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

Summary 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.     

Acute dieldrin toxicity: effect on the uptake of glucose and leucine and on brush border enzymes in monkey intestine

Administration of a single oral dose of dieldrin (20 mg/kg body wt.) to rhesus monkeys considerably elevated the uptake of glucose and the activities of brush border sucrase, lactase, maltase and alkaline phosphatase in intestine compared to control animals. Leucine uptake and leucine amino peptidase activity was significantly depressed in pesticide-treated animals. Kinetic studies with brush border sucrase revealed that augmentation of enzyme activity in pesticide-fed animals was due to an increase in the disaccharidase content.     

Expression of sodium-glucose co-transporter and brush border disaccharidases in Giardia lamblia infected rat intestine

The absorption of D-glucose and brush border membrane disaccharidases in the intestine of rat during infection by Giardia lamblia has been studied. The level of mRNA encoding Na+/glucose co-transporter (SGLT1) and brush border sucrase and lactase activities were also analyzed. At the peak of infection, i.e, day 7, 11 and 15 post-infection, there was a marked decrease in the signal of 4.5 kb and 2.8 kb mRNAs encoding SGTL1 compared to the controls. A similar decrease in sucrase and lactase mRNA's (6.5 kb and 6.8 kb respectively) was also observed under these conditions. This corresponds to observed decrease in the rate of Na(+)-dependent D-glucose uptake and low activities of brush border sucrase and lactase under these conditions. There was no change in Na(+)-independent D-glucose uptake in giardia infected rat intestine. These findings suggest that the down regulation of the expression of SGLT1 and brush border sucrase and lactase activities may be responsible for the observed malabsorption in G. lamblia infection.     

Sugar hydrolases of the infant rat intestine and their arrangement on the brush border membrane

Lactase and maltase, the predominant sugar hydrolases associated with the intestinal brush border membrane of the suckling rat, were purified essentially free of the other to near homogeneity (lactase at specific activity 23, maltase at specific activity 58), and their specific physicochemical properties determined. Antisera prepared to each showed by immunodiffusion a single common precipitin line with pure enzyme and solubilized proteins of the brush border membrane. Brush border membranes were purified 26–35-fold from infant rat intestine. Membranes prepared from 10-day-old rats contained 32% protein, 43% lipid and 25% carbohydrate with lactase and maltase estimated to comprise in excess of 10% and 2%, respectively, of the membrane protein.Immunotitration curves of lactase and maltase showed equivalent antibody binding by the membrane-bound and free enzyme forms. Furthermore, antibody binding to one enzyme did not affect the immunotitration curve or the extractability (by papain or Triton X-100) of the other membrane-bound enzyme. It was concluded that the lactase and maltase molecules are attached singly on the external membrane surface in a spatially independent manner with their antigenic sites as freely available to antibody binding as exhibited by their papain-solubilized counterparts.     

Sugar hydrolases of the infant rat intestine and their arrangement of the brush border membrane.

Lactase and maltase, the predominant sugar hydrolases associated with the intestinal brush bordermembrane of the suckling rat, were purified essentially free of the other to near homogeneity (lactase at specific activity 23, maltase at specific activity 58), and their specific physiocochemical properties determined. Antisera prepared to each showed by immunodiffusion a single common precipitin line with pure enzyme and solubilized proteins of the brush border membrane. Brush border membranes were purified 26--35-fold from infant rat intestine. Membranes prepared from 10-day-old rats contained 32% protein, 43% lipid and 25% carbohydrate with lactase and maltase estimated to comprise in excess of 10% and 2%, respectively, of the membrane protein. Immunotitration curves of lactase and maltase showed equivalent antibody binding by the membrane-bound and free enzyme forms. Furthermore, antibody binding to one enzyme did not affect the immunotitration curve or the extractability (by papain or Triton X-100) of the other membrane-bound enzyme. It was concluded that the lactase and maltase molecules are attached singly on the external membrane surface in a spatially independent manner with their antigenic sites as freely available to antibody binding as exhibited by their papain-solubilized counterparts.     

Stimulation of lactase synthesis induced by starvation in the jejunum of adult rat

The effects of actinomycin D and of cycloheximide administration have been investigated on the enzyme activities of the jejunal brush border membrane in adult rats after a 48-hour period of starvation. The modifications in the protein and enzyme patterns of the brush border membrane and the incorporation of radiolabelled amino acid in the protein band corresponding to lactase have been studied in the nourished and in the starved animal. The results show that actinomycin D administration did not modify the stimulation of lactase activity caused by starvation whereas cycloheximide completely inhibited this process. The stimulation of lactase activity, in the starved animal, is related to a quantitative increase of the corresponding protein band and with enhanced incorporation of L-[3H]valine in this protein band after separation of brush border proteins by gel electrophoresis. It is concluded that the stimulation of lactase activity observed during starvation is the consequence of de novo synthesis of lactase molecules and that this process is regulated at a translational level. A general hypothesis is proposed in order to clear up partly the mechanism involved in the stimulation of lactase activity by food deprivation in the adult rat.     

