GATA4 represses an ileal program of gene expression in the proximal small intestine by inhibiting the acetylation of histone H3, lysine 27.

GATA4 is expressed in the proximal 85% of small intestine where it promotes a proximal intestinal ('jejunal') identity while repressing a distal intestinal ('ileal') identity, but its molecular mechanisms are unclear. Here, we tested the hypothesis that GATA4 promotes a jejunal versus ileal identity in mouse intestine by directly activating and repressing specific subsets of absorptive enterocyte genes by modulating the acetylation of histone H3, lysine 27 (H3K27), a mark of active chromatin, at sites of GATA4 occupancy. Global analysis of mouse jejunal epithelium showed a statistically significant association of GATA4 occupancy with GATA4-regulated genes. Occupancy was equally distributed between down- and up-regulated targets, and occupancy sites showed a dichotomy of unique motif over-representation at down- versus up-regulated genes. H3K27ac enrichment at GATA4-binding loci that mapped to down-regulated genes (activation targets) was elevated, changed little upon conditional Gata4 deletion, and was similar to control ileum, whereas H3K27ac enrichment at GATA4-binding loci that mapped to up-regulated genes (repression targets) was depleted, increased upon conditional Gata4 deletion, and approached H3K27ac enrichment in wild-type control ileum. These data support the hypothesis that GATA4 both activates and represses intestinal genes, and show that GATA4 represses an ileal program of gene expression in the proximal small intestine by inhibiting the acetylation of H3K27.

Postprandial plasma retinyl ester response is greater in older subjects compared with younger subjects. Evidence for delayed plasma clearance of intestinal lipoproteins.

Postprandial vitamin A and intestinal lipoprotein metabolism was studied in 86 healthy men and women, aged 19-76 yr. Three independent experiments were carried out. In the first experiment, a supplement dose of vitamin A (3,000 retinol equivalents [RE]) was given without a meal to 59 subjects, aged 22-76 yr. In the second experiment, 20 RE/kg body wt was given with a fat-rich meal (1 g fat/kg body wt) to seven younger subjects (aged less than 50 yr) and seven older subjects (aged greater than or equal to 50 yr). In both experiments, postprandial plasma retinyl ester response increased significantly with advancing age (P less than 0.05). In the third experiment, retinyl ester-rich plasma was infused intravenously into nine young adult subjects (aged 18-30 yr) and nine elderly subjects (aged greater than or equal to 60 yr), and the rate of retinyl ester disappearance from plasma during the subsequent 3 h was determined. Mean (+/- SE) plasma retinyl ester residence time was 31 +/- 4 min in the young adult subjects vs. 57 +/- 8 min in the elderly subjects (P less than 0.05). These data are consistent with the concept that increased postprandial plasma retinyl ester concentrations in older subjects are due to delayed plasma clearance of retinyl esters in triglyceride-rich lipoproteins of intestinal origin.

Isolation, characterization, and developmental expression of the rat peptide-YY gene.

In the present study we describe the isolation, structural characterization, and developmental expression of the gene encoding the intestinal hormone peptide-YY. Examination of the nucleotide sequence of the peptide-YY gene reveals that each of the four exons encodes a functional domain of its mRNA that is analogous to the corresponding exons of the genes encoding two closely related peptides neuropeptide-Y and pancreatic polypeptide. The highly conserved structural organization of the genes encoding this family of three peptides suggests that each gene arose from the duplication of a common ancestral gene. Developmental studies reveal that the peptide-YY gene exhibits a complex pattern of tissue-specific expression in the gastrointestinal tract. Unlike many gastrointestinal hormones, peptide-YY mRNA levels are highest before birth. The pancreas appears to be the major site of peptide-YY gene expression in the fetus, exceeding colonic expression by 7-fold. The abundance of peptide-YY mRNA in the pancreas declines rapidly after birth, in contrast to the colon, where mRNA levels are maintained throughout development into adulthood. Expression of the peptide-YY gene before birth antedates the presence of known enteral secretagogues for this hormone, suggesting alternate mechanisms that control its biosynthesis during development.

Sex differences in postabsorptive plasma vitamin A transport.

