Lactase mRNA Research Articles

Influence of dietary fructose supplementation on visceral organ mass, carbohydrase activity, and mRNA expression of genes involved in small intestinal carbohydrate assimilation in neonatal calves

The hypothesis of this experiment was that dietary fructose would influence visceral organ mass, carbohydrase activity, and mRNA expression of carbohydrases and nutrient transporters in the small intestine in neonatal calves. Therefore, our objective was to use the neonatal calf as a model to evaluate the effects of postruminal fructose supply on small intestinal carbohydrate assimilation. Ten calves (<7 d of age; 41.2 ± 1.46 kg of body weight) were fed milk replacer at 2.0% of body weight daily (816 ± 90.5 g/d; 272 ± 30.1 g/L; dry-matter basis) in 2 equal portions and assigned to the following dietary treatment groups: (1) milk replacer (control; n = 6) or (2) milk replacer + 2.2 g of fructose/kg of body weight (fructose; n = 4). Calves were fed dietary treatments for 28 d, with jugular blood sampled every 7 d before and after the morning feeding. Calves were slaughtered, and visceral weights were recorded. Postruminal carbohydrase activities were assayed. Quantitative real-time PCR was conducted for small intestinal mRNA expression of nutrient transporters [solute carrier family 2 member 5 (GLUT5), solute carrier family 2 member 2 (GLUT2), and solute carrier family 5 member 1 (SGLT1)], carbohydrases (lactase, maltase-glucoamylase, and sucrase-isomaltase), and ketohexokinase (KHK). Data were analyzed using MIXED procedures in SAS version 9.4 (SAS Institute Inc, Cary, NC). Dietary fructose supplementation decreased serum glucose concentration. Small intestinal mass was greater in calves supplemented with fructose. Dietary fructose supplementation did not influence pancreatic α-amylase, small intestinal isomaltase, or maltase activities. Sucrase activity was undetected in the small intestine. Dietary fructose supplementation increased small intestinal glucoamylase activity per gram of tissue by 30% and increased maltase-glucoamylase mRNA expression by 6.8-fold. Dietary fructose supplementation did not influence mRNA expression of GLUT5, SGLT1, GLUT2, or KHK. Dietary fructose supplementation increased small intestinal lactase mRNA expression by 3.1-fold. Sucrase-isomaltase mRNA expression in the small intestine decreased 5.1-fold with dietary fructose supplementation. Dietary fructose supplementation does not induce sucrase activity in neonatal calves; however, sucrase-isomaltase may be transcriptionally regulated by dietary fructose in neonatal calves. More research is needed to compare glucose and fructose at isocaloric intakes to examine effects of dietary fructose at equal metabolizable energy intake.

Lactase nonpersistence is directed by DNA-variation-dependent epigenetic aging.

Inability to digest lactose due to lactase non-persistence is a common trait in adult mammals, with the exception of certain human populations that exhibit lactase persistence. It is not clear how the lactase gene can be dramatically downregulated with age in most individuals, but remains active in some. We performed a comprehensive epigenetic study of the human and mouse intestine using chromosome-wide DNA modification profiling and targeted bisulfite sequencing. Epigenetically-controlled regulatory elements were found to account for the differences in lactase mRNA levels between individuals, intestinal cell types and species. The importance of these regulatory elements in modulating lactase mRNA levels was confirmed by CRISPR-Cas9-induced deletions. Genetic factors contribute to epigenetic changes occurring with age at the regulatory elements, as lactase persistence- and non-persistence-DNA haplotypes demonstrated markedly different epigenetic aging. Thus, genetic factors facilitate a gradual accumulation of epigenetic changes with age to affect phenotypic outcome.

Ontogenic Changes of Villus Growth, Lactase Activity, and Intestinal Glucose Transporters in Preterm and Term Born Calves with or without Prolonged Colostrum Feeding.

