period, the activities of the disaccharidases and of aminopeptidase N returned to values measured in active animals. In contrast to the nutritional state, which has an important impact on the activities of intestinal enzymes, season has little effect on the intestine of the active animal under a controlled environment. The pattern of enzyme activities which occurs along the small intestine in the hibernating animal may be a prerequisite for optimum digestion during the short phases of waking during the hibernation period of the European hamster.

Prenatal exposure to alcohol in rat: effect on intestinal enzymes in offspring.

Intestinal hydrolase activities were studied during postnatal development in the offspring of rats exposed to 20% ethanol during gestation; alcohol was withdrawn at birth. Controls received water during gestation. Sucrase, lactase, glucoamylase and aminopeptidase activities were determined 2 and 4 weeks after birth in the proximal jejunum. Offspring prenatally exposed to ethanol showed a deficit in body weight and lower aminopeptidase activity during the suckling period (2 weeks). These effects were reversible by 4 weeks when alcohol was withdrawn at birth. The prenatal exposure to ethanol did not change the pattern of sucrase maturation in the intestine of offsprings. The activities of lactase and glucoamylase were not modified following prenatal exposure to ethanol. In conclusion, exposure to ethanol during gestation caused decreased abilities for the intestine of the offspring to digest protein.

A comparison of intestinal adaptation to short-term intravenous versus intragastric diet in adult rats.

This study was designed to determine, on intestinal function, the comparative effects of a fat emulsion, a carbohydrate solution, and a mixture of lipids and carbohydrates given for 4 days to adult rats either intragastrically or intravenously. The rats were separated into three groups (n = 24 in each group). Each group was divided into two populations fed either intragastrically or intravenously. Each group received one of the following nutrients: a 20% Intralipid emulsion, a mixture (1:1, V/V) of Intralipid 20% and Vamine N containing 25% glucose (W/V), a solution of Vamine-glucose supplemented with fructose to reach a final concentration of 20% (W/V). Sham-operated rats that received laboratory chow orally were used as controls. The daily caloric intake was 0.21 to 0.22 kcal/g body weight. The studies on villus morphology and on brush border enzyme activities were performed on the proximal part of the jejunum. For all nutrients, intragastric infusion provoked an increase in the villus height. The lipids were the only nutrients to cause villus lengthening by the intravenous route. Intragastric or intravenous infusion of fat provoked a deficiency in intestinal disaccharidases; the presence of carbohydrates in the diet inhibited this effect slightly. Carbohydrates given alone, either intragastrically or intravenously, caused an elevation of lactase activity. Independent of diet composition, aminopeptidase activity was reduced after intravenous feeding. In conclusion, the disaccharidase activities are largely dependent on changes occurring in the nutrient composition given either intragastrically or intravenously, whereas amino-peptidase activity was related to the route of diet administration.