Mucin dynamics in the chick small intestine are altered by starvation

The absorptive surface of the small intestine is covered by a layer of mucus secreted by goblet cells. The secreted mucins and thickness of the adherent layer influence nutrient digestion and absorption processes as well as the functionality of the mucosa. In this study, methods for the analysis of mucin synthesis and dynamics in the chick small intestine are described. A fragment of chicken mucin cDNA was isolated and characterized; this fraction had 60% homology to human mucin MUC-5AC. The thickness of the mucus adherent layer and the relative amounts of mucin glycoprotein and mRNA were also examined in the small intestines of control and starved chicks. Relative amounts of intestinal mucin mRNA and protein increased in the duodenum and jejunum of starved chicks, and mucus adherent layer thickness decreased throughout the small intestine. In starved chicks, higher mRNA expression and protein concentrations with lower amounts of adherent mucus may be related to a higher rate of degradation of the mucus layer, a lower rate of mucus secretion, or an altered rate of mucin turnover. It thus appears that starvation alters mucus dynamics in the small intestine, and this may affect intestinal digestive function and defense.     

Early response of ornithine decarboxylase activity and energy metabolism to postsurgery refeeding in rat small intestine.

INTRODUCTION: Enterocyte proliferation and cellular energy status are important to intestinal integrity after starvation and trauma. The proliferative response to nutrients is expressed in the activity of ornithine decarboxylase (ODC), but ODC activity and ATP level in the intestinal mucosa the first hours after surgery and immediate refeeding are not known. METHODS: Male Wistar rats (240-280 g) were starved for 48 h and submitted to laparotomy with distal ileal transection, gastrostomy and jejunal instillation of either enteral formula or saline. The ODC activity and ATP content of the jejunal mucosa were analysed in samples taken at 1, 2, 4 and 6 h after surgery. RESULTS: ODC activity increased and reached the highest peak at 2 h in the refed animals. ATP concentration and energy charge of jejunal mucosa were significantly reduced 6 h after surgery compared to initial levels, but there were no differences between animals that were refed or not. Intestinal transection did not stimulate ODC activity. CONCLUSION: ATP levels in intestinal mucosa decreased after surgery, and early enteral feeding did not seem to prevent this decrease during the first 6 h. Refeeding immediately after surgery elicits an early but transient increase of ODC activity in rat jejunal mucosa.     

禁食和再喂养期间大鼠小肠胰岛素和胰岛素样生长因子-1受体的比较

胰岛素样生长因子-1（IGF-1）和胰岛素可能是肠道生长的重要调节因子。为了研究肠细胞萎缩和再生期间小肠IGF－1受体（IGF-IR）和胰岛素受体（IR），我们比较了禁食72小时和肠内再喂养24～72小时大鼠空肠IGF-IR和IR表达的指标。禁食引起肠萎缩，血浆胰岛素和IGF－1浓度降低以及空肠IGF-1信使RNA（mRNA）水平的明显降低，再喂养可逆转这些改变。禁食明显增加胰岛素与空肠特异性地结合，IR含量（达对照组的230％）和9．6kb和7．4kbIRmRNA转录本水平（分别达对照组的202％和218％）。再喂养时，这些IR指标迅速降到对照组水平。禁食时IGP-IR（用Scatchard分析）和IGF－1－RmRNA无明显的改变。再喂养后的前24小时间11-kbIGF－IRmRNA转录本明显增加（达对照组水平166％），IGF-IR数量增加3倍。我们的结论是：大鼠空肠的IR和IGF－IR受到不同营养物利用状态的调节。再喂养时空肠IGF－1和IGF－IR表达的向上调节表明，IGF作用途径在对肠内营养物产生肠道营养反应的过程中起作用。     

The effects of starvation and refeeding on muscle protein synthesis and catabolism in the young rat

We studied the effects of acute starvation and refeeding on muscle protein synthesis and degradation in young rats. As measures of synthesis, we determined muscle RNA concentration and the rate of incorporation of [14C]leucine into skeletal muscle protein (Sm). As an estimate of nitrogen retention we measured urea production (UrP). Starvation reduced these variables significantly. One refeeding period returned Sm to control values, only partially restored RNA concentration, and increased UrP. We determined the urinary excretion rate of 3-methylhistidine (3-MH) as a measure of the rate of myofibrillar protein degradation. Excretion of 3-MH was lowest in control and highest in starved rats. Refeeding decreased 3-MH excretion to a level midway between control and starved animals. Growth was attended by high rates of synthesis and low rates of degradation. Starvation depressed synthesis and increased degradation. With refeeding, synthesis increased and degradation decreased, compared with the starved state.     