Oral glucose supply is important for neonatal calves to stabilize postnatal plasma glucose concentration. The objective of this study was to investigate ontogenic development of small intestinal growth, lactase activity, and glucose transporter in calves (n = 7 per group) that were born either preterm (PT; delivered by section 9 d before term) or at term (T; spontaneous vaginal delivery) or spontaneously born and fed colostrum for 4 days (TC). Tissue samples from duodenum and proximal, mid, and distal jejunum were taken to measure villus size and crypt depth, protein concentration of mucosa and brush border membrane vesicles (BBMV), total DNA and RNA concentration of mucosa, mRNA expression and activity of lactase, and mRNA expression of sodium-dependent glucose co-transporter-1 (SGLT1) and facilitative glucose transporter 2 (GLUT2) in mucosal tissue. Additionally, protein expression of SGLT1 in BBMV and GLUT2 in crude mucosal membranes and immunochemical localization of GLUT2 in the enterocytes were determined. Villus height in distal jejunum was lower in TC than in T. Crypt depth in all segments was largest and the villus height/crypt depth ratio in jejunum was smallest in TC calves. Concentration of RNA was highest in duodenal mucosa of TC calves, but neither lactase mRNA and activity nor SGLT1 and GLUT2 mRNA and protein expression differed among groups. Localization of GLUT2 in the apical membrane was greater, whereas in the basolateral membrane was lower in TC than in T and PT calves. Our study indicates maturation processes after birth for mucosal growth and trafficking of GLUT2 from the basolateral to the apical membrane. Minor differences of mucosal growth, lactase activity, and intestinal glucose transporters were seen between PT and T calves, pointing at the importance of postnatal maturation and feeding for mucosal growth and GLUT2 trafficking.

First-pass uptake and oxidation of glucose by the splanchnic tissue in young goats fed soy protein-based milk diets with or without amino acid supplementation: Glucose metabolism in goat kids after soy feeding

The study was designed to examine whether feeding soy protein isolate as partial replacement of casein (CN) affects glucose metabolism in young goats and whether effects may be ameliorated by supplementation of those AA known to be lower concentrated in soy than in CN. Goat kids (d 20 of age) were fed comparable milk protein diets, in which 50% of the crude protein was either CN (control, CON), soy protein isolate (SPI), or soy protein isolate supplemented with AA (SPIA) for 43d (n=8 per group). On d 62 of age, a single bolus dose of d-[13C6]glucose (10mg/kg of BW) was given with the morning diet, and simultaneously, a single bolus dose of d-[6,6-2H2]glucose (5mg/kg of BW) was injected into a jugular vein. Blood samples were collected between −30 and +420min relative to the tracer administration to measure the 13C and 2H enrichments of plasma glucose and the 13C enrichment of blood CO2. Glucose first-pass uptake by the splanchnic tissues was calculated from the rate of appearance of differentially labeled glucose tracer in plasma. Glucose oxidation was calculated from 13C enrichment in blood CO2. In addition, plasma concentrations of triglycerides, nonesterified fatty acids, glucose, insulin, and glucagon were measured. On d 63 of age, kids were killed and jejunal mucosa and liver samples were collected to measure lactase mRNA levels and lactase and maltase activities in the jejunum and activities of pyruvate carboxylase and phosphoenolpyruvate carboxykinase (PEPCK) in the liver. Basal plasma glucose concentration tended to be higher in the CON than the SPIA group, whereas basal insulin was higher in the CON group than the SPI and SPIA groups, and glucagon was higher in the CON than the SPIA group. Plasma glucose and insulin concentrations increased during the first hour after feeding, whereas plasma glucagon increased immediately after feeding and after 1h of feeding. First-pass uptake and glucose oxidation were not affected by diet. Maltase activities in proximal and mid jejunum and lactase activities in mid jejunum were lower in the CON than in the SPIA group. Activities of PEPCK were higher in the SPIA than in the SPI group. In conclusion, feeding milk diets with soy protein isolate seems to affect glucose status in kids, but has no effect on first-pass uptake and oxidation of glucose. The highest activities of lactase and maltase were observed after supplementation with AA. Higher PEPCK activities in the liver may point at elevated gluconeogenic activities after AA supplementation in soy-fed kids.