This study examined postabsorptive plasma vitamin A after doses of retinyl palmitate in healthy men (n = 28) and women (n = 31). On consecutive days one physiologic [3000 retinol equivalents (RE)] and one pharmacologic dose (105,000 RE) were administered and blood samples collected. Plasma retinol and retinyl esters were measured by HPLC. Tolerance curves were constructed by plotting plasma retinyl ester concentration vs time. Postprandial retinyl ester response was measured as peak rise in retinyl ester concentration and area under the curve (AUC). Peak plasma retinyl ester concentration occurred earlier for females but the earlier peak was significant only for younger subjects (< or = 50 y, P < 0.02) given the low dose and older subjects (> 50 y, P < 0.02) given the high dose. Peak rise and AUC were lower in females than in males, but this difference was significant for the high dose only (P < 0.05). In the high-dose experiment, when each age group was evaluated for sex differences the peak rise was significantly greater in males than in females in the older subjects (P < 0.05). Postabsorptive plasma retinol did not change from fasting concentrations. A lower plasma response in retinyl esters in women could be due to a more efficient chylomicron-remnant clearance.

Relationship of vitamin A and vitamin E intake to fasting plasma retinol, retinol-binding protein, retinyl esters, carotene, alpha-tocopherol, and cholesterol among elderly people and young adults: increased plasma retinyl esters among vitamin A-supplement users.

We studied the relationships of supplemental and total vitamin A and supplemental vitamin E intake with fasting plasma biochemical indicators of vitamin A and vitamin E nutritional status among 562 healthy elderly people (aged 60-98 y) and 194 healthy young adult (aged 19-59 y) volunteers. All subjects were nonsmokers. For the young adults, plasma retinol was significantly greater in males than in females (p less than 0.01); retinol was not related to supplemental vitamin A intake for either group. Fasting plasma retinyl esters demonstrated a significant increase with vitamin A supplement use. For supplemental vitamin A intakes of 5001-10,000 IU/d, a 2.5-fold increase over nonusers in fasting plasma retinyl esters was observed for elderly people (p less than 0.05) and a 1.5-fold increase for young adults (p greater than 0.20). For elderly people, greater fasting plasma retinyl esters were associated with long-term vitamin A supplement use (greater than 5 y) and biochemical evidence of liver damage. Elderly people who take vitamin A supplements may be at increased risk for vitamin A overload.

The prevalence of vitamin K deficiency in chronic gastrointestinal disorders.

Vitamin K deficiency results in the appearance of abnormal prothrombin, deficient in gamma-carboxyglutamic acid, in the blood. The presence of abnormal prothrombin can be eliminated or lowered by the administration of vitamin K. Since the abnormal prothrombin antigen assay is approximately 1000-fold more sensitive than the prothrombin time for the diagnosis of vitamin K deficiency, this assay was used to evaluate patients with intestinal abnormalities. Vitamin K deficiency was found in 18 of 58 patients (31%) with chronic gastrointestinal disease and/or resection. All patients with vitamin K deficiency had either Crohn's disease involving the ileum or ulcerative colitis treated with sulfasalazine or antibiotics. Abnormal prothrombin levels returned toward normal in patients treated with vitamin K but not in patients who were not treated with vitamin K. The mean plasma vitamin E level in patients with vitamin K deficiency was significantly lower than in vitamin-K sufficient patients (p less than 0.01). We conclude that certain chronic forms of gastrointestinal disorders are associated with vitamin K deficiency.

Evaluation of vitamin A absorption by using oil-soluble and water-miscible vitamin A preparations in normal adults and in patients with gastrointestinal disease.

We evaluated vitamin A absorption in 50 healthy adults and 26 gastrointestinal-disease patients by measuring the postabsorptive response in plasma retinyl esters after oral doses of the vitamin. On 3 consecutive days, two physiologic-dose tests of 2000-2400 retinol equivalents (RE) and one pharmacologic-dose test (84,000 RE) were administered. The physiologic doses were given as an oil-soluble or a water-miscible preparation. In gastrointestinal-disease patients the physiologic-dose test was highly correlated with the pharmacologic-dose test for the oil-soluble preparation as determined by peak rise (r = 0.50, P less than 0.05) and area under the curve (r = 0.56, P less than 0.01), suggesting that the physiologic dose is valid for investigating vitamin A absorption. Intestinal-disease or resection patients absorbed preparations poorly. Pancreatic-disease patients absorbed the oil-soluble preparation poorly. Physiologic rather than pharmacologic doses of vitamin A can be used to study vitamin A absorption.