Ischemia/reperfusion-induced disruption of rat small intestine transit is reversed by total enteral nutrition

OBJECTIVES: A reduced blood flow to the gut is a consistent event after traumatic shock. Enteral nutrition support has been shown to reduce the septic morbidity after major trauma. We evaluated the effects of a transient ischemia followed by reperfusion (I/R) and an enteral nutrition support regimen on the motility of the small intestine of the rat. METHODS: A catheter was placed in the upper duodenum and the small intestine was then made ischemic by clamping the superior mesenteric artery for 45 min; the arteries of sham rats were isolated but not clamped. Intestinal transit was evaluated by measuring the amount of fluorescein isothiocyanate-dextran (12 000 MW) in each of 10 intestinal sections at 30 min after injection through the duodenal catheter. The mean geometric center of marker distribution (MGC) was calculated for each group and compared. In a second study, I/R was followed by infusion of saline or a complete nutrient solution overnight, and transit was determined. RESULTS: Intestinal transit (as the MGC) of I/R rats at 24 h after the beginning of reperfusion (3.5 +/- 0.2) and 48 h after the beginning of reperfusion (4.5 +/- 1.1) was significantly lower than that in the respective sham controls (5.1 +/- 0.3 and 5.9 +/- 0.5). The MGC for rats receiving a nutrient solution overnight during the reperfusion phase (6.0 +/- 1.1) was significantly increased compared with the MGC of 4.8 +/- 0.3 for rats receiving saline during the same period. CONCLUSIONS: These results demonstrate a long-term deleterious effect of a non-lethal ischemia on intestinal transit and may be one explanation for many of the sequelae occurring after ischemia. In addition, these results demonstrate that a nutrient infusion will prevent the delayed transit. This may provide a partial explanation for the beneficial effects of total enteral nutrition in the clinical situation of posttraumatic injury.     

Retinoid metabolism in the rat small intestine

Vitamin A (retinol) is essential for epithelial cell growth, differentiation and proliferation. The absorption of retinol occurs in the small intestine, and the metabolism of this vitamin is not well studied in this organ. The intestinal epithelium has a high rate of cell proliferation and differentiation, and the present study looked at the level of retinoids and metabolizing enzymes involved in their interconversion along the villus-crypt axis under normal conditions. Intestine was removed from control rats, and enterocytes at various stages of maturation and differentiation were quantified by the metal chelation method. Using HPLC, various retinoid concentrations in the cell homogenate and the metabolizing enzymes in the cytosol were quantified. The proliferating crypt cells were found to have a higher level of retinoic acid as well as of the enzymes involved in its formation, such as retinaldehyde oxidase and retinol dehydrogenase, compared with the villus cells, suggesting a possible role for this compound in intestinal epithelial cell proliferation and differentiation. The high level of retinol and high retinaldehyde reductase activity in the villus cells suggest the important role played by this enzyme in the conversion of dietary beta-carotene to retinol via retinaldehyde. In summary, this study has given for the first time a detailed analysis of the retinoid levels and metabolizing enzymes in different cell populations in the rat small intestinal epithelium.     