Effect of C/T −13910 cis-acting regulatory variant on expression and activity of lactase in Indian children and its implication for early genetic screening of adult-type hypolactasia

Background Absorption of milk sugar (lactose) is regulated by the activity of lactase enzyme in gut wall. Intestinal lactase activity declines during childhood in majority of human populations leading to adult-type hypolactasia (primary lactose malabsorption), limiting the use of fresh milk due to lactose intolerance. Aim of this study was to correlate lactase expression and activity with C/T −13910 variant in Indian children, determine the age of onset of down-regulation of lactase activity and assess the applicability of the C/T −13910 variant as a diagnostic marker for identifying children genetically inclined to develop adult-type hypolactasia. Methods Intestinal biopsies were obtained from 176 children aged 1–16 years undergoing routine endoscopy for various abdominal complaints. The biopsies were assayed for lactase, sucrase and maltase activities and genotyped for C/T 13910 variant using PCR-RFLP analysis. The functional effect of the C/T − 13910 variant on expression of lactase mRNA and protein in these children was examined using reverse-transcription PCR and western blotting. Results Among the 176 children investigated in our study, 56.8% (100/176) carried the C/C − 13910 genotype, which has been associated with the onset of adult-type hypolactasia, while 40.9% (72/176) carried the C/T − 13910 genotype and 2.3% (4/176) the T/T − 13910 genotype. There was a significant correlation between lactase activity and C/T − 13910 variant ( P < 0.001). The mean level of lactase activity among children with C/C − 13910 genotype was 15.9 U/g protein and with C/T and T/T − 13910 genotypes was 30.9 U/g protein. The age of onset of down-regulation of lactase activity in children with C/C − 13910 genotype was between 3 and 5 years and keeping 10 U/g protein lactase activity as cut off, adult-type hypolactasia was evident in all the individuals > 8 years of age for this genotype. C/C − 13910 genotype was associated with low expression of lactase mRNA and protein compared with C/T genotype. Considering lactase activity of 10 U/g protein as gold standard, predictive value of genetic test based on C/T − 13910 variant for adult-type hypolactasia was 100% in children > 8 years of age. Conclusion C/T − 13910 cis-acting regulatory variant located ≈ 14 kb upstream of lactase gene (LCT) completely correlates with lactase phenotype in Indian children. The genetic testing for the C/T − 13910 variant may be helpful in the diagnosis of adult-type hypolactasia in Indian children.

The small intestinal lactase expression is reduced in the piglet with bowel inflammation

The effects of dextran sodium sulfate (DSS) induced bowel inflammation on jejunal lactase activity and expression were examined in 12 Yorkshire piglets at 5 d of age with body weight (BW) of 3 kg were fitted with ig catheters and divided into control (CON) and treatment groups (DSS). Both groups of pigs were infused with an equal volume of either saline or 1.25 g DSS/kg BW.d in saline, respectively, for 10 d. Lactase protein abundance was carried out by Western blotting and the mRNA abundance was measured by quantitative real time RT‐PCR. â‐actin was used as the housekeeping gene. DSS treatment decreased (P&lt;0.05) jejunal lactase maximal specific activity (Vmax, µmol/mg protein.min) in the tissue homogenate (CON, 87.66±0.14 vs. DSS, 24.05±0.30) by 73%, cytosol (CON, 45.83±0.68 vs. DSS, 12.58±1.15) by 73% and apical membrane (CON, 422.67±3.29 vs. DSS, 92.22±2.42) by 78%, respectively, with orresponding decreases (P&lt;0.05) of 26, 48, and 32% in the jejunal lactase protein abundance. The relative abundance of the jejunal lactase mRNA was lower by 25% in the DSS piglets. Bowel inflammation reduced the lactase activity in the small intestine by lowering its expression at the transcriptional and translational levels. Supported by AFMNet, OMAFRA and NSERC of Canada.