Asymmetrical localization of mRNAs in enterocytes of human jejunum.

Intracellular localization of specific mRNAs is known to be a mechanism for targeting proteins to specific sites within the cell. Previous studies from this laboratory have demonstrated co-localization of mRNAs and proteins for a number of genes in absorptive enterocytes of fetal rat intestine. The present study was undertaken to examine in human enterocytes the intracellular localization patterns of mRNAs for the microvillous membrane proteins lactase-phlorizin hydrolase (LPH), sucrase-isomaltase (SI), and intestinal alkaline phosphatase (IAP), and the cytoskeletal protein beta-actin. In sections of human jejunum, mRNAs were localized by in situ hybridization using digoxigenin-labeled anti-sense RNA probes. Both LPH and SI mRNAs were localized to the apical region of villous enterocytes, whereas IAP and beta-actin mRNAs were detected both apically and basally relative to the nucleus. Therefore, in contrast to LPH, SI, and beta-actin mRNAs, which co-localize with their encoded proteins, that of IAP is present in the basal region of the cell where IAP protein has not directly been demonstrated to be present. Absorptive enterocytes from humans possess the mechanisms for intracellular mRNA localization, but not all mRNAs co-localize with their encoded proteins.

Fundic atrophic gastritis in an elderly population. Effect on hemoglobin and several serum nutritional indicators.

The ratio of pepsinogen I to pepsinogen II in the circulation decreases progressively with increasing severity of atrophic gastritis of the fundic gland mucosa. Fasting blood was obtained from 359 free-living and institutionalized elderly people (age range, 60 to 99 years). A pepsinogen I/pepsinogen II ratio less than 2.9, indicating atrophic gastritis, was found in 113 (31.5%) subjects. The prevalence of atrophic gastritis increased significantly with advancing age (P less than .05). Within the atrophic gastritis group, 84 had a pepsinogen I level greater than or equal to 20 micrograms/L, indicating mild to moderate atrophic gastritis, and 29 had a pepsinogen I level less than 20 micrograms/L, indicating severe atrophic gastritis or gastric atrophy. A significant increase in the prevalences of elevated serum gastrin levels (P less than .005), low serum vitamin B12 levels (P less than .005), circulating intrinsic factor antibody (P less than .005), and anemia (P less than .025) was observed with stepwise increases in severity of atrophic gastritis. Subjects with atrophic gastritis exhibited a lower mean serum vitamin B12 level (P less than .05) and a higher mean folate level (P less than .05), but no difference was detected in mean hemoglobin levels or serum levels of iron, ferritin, retinol or alpha-tocopherol. It is concluded that serum pepsinogen I and pepsinogen II levels can be used to determine the prevalence and severity of atrophic gastritis, that atrophic gastritis is common in an elderly population, and that atrophic gastritis is associated with vitamin B12 deficiency and anemia. Further, higher folate levels in atrophic gastritis may be related to an accumulation of 5-methyl tetrahydrofolate in serum due to vitamin B12 deficiency and/or greater folate synthesis by the intestinal flora resulting from bacterial overgrowth secondary to hypo- or achlorhydria.

Human adult-onset lactase decline: an update.

Human adult-onset lactase decline is a biologic feature characteristic of the maturing intestine in the majority of the world's population. The digestion and absorption of lactose, the major carbohydrate in milk and also the main substrate for lactase, is often variable, a consequence of lactase levels, gastric emptying rate, and colonic salvage. Although commercially available "lactase" products alleviate symptoms in many lactose-intolerant people, a greater understanding of this variability in lactose tolerance could lead to interventions that reduce the rate of gastric emptying and/or increase the proliferation of lactose-metabolizing bacteria in the colon, leading to more efficient lactose utilization. Adult-onset lactase decline appears to be a risk factor for developing osteoporosis, owing to avoidance of dairy products or interference of undigested lactose with calcium absorption. Elderly with both adult-onset lactase decline and atrophic gastritis or those undergoing anti-ulcer treatment may have an increased risk of low calcium absorption owing to the lack of gastric acid that facilitates calcium uptake. Thus, lactose-intolerant elders should monitor their calcium nutrition status carefully.