Evaluation of polyphenol bioavailability in rat small intestine

Summary Background Dietary polyphenols, which are contained in several foods of plant origin, have been reported to be effective protective agents against cardiovascular diseases and cancer. However, data on their absorption from the gastrointestinal tract are still scarce and, often, contradictory. Aim of the study In this report, evaluation of polyphenol bioavailability was carried out by using segments of the small intestine from rat. The extent of absorption throughout the small intestine of rat was evaluated with two model compounds, tannic acid and catechin, as representatives of high and low molecular weight polyphenols, respectively. The consequence of the binding of tannic acid to BSA on both tannic acid absorption and in vivo protein digestibility was also examined. Methods Polyphenol solutions of different concentrations were injected into the lumen of ligated segments (6 cm) of the small intestine and the segments incubated in buffer for 5 min. The residual amount of polyphenol in the lumen of each segment was assayed by maximum absorption in the UV/VIS optical spectrum as was the amount of compound that had crossed the gut wall into the incubation buffer. Digestibility of BSA and of a BSA-tannic acid complex was assayed with rats. Results The results indicated a significant, concentration-dependent, disappearance of both polyphenols from the small intestine of the rat, with higher uptake levels being evident for tannic acid (50 %) than catechin (30 %). However, complete transfer through the gut wall was not observed with tannic acid whilst low but significant amounts (10 %) were detected in incubation buffers with catechin. Partial binding of polyphenols by endogenous proteins in the intestinal lumen was also demonstrated. Complexing tannic acid with BSA (1:10 mol/mol) was not found to affect either the extent of interaction of tannic acid with the small intestine or the in vivo digestibility of the protein. Conclusions Our experiments show that tannic acid and catechin both interact with the gut but only catechin appears able to traverse the gut. In addition, they provide evidence for binding of tannic acid and catechin by endogenous proteins in the intestinal lumen. This may limit their absorption from the small intestine. BSA complexed with tannic acid was as readily digested as BSA alone. This may suggest that tannic acid exerts anti-nutritional effects by binding to proteins of the gut wall and interfering with gut function rather than by inhibition of dietary protein digestion. Received: 1 February 2001 / Accepted: 14 May 2001     

Absorption and metabolism of anthocyanin cyanidin-3-glucoside in the isolated rat small intestine is not influenced by ethanol

Summary. Anthocyanins are receiving renewed attention for their positive health attributes. High intakes and an adequate absorption rate of anthocyanins are necessary for efficient protection, though other dietary agents might influence absorption efficacy. The aim of this study was to investigate intestinal handling of luminally administered cyanidin-3-glucoside in the absence and presence of ethanol in an isolated preparation of luminally and vascularly perfused rat small intestine. A synthetic perfusate free from blood components was used as vascular medium, with a perfluorocarbon as the oxygen carrier. Luminal media consisted of a bicarbonate buffered sodium chloride solution in water or in water/ethanol (95/5), spiked with cyanidin-3-glucoside. Absorption rate of cyanidin-3-glucoside was 4.3% ± 3.2 (n=5). Ethanol (5 %) had no significant influence on absorption rate (2.9% ± 1.8, n=5). Irrespective of the presence of ethanol, the majority of the absorbed cyanidin-3-glucoside appeared unchanged, besides some cyanidin-3-glucoside-conjugate.     

Beta-carotene 15,15'-dioxygenase activity is responsive to copper and iron concentrations in rat small intestine

OBJECTIVE: Previous in vitro studies have suggested that beta-carotene 15,15'-dioxygenase is an iron-dependent enzyme. However, in vivo, it is difficult to alter iron tissue concentration by varying dietary iron because of homeostatic control. On the other hand, an interaction between iron and copper has been shown, i.e., copper-deficiency results in an increase of iron in rat liver. Therefore, we hypothesized that intestinal iron concentration could be increased by copper-deficiency. Our objective was to examine the effects of iron as affected by dietary copper on beta-carotene 15,15'-dioxygenase activity in the small intestine. METHODS: Weanling male Sprague-Dawley rats (40 to 45g) were divided into four dietary groups: two copper-adequate groups (6.0 microg Cu/g diet) and two copper-deficient groups (0.6 microg Cu/g) combined with either normal iron (44 microg Fe/g) or high iron (87 microg Fe/g). Iron and copper concentrations were determined by atomic absorption spectrophotometry and the dioxygenase activity by reverse phase HPLC. RESULTS: Intestinal copper concentration was significantly reduced (40%) by the consumption of the copper-deficient diets, but intestinal iron was not changed by doubling dietary iron in rats fed either copper-adequate or copper-deficient diets. However, as hypothesized, the two copper-deficient groups exhibited higher intestinal iron concentration (> or =137%, p<0.001) than the copper-adequate controls. In addition, intestinal beta-carotene 15,15'-dioxygenase activity was increased by 27% and 106%, respectively, for copper-deficient rats fed either normal or high iron diets, compared to the respective copper-adequate controls (p<0.01). The dioxygenase activity was not significantly affected by dietary iron in either copper-adequate or copper-deficient groups. Finally, the enzyme activity was positively correlated (r=0.67, p<0.0001) with iron concentration and negatively correlated (r=-0.49, p<0.01) with copper concentration in small intestine. CONCLUSIONS: Intestinal beta-carotene 15,15'-dioxygenase may be an iron-dependent enzyme sensitive to copper status in vivo.     