M1731 Sphingosine-1-Phosphate Regulates Barrier Function in Cultured Intestinal Epithelial Cells and Murine Intestinal Tissue

assessed by stable isotope methodology after administration of U-13C-glucose and 1-13Clactose. Concentrations of the EFA linoleic acid (LA), and the enzyme activity and mRNA expression of lactase, were measured in the mucosa of proximal, mid and distal small intestine. Results: Mice fed the EFA-deficient diet were markedly EFA-deficient (triene/ tetraene ratio 0.23±0.06 vs. 0.01±0.00 in controls, p<0.05) with a profound lower fat absorption of dietary fat (81±3 vs. 99±0% in controls, of ingested amount, p<0.05). EFA deficiency did not affect the histology or proliferative capacity of the small intestine as demonstrated by the histological and proliferative staining, and by the morphometrcal measurements of the small intestinal villi. Blood C6-glucose appearance and disappearance were similar in both groups, indicating unaffected monosaccharide absorption. In contrast, blood appearance of 13C-glucose, originating from 13C-lactose, was delayed in EFA-deficient mice. EFA deficiency profoundly reduced lactase activity (-58%, p<0.01) and mRNA expression (-55%, p<0.01) in mid small intestine. Both lactase activity and its mRNA expression strongly correlated with mucosal LA concentrations (r=0.89 and 0.79, resp., p< 0.01). Conclusions: EFA deficiency in mice inhibits the capacity to digest lactose, but does not affect small intestinal histology. These data underscore the observation that EFA deficiency functionally impairs the small intestine, possibly mediated by low LA levels in the enterocytes. This research was funded by the Dutch Digestive Foundation (MWO 04-38).

Study on the regulation of intestinal lactase gene during postnatal development in rats

Maturation decline in lactase activity is complex phenomenon which is either due to shift in lactase synthesis and processing to the inactive 100Kda and Increased degradation of the lactase activity of 130kda subunit, or decline in the enterocyte life span.lactase phlorizin hydrlase shifts from a high sialylation status during weaning to high fucosylation level in adult rats. The activities of brush border lactase were assayed. sialic acid and fucose were estimated. Membrane protein were resolved by SDS‐PAGE on to nitocellulose paper/PVD membrane and immunostained with polyclonal antibodies against lactase. Total RNA was isolated and transfered RNA to nylon membrane.Determination of fucose to sialic acid ratio revealed close parllelism between fucosylation and sialylation of microvillus membranes with decline in intestinal lactase activity with development age in rat. Upstream region of prolactase gene, suggesting the absence of functional role in mature lactase mRNA and also mRNA is poorly translated to active lactase in the adult intestine which may be due to its interaction with 100kda protein product of prolactase hydrolysis. Western blot analysis revealed the presence of 100kda protein in rat intestine during postnatal development.

Decreased sucrase and lactase activity in iron deficiency is accompanied by reduced gene expression and upregulation of the transcriptional repressor PDX-1

Disaccharidases are important digestive enzymes whose activities can be reduced by iron deficiency. We hypothesise that this is due to reduced gene expression, either by impairment to enterocyte differentiation or by iron-sensitive mechanisms that regulate mRNA levels in enterocytes. Iron-deficient Wistar rats were generated by dietary means. The enzyme activities and kinetics of sucrase and lactase were tested as well as the activity of intestinal alkaline phosphatase (IAP)-II because it is unrelated to carbohydrate digestion. mRNA levels of beta-actin, sucrase, lactase, and the associated transcription factors pancreatic duodenal homeobox (PDX)-1, caudal-related homeobox (CDX)-2, GATA-binding protein (GATA)-4, and hepatocyte nuclear factor (HNF)-1 were measured by real-time PCR. Spatial patterns of protein and gene expression were assessed by immunofluorescence and in situ hybridization, respectively. It was found that iron-deficient rats had significantly lower sucrase (19.5% lower) and lactase (56.8% lower) but not IAP-II activity than control rats. Kinetic properties of both enzymes remained unchanged from controls, suggesting a decrease in the quantity of enzyme present. Sucrase and lactase mRNA levels were reduced by 44.5% and 67.9%, respectively, by iron deficiency, suggesting that enzyme activity is controlled primarily by gene expression. Iron deficiency did not affect the pattern of protein and gene expression along the crypt to villus axis. Expression of PDX-1, a repressor of sucrase and lactase promoters, was 4.5-fold higher in iron deficiency, whereas CDX-2, GATA-4, and HNF-1 levels were not significantly different. These data suggest that decreases in sucrase and lactase activities result from a reduction in gene expression, following from increased levels of the transcriptional repressor PDX-1.