Postprandial plasma vitamin A metabolism in humans: a reassessment of the use of plasma retinyl esters as markers for intestinally derived chylomicrons and their remnants.

We investigated postprandial vitamin A metabolism by measuring retinyl ester, triglyceride, and apolipoprotein (apo)B-48 in the plasma lipoproteins of human subjects before and after fat-feeding. Following a 14-hour fast, eight healthy subjects (two men, six women, 28 to 79 years) were given a fat-rich meal (1 g fat/kg body weight) containing vitamin A (40 retinol equivalents per kilogram body weight). Blood was collected every 3 hours for 12 hours and lipoproteins were isolated by sequential ultracentrifugation. Mean plasma retinyl ester concentration peaked 6 hours after the fat-rich meal, whereas mean plasma triglyceride peaked at 3 hours. Data obtained from hourly samples in 3 subjects showed that changes in the postprandial plasma concentration of retinyl ester occurred 1 to 2 hours after changes in the plasma triglyceride concentration. In triglyceride-rich lipoproteins (TRL) of d less than 1.006 g/mL, retinyl ester similarly peaked at 6 hours, whereas triglyceride as well as apoB-48 peaked at 3 hours. Although retinyl esters were found mainly in TRL in the initial postprandial period (84%, 3 hours; 83%, 6 hours), in fasting and postprandial plasma, particularly 9 or more hours after fat-feeding, a large percentage of plasma retinyl esters were in low-density lipoproteins (LDL) (44%, fasting; 9%, 3 hours; 9%, 6 hours; 19%, 9 hours; 32%, 12 hours). A small percentage of retinyl esters were also found in postprandial high-density lipoproteins (HDL) (2% to 7%). ApoB-48 was not detected in LDL of fasting or postprandial plasma.(ABSTRACT TRUNCATED AT 250 WORDS)

Role of triglyceride-rich lipoproteins from the liver and intestine in the etiology of postprandial peaks in plasma triglyceride concentration.

Plasma triglyceride concentration in human subjects peaks once, twice or three times in the twelve-hour period following the ingestion of a fat-rich meal. Triglyceride-rich lipoproteins (TRL) containing apolipoprotein (apo)B-48 (of intestinal origin), and TRL containing apoB-100 (predominantly of hepatic origin) both contribute to postprandial changes in plasma triglyceride concentration. To test the hypothesis that earlier peaks in postprandial triglyceridemia are due predominantly to the secretion of TRL from the intestine, while later peaks are due to the secretion of TRL from the liver, TRL apoB-48, TRL apoB-100 and retinyl ester (a marker of intestinal lipoproteins) were measured in plasma samples from subjects fed a fat-rich meal (1 g fat/kg body wt). Data from seven subjects (four fed 40 retinol equivalents vitamin A/kg body wt, three fed 20 retinol equivalents vitamin A/kg body wt, with the fat meal), showed that postprandial peaks in plasma triglyceride were always associated with increases in plasma retinyl ester concentration. In four subjects, who were selected because they had two clearly defined postprandial triglyceride peaks, the plasma concentration of TRL triglyceride, apoB-48, apoE and apoC increased in conjunction with both the earlier (three hour) and later (nine hour) peaks in plasma triglyceride. Increase in TRL apoB-100 was associated with both peaks in two of the four subjects. Our data suggest that 1) TRL from the liver and intestine contribute to both earlier and later peaks in postprandial triglyceridemia; and 2) the rate of appearance of TRL from the intestine is not constant after dietary fat absorption.

Total intestinal lactase and sucrase activities are reduced in aged rats.