Enzymic solubilization of the human intestinal brush border membrane enzymes

The releases of proteins, maltase, lactase, sucrase, trehalase, alkaline phosphatase, γ-glutamyltransferase and leucylnaphthylamide-hydrolyzing activity from human intestinal brush border membrane vesicles by various enzymes (especially pancreatic proteases) have been studied.The brush border membrane enzymes are not solubilized by digestion with trypsin and chymotrypsin but are largely released after treatment with papain or elastase. Most of the enzymes are fully active after the proteolytic treatment. All proteins released by papain and elastase have been identified by electrophoresis to already known intestinal hydrolases.Electron microscopy of brush border membrane vesicles demonstrates “knob-like” structures (particles) attached to the external side of the membrane. During papain treatment, enzyme removal runs parallel with the disappearance of the particles. During elastase treatment it is not possible to correlate the release of th enzymic activities with the removal of the particles.The results indicate that most of the intestinal hydrolases are surface components attached to the external side of the membrane. They are in accord with the concept that the brush border membrane enzymes are organized within the membrane in a mosaic-like pattern.     

Effect of tannic acid on brush border disaccharidases in mammalian intestine

Tannic acid is a glucoside (penta-m-digallolyl-glucose), which exhibits a wide variety of physiological functions. Around neutral pH, 0.4 mM tannic acid produced 84% inhibition of rat brush border sucrase activity, but 35-40% enzyme inhibition was observed in the rabbit intestine at 0.08 mM concentration. In the mice, 74-77% enzyme inhibition was observed at 0.05 mM concentration of tannic acid. The observed inhibition was reversible in rat intestine. Tannic acid (0.2 mM) also inhibited lactase (18% in adult and 71% in suckling animals), maltase (76%) and trehalase (88%) activities in rat intestine. pH versus activity curves showed that 0.2 mM tannic acid inhibited enzyme activity in rat by 91% at pH 5.5 which was reduced to 14% at pH 8.5 compared to the respective controls. In the rabbit 18-60% enzyme inhibition was noticed below pH 7.0, however at pH 8.5, it was of the order of 38%. Kinetic analysis revealed that tannic acid is a competitive inhibitor of rat brush border sucrase at pH 6.8. Effect of tannic acid together with various -SH group reacting reagents revealed that the enzyme inhibition is additive in nature, suggesting the distinct nature of binding sites on the enzyme for these compounds. The results suggest that tannic acid is a potent inhibitor of intestinal brush border disaccharidases, and could modulate the intestinal functions.     

Enzymic solubilization of the human intestinal brush border membrane enzymes.

The releases of proteins, maltase, lactase, sucrase, trehalase, alkaline phosphatase, gamma-glutamyltransferase and leucylnaphthylamide-hydrolyzing activity from human intestinal brush bborder membrane vesicles by various enzymes (especially pancreatic proteases) have been studied. The brush border membrane enzymes are not solubilized by digestion with trypsin and chymotrypsin but are largely released after treatment with papain or elastase. Most of the enzymes are fully active after the proteolytic treatment. All proteins released by papain and elastase have been identified by electrophoresis to already known intestinal hydrolases. Electron microscopy of brush border membrane vesicles demonstrates "knob-like" structures (particles) attached to the external side of the membrane. During papain treatment, enzyme removal runs parallel with the disappearance of the particles. During elastase treatment it is not possible to correlate the release of the enzymic activities with the removal of the particles. The results indicate that most of the intestinal hydrolases are surface components attached to the external side of the membrane. They are in accord with the concept that the brush border membrane enzymes are organized within the membrane in a mosaic-like pattern.     

Developmental changes in intestinal brush border enzymes of rats prenatally exposed to ethanol

The activities of lactase, sucrase, alkaline phosphatase (AP) and y-glutamyl transpeptidase (gamma-GTP) were studied in the intestinal brush border membranes of pups born to rat mothers exposed to ethanol (1 ml of 30% ethanol daily during gestation) at different days of postnatal development. The activities of lactase (at day 4-20) and sucrase (at day 20-30) were considerably reduced in response to prenatal exposure to ethanol, while AP (at day 4-30) and gamma-GTP activities were significantly enhanced (p < 0.05) at day 4, 8, 14 and 20, but there was no significant difference by day 30 of postnatal development. The observed changes in enzyme activities were corroborated by Western blot analysis of lactase, sucrase and AP. Kinetic studies revealed a change in Vmax without affecting apparent Km of enzymes under these conditions. The present findings suggest that in utero ethanol exposure to rats is embryotoxic and affects the postnatal development of various brush border enzymes, which persist long after the ethanol was withdrawn prior to birth.     