In vitro translation of RNA to lactase during postnatal development of rat intestine

mRNA levels encoding lactase were detected by Northern blot analysis using two different probes in developing rat intestine. Probe I and probe II corresponding to second half of prolactase gene showed a 6.8 kb mRNA transcript in 7, 14, 21 and 30 day old rat intestine. There was no change in quantity of lactase mRNA detected using probe II, but hybridization with probe I showed a progressive decrease in mRNA transcript encoding lactase with age. At day 7 and 14 of postnatal development, the lactase mRNA was quite high, but it reduced upon weaning. The in vitro translation products of RNA detected by Western blot analysis using brush border lactase antibodies showed several isoforms of lactase antigen with molecular weight ranging from 100-220 kDa. Analysed at days 7 and 30 of postnatal development, lactase isoforms of molecular weight 130 kDa and 220 kDa were similar to those found in purified brush border membranes. The translation of RNA to 220 kDa lactase protein was high in 7 and 14 day old pups, but it was markedly reduced in 30 day old animals. These findings support the contention that translation of mRNA to lactase is impaired in weaned animals, which may also be responsible for the maturational decline in lactase activity in adult rat intestine.

Real-Time PCR Quantification of Bovine Lactase mRNA: Localization in the Gastrointestinal Tract of Milk-Fed Calves

Lactase is a disaccharidase that is present in the brush-border membrane of the small intestine, hydrolyzes lactose to glucose and galactose, and is therefore important in milk-fed animals. Assays based on quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR) in the bovine species have not yet been described. Therefore, we have developed an RT-PCR assay for the quantification of lactase mRNA levels and have tested its suitability in the bovine gastrointestinal tract of seven 5-d-old milk-fed calves. Primers for RT-PCR amplification of bovine lactase mRNA were designed in the 100% identical regions of species (rats, rabbits, humans) from which lactase sequences were available. Lactase mRNA was expressed relative to mean levels of 4 housekeeping genes (glyceraldehyde-3-phosphate dehydrogenase, β-actin, ubiquitin, and 18S). The presence of lactase mRNA along the entire gastrointestinal tract was evaluated in samples that consisted of whole gut walls (mucosa plus submucosa). Furthermore, mRNA levels of lactase were measured in fractionized layers of jejunal and ileal mucosa (mainly containing villus tips or crypts) and ileal lamina propria (mainly containing Peyer's patches). Agarose gel electrophoresis of the lactase PCR product revealed a single band that corresponded to the single-amplified product as predicted by the melting curve analysis of the PCR. The amplified partial-bovine lactase sequence showed 87% similarity with human and rabbit sequences and 82% similarity with the rat sequence. Lactase mRNA was present in whole walls (consisting of mucosa and submucosa) of the entire small intestine, but was absent in esophagus, rumen, fundus, pylorus, and colon. Furthermore, lactase mRNA was detected in fractionized villus and crypt layers of jejunum and ileum, but levels were higher in the jejunum in villus than in crypt fractions. No lactase mRNA was detectable in the lamina propria fraction of the ileum containing mainly Peyer's patches. In conclusion, the developed RT-PCR method allows study of lactase mRNA levels.