Digestion and absorption of NAD by the small intestine of the rat

A number of preparations of varying complexity have been used in an effort to elucidate the reactions by which NAD is hydrolyzed to nicotinamide during intestinal digestion. NAD labeled with 14C in the adenine or pyridine moiety was the substrate used with perfused rat intestine, live rats, perfused live rats, with collection of portal flow, intestinal contents, mucosal tissue, or pancreatic juice. The conclusions reached are that a pyrophosphatase present in the intestinal juice and to a much lesser extent in the pancreatic juice releases 5'-AMP and nicotinamide ribonucleotide. The 5'-AMP was rapidly converted to adenosine then to inosine by bacteria-free intestinal contents. Perfused or intact intestine rapidly hydrolyzed NMN to nicotinamide riboside, which accumulated, but was not absorbed. It was slowly cleaved by an enzyme associated with the mucosal cells to nicotinamide, which was the major if not the only labeled compound absorbed.     

In vitro uptake and metabolism of pteroylpolyglutamate by rat small intestine.

The digestion and cellular uptake of 14C-pteroylheptaglutamate (14C-PG-7) was studied using an isolated cell preparation from rat small intestine and by assay of folate conjugase (EC 3.4.23.10) in whole homogenates and brush border fractions of rat small intestinal mucosa. In the cell preparation, 14C-pteroylmonoglutamate (14C-PG-1), the principal degradation product of 14C-PG-7, was concentrated earlier in the cell fraction than in the extracellular fraction, while the concentrations equalized after 15 minutes. Incubation of cells with equimolar 14C-PG-7 and 3H-PG-1 was followed by a threefold greater cellular concentration of 14C-PG-1 than 3H-PG-1. In subcellular fractionation studies, folate conjugase was minimally present in the brush border relative to the supernatant fraction. These data suggest that 14C-PG-7 is transported into the epithelial cell of the intestinal mucosa prior to its hydrolysis by intracellular folate conjugase.     

Effect of starvation and refeeding on activity of a Ca2+-dependent protease in rat skeletal muscle

The effects of starving, refeeding, and restarving rats for different periods on content of sarcoplasmic and contractile proteins and on activity of the Ca2+-dependent proteinase (CAF) in skeletal muscle was determined. Groups of five to six male rats, 8 to 11 weeks old, were starved up to 8 days, refed up to 6 days, and in two experiments, restarved up to 10 days. CAF activity was assayed in P 0-45 crude CAF fractions prepared so as to remove a protein inhibitor of CAF; the assays were demonstrated to be specific for CAF. Sarcoplasmic protein content of rat skeletal muscle changed little until after 6 days of restarvation when it decreased to 68-89% of control level (P less than 0.05). Contractile protein content decreased to 65% of control level (P less than 0.01) after 8 days of starvation, remained at this level for 4 days of refeeding, then increased to approximately 80% of control level after 6 days of refeeding, remained at this level for 2 days of restarvation, and then decreased to approximately 65% of control level (P less than 0.01) after 6 and 8 days of restarvation. Muscle CAF activity did not change during the first 8 days of starvation but increased to 113% above control level (P less than 0.01) after 6 days refeeding and then decreased to only 29% of control level (P less than 0.01) after 8 days of restarvation. These changes in muscle CAF activity are consistent with the proposed role for CAF in initiating metabolic turnover of contractile proteins, but because actual measurements of myofibrillar protein were not made and because in vivo CAF activity is difficult to assess, they do not prove this role for CAF nor do they exclude participation of other proteinases.     