Lactase-phlorizin hydrolase (LPH) and sucrase-isomaltase (SI) are intestinal microvillus membrane hydrolases that play important roles in carbohydrate digestion. Although the expression of these enzymes during postnatal development has been characterized, the effect of old age on disaccharidase activity is poorly understood. In the present investigation, we examined the effect of aging on lactase and sucrase activities and their mRNA levels in the small intestines of 3-, 12- and 24- mo-old rats by sampling from nine equidistant segments of small intestine. Total intestinal disaccharidase activity or mRNA abundance was determined from areas under the proximal-to-distal curves. Rats 24 mo of age had total intestinal lactase and sucrase activities that were 12 and 38% lower, respectively, than the 3-mo-old animals (P < 0.05). In contrast, total LPH and SI mRNA abundance did not change significantly. Thus, total intestinal lactase and sucrase activities decrease with age in a manner that likely involves a posttranscriptional process. The age-related decline in disaccharidase activity, if extrapolated to humans, may have important implications for the digestion of carbohydrate contained in the diet of the elderly.

Defective intracellular processing of lactase-phlorizin hydrolase protein in rats prenatally exposed to ethanol.

We have previously shown that fetal exposure to ethanol in rats produces both structural and biochemical abnormalities in absorptive enterocytes. Among the indicators of injury are derangements in the expression of lactase-phlorizin hydrolase (LPH), which is an essential enzyme for the assimilation of milk. In an animal model of fetal alcohol syndrome, unsuckled newborn rats prenatally exposed to maternal ethanol revealed a 10- to 15-fold increase in the number of LPH mRNA molecules per absorptive enterocyte, compared with controls (Estrada et al., Alcohol. Clin. Exp. Res. 20:1662-1668, 1996). However, lactase activity per cell was similar in both groups. The aim of this study was to characterize the effect of prenatal exposure to ethanol on the processing of LPH mRNA and protein. RNase protection assays using 3'- and 5'-directed antisense RNA probes revealed that the LPH mRNA from ethanol-exposed pups is full length. However, metabolic labeling, followed by immunoprecipitation using an anti-LPH monoclonal antibody, demonstrated a significant alteration in LPH protein processing. Intestinal explants from 21-day ethanol-exposed fetuses that were chased 30 min after a [35S]methionine pulse showed greater amounts of newly synthesized LPH precursors (205 and 220 kDa) and low molecular weight degradation products than controls. However, despite the increases in LPH precursor, the amount of 130 kDa mature LPH was similar in ethanol-exposed and control explants. These data suggest an increase in intracellular degradation of LPH precursor in rats prenatally exposed to ethanol, which occurs before its insertion into the microvillus membrane. Biosynthesis of LPH appears to be upregulated at the transcriptional level, which overcomes the degradation of LPH precursor during processing.

Differential activation of intestinal gene promoters: functional interactions between GATA-5 and HNF-1 alpha.

The effects of GATA-4, -5, and -6, hepatocyte nuclear factor-1 alpha (HNF-1 alpha) and -beta, and Cdx-2 on the rat and human lactase-phlorizin hydrolase (LPH) and human sucrase-isomaltase (SI) promoters were studied using transient cotransfection assays in Caco-2 cells. GATA factors and HNF-1 alpha were strong activators of the LPH promoters, whereas HNF-1 alpha and Cdx-2 were strong activators of the SI promoter, although GATA factors were also necessary for maximal activation of the SI gene. Cotransfection of GATA-5 and HNF-1 alpha together resulted in a higher activation of all three promoters than the sum of the activation by either factor alone, demonstrating functional cooperativity. In the human LPH promoter, an intact HNF-1 binding site was required for functional synergy. This study is the first to demonstrate 1) differential activation of the LPH and SI promoters by multiple transcription factors cotransfected singly and in combination and 2) that GATA and HNF-1 transcription factors cooperatively activate intestinal gene promoters. Synergistic activation is a mechanism by which higher levels of tissue-specific expression might be attained by overlapping expression of specific transcription factors.

Three distinct messenger RNA distribution patterns in human jejunal enterocytes.