Abundance of mRNA encoding for components of the somatotropic axis and insulin receptor in different layers of the jejunum and ileum of neonatal calves1,2

Insulin-like growth factors-1 and -2, IGFBP-2 and -3, and receptors for IGF type-1 and type-2 (IGF-1R, IGF-2R), growth hormone (GHR), and insulin (InsR) in neonatal calves are variably expressed among gastrointestinal sites and thought to exert site-specific physiological functions. We studied by real-time reverse-transcription PCR, whether there are differences in the abundance of mRNA coding for IGF-I, IGF-2, IGFBP-2, IGFBP-3, IGF-1R, IGF-2R, GHR, and InsR in compartmentalized layers (fractions) of jejunum and ileum of 5-d-old calves fed colostrum. Samples of jejunum consisted primarily of villi and crypts; samples from ileum consisted mainly of villus tips, crypts, and lamina propria (LP; containing mainly Peyer's patches). After slaughter, segments of middle areas of jejunum and ileum were flushed with 154 mM NaCl. Pieces (5 mm x 5 mm) of jejunal (n = 9) and ileal walls (n = 5) were placed on glass slides and snap-frozen in liquid N before being cut horizontally into 10-mum-deep slices using a cryotome at -20 degrees C. Fifteen consecutive and morphologically similar slices were collected as fractions of villus, crypt, and LP layers, respectively. Fractions were characterized by use of 5'-bromo-2-deoxyuridine (BrdU) that labeled proliferating cells, and by expression of lactase mRNA. The BrdU-labeled cells were present in crypts and LP, but not in tips of villi. Lactase mRNA levels were greater in villus than crypt fractions, but lactase mRNA was absent in LP. In jejunum, mRNA levels, relative to levels of housekeeping genes (sum of levels of mRNA coding for ubiquitin, glyceraldehyde phosphate dehydrogenase, beta-actin, and ribosomal RNA), differed (P < 0.05) between fractions for InsR (crypts > villi), IGFBP-2 (crypts > villi), and IGFBP-3 (crypts > villi), and total RNA levels were greater (P < 0.05) in crypt than villus fractions. In ileum, mRNA levels, expressed relative to housekeeping genes, differed (P < 0.05) between fractions for IGF-I (LP > villi, crypts), IGF-2, and IGFBP-3 (villi > crypts, LP), GHR and InsR (crypts > LP), IGFBP-2 (crypts > villi, LP), and total RNA levels were greater (P < 0.05) in LP and crypt than in villus fractions. In conclusion, the tested fractionation technique is quite applicable for gene expression studies in the intestine of calves. Members of the somatotropic axis and of the insulin receptor are not equally expressed in different jejunal and ileal layers of neonatal calves.

Lactase Gene Transcription Is Activated in Response to Hypoxia in Intestinal Epithelial Cells

Lactase-phlorizin hydrolase, a brush-border membrane disaccharidase, is a marker of intestinal epithelial cell differentiation and digestive function. The intestine is susceptible to conditions of hypoxia resulting from vascular perfusion deficits. We hypothesized that lactase gene induction may provide a mechanism to efficiently increase nutrient energy substrates during gut hypoxia. These studies sought to characterize expression of the lactase gene in response to hypoxia and to characterize a role for hypoxia-inducible factor (HIF-1) in mediating the hypoxic response. Microarray analysis and confirmatory RT-PCR identified a 4-fold induction of lactase mRNA abundance in intestinal epithelial Caco-2 cells exposed to hypoxia. Lactase promoter activity was similarly induced by hypoxia in cells stably transfected with a 2.0-kb 5′-flanking region of the rat lactase gene linked to a reporter gene. Transient cotransfection with HIF-1α and β stimulated lactase promoter activity 2.4- and 3.5-fold under conditions of normoxia and hypoxia, respectively. We conclude that HIF-1 can activate the lactase promoter in intestinal epithelial cells exposed to hypoxia. Induction of lactase transcription may represent an adaptive response to gut hypoxia.

Changes in cell proliferation and differentiation of adult rat small intestine epithelium after adrenalectomy: kinetic, biochemical, and morphological studies.