Zinc uptake by basolateral membrane vesicles from rat small intestine

Zinc uptake (transport and binding) by basolateral membrane vesicles was investigated using membranes derived from small intestines of rats fed zinc-adequate and zinc-deficient diets. Uptake was separated into saturable and nonsaturable (diffusion) components. Kinetic analysis of initial rates of the saturable component of uptake (at 1 min) indicates half maximal uptake (Km) by vesicles from zinc-adequate rats was observed at a medium Zn2+ concentration of 24 microM. The maximum Zn2+ uptake rate (Jmax) of the saturable component was 17 nmol/(mg protein.min). Dietary zinc intake did not affect these parameters. Zn2+ transport by the basolateral membrane may involve an ATP-driven mechanism.     

Catechin is metabolized by both the small intestine and liver of rats

Flavan-3-ols are the most abundant flavonoids in the human diet, but little is known about their absorption and metabolism. In this study, the absorption and metabolism of the monomeric flavan-3-ol, catechin, was investigated after the in situ perfusion of the jejunum + ileum in rats. Five concentrations of catechin were studied, ranging from 1 to 100 micromol/L. The absorption of catechin was directly proportional to the concentration, and 35 +/- 2% of the perfused catechin was absorbed during the 30-min period. Effluent samples contained only native catechin, indicating that intestinal excretion of metabolites is not a mechanism of catechin elimination. Catechin was absorbed into intestinal cells and metabolized extensively because no native catechin could be detected in plasma from the mesenteric vein. Mesenteric plasma contained glucuronide conjugates of catechin and 3'-O-methyl catechin (3'OMC), indicating the intestinal origin of these conjugates. Additional methylation and sulfation occurred in the liver, and glucuronide + sulfate conjugates of 3'OMC were excreted extensively in bile. Circulating forms were mainly glucuronide conjugates of catechin and 3'OMC. The data further demonstrate the role of the rat small intestine in the glucuronidation and methylation of flavonoids as well as the role of the liver in sulfation, methylation and biliary excretion.     

Absorption and metabolism of genistein in isolated rat small intestine

Studies suggest a variety of biological effects for the isoflavonoid genistein, but there is little information regarding small intestinal absorption and metabolism. The aim of this study was to investigate the absorption and metabolism of luminally administered genistein in an isolated preparation of luminally and vascularly perfused rat small intestine. A synthetic perfusate free from blood components was used as vascular medium, with a perfluorocarbon as oxygen carrier. Luminal media consisted of a bicarbonate buffered sodium chloride solution spiked with genistein (12 micromol/L). Viability was maintained during the entire perfusion as indicated by no significant differences between genistein and control perfusions for perfusion pressure, lactate-pyruvate ratio, oxygen uptake and acid-base homeostasis. Luminal disappearance rate of genistein did not change throughout the entire perfusion time. After a significant increase until about 30 min, vascular appearance rate of total genistein reached an equilibrium. The intestinal absorption of luminally administered genistein was 40.6%, corresponding to an average uptake of 2.9 nmol. min(-1). g dry intestine(-1). The majority (31.3%) of the absorbed genistein appeared as genistein glucuronide, also recovered as the main metabolite on the luminal side (13.3%). Only small amounts of genistein (2.6%) and genistein glucuronide (2.9%) were found in the intestinal tissue. The results demonstrate a favorable uptake of genistein, a pertinent addition to the ongoing discussion about health benefits of isoflavones.     