BACKGROUND & AIMS: The importance of messenger RNA (mRNA) localization in human enterocytes is poorly understood. Previous studies from our laboratory have indicated that mRNAs are asymmetrically distributed in human intestinal epithelial cells, but in general colocalized with their encoded proteins. The aim of this study was to characterize, in human enterocytes, mRNA localization patterns of three genes with distinctly different functions. METHODS: mRNA distribution was determined by in situ hybridization with digoxigenin-labeled RNA probes in tissue sections of human jejunum. RESULTS: The mRNA for villin, a well-characterized microvillus cytoskeletal protein, was sorted to the basal region of the enterocyte. The mRNA for human sodium glucose cotransporter 1 was localized to the apical region, and the mRNA for human liver fatty acid-binding protein was distributed diffusely in the cytoplasm. CONCLUSIONS: The three distinct mRNA distribution patterns suggest that active mRNA sorting mechanisms exist in human enterocytes. This study also reveals for the first time that dichotomies may occur between the distribution patterns of sorted mRNAs and their encoded proteins.

Protein-energy malnutrition and liver dysfunction limit the usefulness of the relative dose response (RDR) test for predicting vitamin A deficiency.

The relative dose response (RDR) test was examined for its value in predicting vitamin A nutriture among chronically ill patients with protein-energy malnutrition and among patients with varying degrees of liver dysfunction. The RDR test was elevated (ie greater than 14 per cent) and, thus, predictive of vitamin A deficiency in only 4 out of 9 patients with combined vitamin A deficiency and protein-energy malnutrition. Among 27 patients with liver dysfunction who had vitamin A determinations performed on liver biopsy material, there was no correlation found between the RDR response and the hepatic vitamin A level. Of 12 patients with low hepatic vitamin A levels (less than 100 micrograms/g wet weight), the RDR test was normal in 9 (75 per cent). Inability of the RDR test to predict vitamin A nutriture in patients with protein-energy malnutrition and in patients with liver dysfunction may, in part, be due to inadequate synthesis or release of retinol-binding protein.

Folic acid malabsorption in atrophic gastritis. Possible compensation by bacterial folate synthesis.

Folic acid absorption was studied in 12 elderly subjects with atrophic gastritis and 10 elderly normal controls using tritium-labeled pteroylmonoglutamic acid. Two folic acid absorption tests were carried out on each subject with 120 ml of either water or 0.1 N HCl. Folic acid absorption was significantly lower in subjects with atrophic gastritis than in normal controls (31% vs. 51%, respectively; p less than 0.01). In subjects with atrophic gastritis, folic acid absorption rose significantly to 54% (p less than 0.001) when administered with acid, but did not change in normal controls (50%). Serum folate levels were normal in all subjects. Proximal small intestinal pH was higher in atrophic gastritis subjects than in normal controls (7.1 vs. 6.7, respectively; p less than 0.05), as were bacterial counts of small intestinal fluid (p less than 0.01). Bacteria cultured from the aspirates of subjects with atrophic gastritis were able to synthesize folate in vitro when incubated in a folate-free medium. Atrophic gastritis results in folic acid malabsorption but not in folate deficiency, possibly due to increased bacterial synthesis of folate in the small intestine.

Prenatal ethanol exposure alters the expression of intestinal hydrolase mRNAs in newborn rats.

To gain insight into the postnatal growth delay induced by ethanol in utero, we characterized functional impairments of the small intestine of neonatal rats prenatally exposed to ethanol using a well-described model of gestational alcoholism (25% ethanol w/v in the drinking water). Expression of the intestinal enzymes-lactase-phlorizin hydrolase (LPH) and intestinal alkaline phosphatase (IAP)-that are critical for enteral nutrition of neonates was studied. Characteristic patterns of LPH and IAP expression along the proximal-distal (horizontal) and crypt-villus (vertical) axes of the small intestine, as well as the intracellular localization of LPH and IAP mRNAs and immunoreactive proteins within absorptive enterocytes, were not altered by prenatal exposure to ethanol. However, a 10- to 15-fold increase in the number of LPH and IAP mRNA molecules per absorptive enterocyte was found throughout the intestine of ethanol-exposed neonates, compared with controls, whereas lactase and alkaline phosphatase activities per enterocyte remained unchanged. These findings suggest that ethanol in utero alters the mRNA abundance of epithelial enzymes in newborn rat small intestine. Changes in mRNA abundance could be an important aspect of enterocyte adaptation to high ethanol concentrations in gastrointestinal amniotic fluid of ethanol-exposed fetuses.