The effects of 10-day bilateral adrenalectomy on morphometry, proliferation, and apoptosis were determined in the small intestine of 3-month-old Sprague-Dawley rats. The activities of sucrase, lactase, and its respective mRNA, aminopeptidase N, and intestinal alkaline phosphatase were also evaluated. Adrenalectomy lead to partial atrophy and disorganization of the epithelium, with an increased number of goblet and Paneth cells and a reduction of crypt cell proliferation paralleled by a marked increase in villus apoptosis. Biochemical assays revealed that aminopeptidase N and intestinal alkaline phosphatase activities were significantly decreased, whereas disaccharidases were increased by adrenalectomy. The corresponding induction of lactase mRNA suggests an active response of the epithelium. In conclusion, adrenalectomy modified maturation and the differentiation processes of the small intestinal mucosa, especially in the proximal part of the small intestine. This result points to an important role of adrenals and glucocorticoids in the trophic status of the adult small intestinal mucosa.

Enteral Insulin‐like Growth Factor‐I Augments Intestinal Disaccharidase Activity in Piglets Receiving Total Parenteral Nutrition

ABSTRACTBackground:Partial enteral nutrition is administered to infants on parenteral nutrition to stimulate intestinal function. Herein, the hypothesis that supplementation of partial enteral nutrition with insulin‐like growth factor‐I would augment intestinal development was investigated.Methods:One‐day‐old piglets (n = 29) were randomly assigned to five dietary treatment groups: 100% of energy as enteral formula, 100% of energy as total parenteral nutrition, or 80% parenteral nutrition/20% enteral formula supplemented with either 0, 0.2, or 1 mg/kg insulin‐like growth factor‐I for 7 days. Weight gain, intestinal weight, morphology, protein, and DNA content and disaccharidase activity and mRNA expression were assessed.Results:Parenterally fed piglets had similar whole body weight gain and serum hormone concentrations but reduced intestinal mucosal weight, villus height, and sucrase and lactase activity compared with 100% enterally fed pigs. Partial enteral nutrition alone increased mucosal weight and protein content, villus height, and disaccharidase activity compared with 100% parenterally fed piglets. No effect of the lower dose of insulin‐like growth factor‐I (0.2 mg/kg per day) was observed, but supplementing partial enteral nutrition with 1 mg insulin‐like growth factor‐I/kg further increased villus width and cross‐sectional area and disaccharidase activity compared with partial enteral nutrition alone. Lactase mRNA expression was not affected by insulin‐like growth factor‐I, suggesting that the primary site of regulation of lactase by insulin‐like growth factor‐I occurs after transcription.Conclusions:Enteral insulin‐like growth factor‐I augmented intestinal morphology and disaccharidase activity in parenterally fed piglets over that observed with partial enteral nutrition alone. Thus enteral insulin‐like growth factor‐I may represent an efficacious clinical adjunct to promote intestinal development of parenterally fed neonates.

Regulation of lactase and sucrase-isomaltase gene expression in the duodenum during childhood.

In children, lactase and sucrase-isomaltase are essential intestinal glycohydrolases, and insufficiency of either enzyme causes diarrhea and malnutrition. Little is known about the regulation of lactase and sucrase-isomaltase expression in the duodenum during childhood. In this study, the mechanisms of regulation of duodenal expression of both enzymes were examined in a study population with ages ranging from 1 to 18 years. Duodenal biopsy specimens from 60 white children were used to analyze tissue morphology and to quantify lactase and sucrase-isomaltase mRNA and protein. Among healthy subjects, high interindividual variability was noted in both mRNA and protein levels for lactase and sucrase-isomaltase. Lactase mRNA level per subject did not correlate with sucrase-isomaltase mRNA level and thus appeared independent. Both lactase and sucrase-isomaltase protein levels correlated significantly with their respective mRNA levels. For each enzyme, a significant inverse correlation was observed between the degree of villus atrophy and mRNA levels. Aging from 1 to 18 years did not result in significant changes in mRNA or protein levels of either enzyme. Immunostaining patterns within the duodenal epithelium for lactase differed from sucrase-isomaltase in adjacent sections, illustrating independent regulation at the cellular level. In the duodenum of white children, lactase and sucrase-isomaltase seem primarily regulated at the transcriptional level. The expression of each enzyme in the intestinal epithelium is regulated by an independent mechanism. Lactase and sucrase-isomaltase exhibit stable mRNA and protein levels in healthy children as they grow to adulthood. Mucosal damage affected levels of both enzymes negatively.