Acute starvation and subsequent refeeding affect lymphocyte subsets and proliferation in cats

Although the early identification of patients with suboptimal nutritional status can allow the implementation of nutritional intervention to enhance the ability of the body to fight infection and disease, currently no definitive test of nutritional status exists. Therefore, this study was conducted to identify possible functional indicators of acute nutritional deprivation. The effects of total nutritional deprivation and subsequent refeeding on lymphocyte functions and subpopulations were examined in 23 healthy cats. Peripheral blood samples were analyzed at various times during food deprivation and refeeding periods. During the food deprivation period, decreases were observed in leukocyte number (P: < 0.05), lymphocyte number (P: < 0.05), percentage of CD4(+) cells [before stimulation with concanavalin-A (Con-A); P: < 0.05] and the CD4/CD8 ratio (before stimulation with Con-A; P: < 0.01) compared with d 0. Increases were observed in the percentage of CD8(+) cells [before (P: < 0.05) and after (P: < 0.01) stimulation with Con-A] and in intracellular calcium (P: < 0.01) during acute starvation. During the refeeding period, increases were observed in the percentage of CD4(+) cells (before and after stimulation with Con-A; P: < 0.01), the percentage of CD8(+) cells (before stimulation with Con-A; P: < 0.05) and lymphocyte number (P: < 0.05) compared with d 7. Lymphocyte proliferative capacity tended to decrease (P: = 0.07) during starvation and increased (P: < 0.01) during the refeeding period. These findings suggest that a 7-d starvation period had immunosuppressive effects on cats and that these effects were not completely normalized during 7 d of refeeding. CD4(+)/CD8(+) subset alterations and CD4/CD8 ratio in conjunction with lymphocyte proliferation may be useful as indices of nutritional status.     

Characterization of thiamine diphosphatase in rat small intestine.

The properties of thiamine diphosphatase (TDPase) and p-nitrophenylphosphatase (p-NPPase) in rat small intestine were investigated. TDPase activity, like p-NPPase activity, was high in the mucosa and in the proximal region. Both activities were high in the membrane-associated fractions of the duodenal mucosa. Furthermore, TDPase had the same properties as intestinal alkaline phosphatase (al-Pase). These results suggest that thiamine diphosphate (TDP) and p-nitrophenylphosphate (p-NPP) are hydrolyzed by a single enzyme, al-Pase, in the intestine.     

Kinetic study of acamprosate absorption in rat small intestine

Acamprosate (calcium bis acetyl-homotaurine), a homotaurine derivative, a structural analogue of gamma-aminobutyric acid (GABA) and an upper homologue of taurine, is a relatively new drug used to prevent relapse in weaned alcoholics. When administered orally as enteric-coated tablets at relatively high doses, this drug has a bioavailability of about 11%; however, the intestinal absorption mechanism has not been studied in depth. The present study was therefore planned to characterize the intestinal transport of acamprosate in the rat and the effect of chronic alcohol treatment on this process, quantifying its kinetic parameters and investigating possible inhibitors. Using an in vitro technique, acamprosate absorption was measured in the rat intestine from three different groups: alcohol group [fed a liquid diet containing 5% (w/v) ethanol for 4 weeks], isocaloric pair-fed control, and a solid diet group. Intestinal acamprosate absorption was found to occur mainly by passive diffusion with a diffusive permeability of 0.213+/-0.004 cm/h in control pair-fed animals, 0.206+/-0.001 cm/h in animals receiving chronic alcohol treatment, and 0.193+/-0.001 cm/h in the solid diet group. Inhibition studies showed that at a 10(-3) M acamprosate concentration, some compounds such as GABA, taurine, proline, and glycine at 40 mM each did not affect acamprosate transport. Nevertheless, when a lower concentration of the drug (10(-4) M) was assayed, a significant reduction of acamprosate transport in the presence of taurine or GABA 40 mM was found. These results suggest that acamprosate in the rat jejunum, could be transported, in part, by a carrier system. Further experiments using different concentrations of taurine (10, 20, and 80 mM) showed that the maximum inhibition (32%) is achieved at 20 mM of taurine. These latter results suggest that acamprosate and taurine share, at least, an intestinal carrier system in rat jejunum. From the above results, it can be concluded that there are probably two pathways involved in the intestinal absorption of acamprosate: passive diffusion and mediated transport, with the former being predominant. Moreover, neither chronic ethanol intake nor the type of diet seems to alter the intestinal absorption of the drug.