Subcellular distribution of digestive enzymes in Ascaris suum intestine

Van den Bossche H. and Borgers M. 1973. Subcellular distribution of digestive enzymes in Ascaris intestine. International Journal for Parasitology3: 59–65. The microvilli of the intestinal cells of Ascaris suum resemble the microvilli of the mammalian intestine in respect to their morphologic structure; their behaviour to homogenization in the presence of a chelating agent; the presence of the disaccharide hydrolases, maltase, sucrase and trehalase and the presence of an enzyme which hydrolyses 5′-AMP at neutral pH. The microvilli of the Ascaris intestinal cells differ completely from those present in mammalian intestine in respect to the presence of non-specific phosphatases. The brush border fraction contains the bulk of acid phosphatase present in the intestinal cells. Although some pinocytotic vesicles have been observed only low endocytotic activity was found. We therefore suggest that the acid hydrolases found on the brush border membrane may be functionally related to extracellular digestion of macromolecules.     

Ontogenic development of intestinal disaccharidases in the precocial rodent Octodon degus (Octodontidae)

We studied the ontogeny of the intestinal brush border disaccharidases sucrase and lactase in the precocial rodent Octodon degus. Sucrase hydrolyze sugars from plants while lactase hydrolyzes sugars from milk. Enzyme expression varied inversely with dietary changes according to the developmental pattern. All new-born pups had high lactase and low sucrase activities. Also, a negative correlation between sucrase and lactase activity was found, supporting the economic design hypothesis for the intestinal tract. Profiles for development of sucrase expression exhibit some differences among precocial species, and in O. degus is correlated with the slower transition from milk to solid food consumption at weaning.     

Influence of duodenal secretions and its components on release and activities of human brush-border enzymes

The in vitro effects of human duodenal secretions and various combinations of its components on activity and release of enzymes from the human brush border were examined. Sucrase retained activity for 90 min in duodenal secretions, and maltase was almost as stable; lactase lost activity rapidly and alkaline phosphatase was of intermediate stability. Inactivation of lactase could only be partly (50%) attributed to luminal proteases, bile salts and phospholipids played no role. Rate of release of an enzyme from the brush border bore no relationship to its rate of inactivation. When individual proteases were studied, elastase was the most potent for releasing disaccharidases from the brush border; trypsin was ineffective alone but augmented the effect of elastase. Sucrase and maltase were activated by proteolytic release, but activation was abolished by simultaneous exposure of brush borders to bile salts. Lactase was released and rapidly inactivated by proteinases, while alkaline phosphatase appeared to be inactivated without significant release. These results show that there are significant interactions between luminal factors which have been inapparent when studying them in isolation. Loss of functionally useful enzyme does not follow release of sucrase or maltase from the brush border into the lumen but does follow release of lactase. Study of the susceptibility of lactase to inactivation by luminal factors in the various forms of lactose intolerance is warranted.     

Modulation by thyroxine of the amount of lactase protein in the jejunum of adult rats

Adult rats starved for 48 h received a daily injection of thyroxine over a 3-day period before they were killed. When compared to nourished animals, starvation provoked a 4- to 5-fold increase in immunoreactive lactase protein, which paralleled a similar stimulation of lactase activity in the brush border membranes of the proximal jejunum. Exogenous thyroxine completely inhibited the starvation-induced increase in immunoreactive lactase protein in both the intracellular and the brush border membranes.     

Cortisone and thyroxine modulate intestinal lactase and sucrase mRNA levels and activities in the suckling rat.

Glucocorticoids and thyroxine modulate postnatal intestinal sucrase and lactase activities. Whether changes in enzyme activity are accompanied by changes in enzyme mRNA levels were determined in day 6 rats given thyroxine, cortisone, or thyroxine plus cortisone and killed 3 days later. Cortisone induced precocious expression of jejunal sucrase activity which was enhanced when cortisone plus thyroxine was administered; sucrase mRNA changed in parallel. Jejunal lactase activity was unaffected by thyroxine and was increased after cortisone, but not after thyroxine plus cortisone. Jejunal lactase mRNA levels increased equally after cortisone or after cortisone plus thyroxine. Thus, cortisone induces coordinated increases in sucrase and lactase activities and in corresponding mRNA levels. Thyroxine only enhances cortisone induced sucrase expression and antagonizes cortisone by depressing lactase activity post-translationally.     

Intestinal lactase (beta-galactosidase) and other glycosidase activities in suckling and adult tammar wallabies (Macropus eugenii)

The activities of various glycosidases in homogenates of the small intestinal mucosa of two adult and 18 suckling tammar wallabies (M. eugenii) aged from 6 to 50 weeks were investigated. Lactase (beta-D-galactosidase), beta-N-acetylglucosaminidase, alpha-L-fucosidase and neuraminidase activities were high during the first 34 weeks post partum and then declined to very low levels. Maltase, isomaltase, sucrase and trehalase activities were very low or absent during the first 34 weeks, and then increased. The lactase activity was unusual in being greater in the distal than the middle or proximal thirds of the intestine, and in its low pH optimum (pH 4.6), inhibition by p-chloromercuribenzene sulfonate but not by Tris, and lack of cellobiase activity. These properties are those of a lysosomal acid beta-galactosidase rather than of a brush border neutral lactase. The maltase activity had the characteristics of a lysosomal acid alpha-glucosidase early in lactation and of a brush border neutral maltase in adult animals. The significance of these findings is discussed in relation to changes in dietary carbohydrates during weaning and to the mode of digestion of milk carbohydrates by the pouch young.