The genetically programmed down-regulation of lactase in children

Background & Aims: Intestinal lactase activity is high in all healthy human babies, but in adults a genetic polymorphism, which acts in cis to the lactase gene, determines high or low messenger RNA (mRNA) expression and activity (lactase persistence and nonpersistence, respectively). Our aim was to investigate the onset of expression of this polymorphism in children. Methods: Activities were analyzed in relation to age in normal biopsy specimens from a 20-year collection of diagnostic specimens. In a smaller set of 32 samples, aged 2–132 months, RNA was extracted for semiquantitative reverse-transcription polymerase chain reaction. Marker polymorphisms were used to determine the allelic origin of lactase mRNA transcripts. Results: Analysis of 866 children showed evidence that the lactase persistence/nonpersistence polymorphism began before 5 years of age. The 32 children tested had high lactase mRNA and activity. Six children aged 2–16 months showed equal expression of two alleles, 2 children aged 7 and 14 months showed slightly asymmetric expression, and 7 children aged 22–132 months showed very asymmetric expression, the second allele being undetectable in the 11-year-old, as previously seen in lactase-persistent heterozygote adults. Conclusions: Genetically programmed down-regulation of the lactase gene is detectable in children from the second year of life, although the onset and extent are somewhat variable. GASTROENTEROLOGY 1998;114:1230-1236

Oral insulin is biologically active on rat immature enterocytes.

Oral insulin is biologically active on rat immature enterocytes.

Lactase persistence versus decline in human adults: Multifactorial events are involved in down-regulation after weaning

BACKGROUND & AIMS: In nonhuman mammals, lactase activity declines during or after weaning. In contrast, about one half of the human species maintains high lactase activity even in adulthood. To clarify this difference, this study examined some parameters for which contrasting observations have been reported in connection with lactase decline. METHODS: Lactase activity, lactase messenger RNA (mRNA) levels, and in vitro lactase biosynthesis were determined in normal jejunal samples from a large group of white adults, all born in or near Naples. RESULTS: Of 44 individuals, 10 were lactase persistent and 34 were hypolactasic. Biosynthesis of prolactase correlated well with lactase mRNA levels, indicating transcriptional control; it did less so with steady-state lactase activity. Examination of lactase mRNA levels and lactase activity/lactase mRNA ratios revealed a heterogeneous pattern of lactase mRNA level, lactase synthesis, and activity in both lactase persistent and hypolactasic subjects. CONCLUSIONS: Both transcriptional and posttranscriptional factors cause the decline of intestinal lactase. This probably explains the multifarious observations that most studies on adult-type hypolactasia have reported. The single overriding factor distinguishing lactase- persistent subjects from hypolactasic subjects is the high rate of lactase biosynthesis. (Gastroenterology 1997 May;112(5):1506-14)

Effects of malnutrition on expression and activity of lactase in children

BACKGROUND & AIMS: Many malnourished infants have reduced lactase specific activity in the small intestine. The aim of this study was to test the hypothesis that the hypolactasia of malnourished infants results from transcriptional suppression of lactase expression.METHODS: Biopsy specimens were studied from two groups of infants: 29 with malnutrition and 10 normally nourished controls with normal morphology and lactase activity.RESULTS: In malnourished infants, lactase messenger RNA (mRNA) was reduced to 32% and sucrase to 61% of normal. Lactase and sucrase enzyme proteins and activities were lower in malnourished infants, and partial villus atrophy was present. The genotype of adult hypolactasia was not present.CONCLUSIONS: Because the hypolactasia of malnourished children was associated with much lower lactase than sucrase mRNA abundance and because the epigenetic suppression, which accounted for the reduction of sucrase mRNA, was inadequate to explain the greater reduction of lactase mRNA, this study concludes that malnutrition suppresses lactase gene transcription or mRNA stability in infants. The reductions of lactase mRNA, distinct from those found in adults with genetic hypolactasia, explain the low lactase activities commonly found in malnourished infants.(Gastroenterology 1997 Mar;112(3):742-51)