Fifty-three years ago, Holzel discovered that the chronic diarrhea of 2 malnourished infants was caused by lactase deficiency (1). The next year, Weijers reported that an identical syndrome could be caused by sucrase and/or maltase deficiencies (2). These conclusions were based on clinical response to dietary elimination and oral sugar loads, resulting in inadequate blood glucose responses (3). Congenital sucrase-isomaltase deficiency (CSID) was at first characterized clinically as a subset of chronic diarrhea. There was onset of watery diarrhea, with low fecal pH and high excretion of lactic acid and low-molecular-weight fatty acids, when ingested food contained the sugar. Soon after the publication of these observations, Prader and Auricchio (Fig. 1) described 5 children between 10 months and 4 years old belonging to 3 different families who were affected by the same disease; the sucrose intolerance was the result of a deficiency of small intestinal sucrase activity (4,5). Diagnosis was based on dependency of the diarrhea on sucrose in the diet and the high fecal excretion of lactic acid when the diet contained sucrose, and on the results of an oral sucrose tolerance test (flat blood glucose response and production of acute diarrhea with acid stools that were enriched with sucrose and lactic acid). Glucose plus fructose tolerance tests, however, were normal (increase of blood glucose and no diarrhea). These patients with CSID also presented symptoms of intolerance to feeding of maltodextrins or starch, but the symptoms were of a milder degree than those after sucrose feeding and with intolerance mainly in the first year of life. The α-amylase activity in duodenal juice was found to be normal for age and tolerance for maltose feeding was normal. It was therefore supposed that starch and maltodextrin maldigestion resulted from a deficiency of isomaltase activity associated with the deficiency of sucrase activity. At that time it was not possible to measure disaccharidase activities in small intestinal biopsies. Isomaltose was unavailable in sufficient amounts for oral tolerance tests. Auricchio therefore tested the in vivo isomaltase activity by oral tolerance tests with disaccharides or oligosaccharides containing the same 1–6 α-glycosidic bond as isomaltose. The first experiments were done with peroral tolerance tests using palatinose (6-α-D-glycopyranoside-D-fructose), a bacterial transformation product of sucrose (5) and afterward, with a mixture of 1–6 α-oligosaccharides obtained by enzymatic hydrolysis of dextran, which contained enriched isomaltose and isomaltotriose (6). In contrast to normal subjects, the patients with CSID had little or no elevation of blood glucose after the test dose and promptly developed diarrhea with large amounts of palatinose or isomaltose in the stools, which had the characteristic high content of lactic acid and low pH. Results indicated that in these patients, sucrase and isomaltase activities were missing simultaneously: “the isomaltase deficiency seems to explain fully the symptoms these patients get on the ingestion of starch or dextrins since isomaltose represents the bond existing in the branch point of starch” (6).
BIOCHEMISTRY OF CSID ENZYME DEFICIENCY
The discovery of this form of diarrhea caused by a congenital deficiency of sucrase and isomaltase enzyme activities generated great interest in the laboratories of Dahlqvist in Sweden and Semenza in Switzerland; they developed biochemical studies on the disaccharidases of the human intestine. Several intestinal maltases (also called α-glucosidases) were identified by heat inactivation studies and confirmed by gel filtration chromatography on Sephadex (7–9). These investigators identified 2 heat-resistant maltases that split only maltosides (maltase-glucoamylase complex [MGAM]), a third maltase that was more heat sensitive and hydrolyzed sucrose, and a fourth maltase that also split isomaltose (sucrase-isomaltase complex, SI). The documented deficiency of the activities of 2 different enzymes, sucrase and isomaltase, in this “inborn error of metabolism” was surprising and a type of close relation between the 2 enzymes was advocated (10,11). The cloning of the SI gene in the laboratory of Semenza demonstrated that sucrase and isomaltase activites resulted from 2 different active centers of the same peptide (12).
Meanwhile, micromethods were developed to measure the disaccharidase activities in small intestinal biopsies. The “normal” values for this enzymatic activity were established (13). Maltase activity was found to be 2 to 4 times higher than that of sucrase and isomaltase. Disaccharidase activities were then measured in biopsies of the small intestines of 15 patients with CSID (14–16). All showed a complete absence of sucrase activity and a severe deficiency of isomaltase activity. The maltase activity was markedly reduced and primarily the result of the heat-resistant maltases. These results were interpreted as a proof of deficiency of maltase-sucrase and maltase-isomaltase. In most patients the residual maltase activity caused by the heat-resistant maltases was sufficient to hydrolyze normal amounts of maltose. It is interesting to note that it was during these studies of the “normal” values of disaccharidase activities in surgical and peroral intestinal biopsies that adults with isolated intestinal lactase deficiency were first identified. They were found to have lactose intolerance in adult life, but normal lactose tolerance as infants and were, therefore, first identified as affected by adult-type hypolactasia (16).
NUTRITION OF PATIENTS WITH CSID
The hydrolysis of starch was studied in more detail in CSID (17). From the first clinical studies of the disease it was recognized that after ingestion of starch, patients with CSID developed a milder degree of fermentative diarrhea than after ingestion of sucrose. In this study, the intraluminal α-amylolytic digestion of starch was studied by analyzing the glucose oligosaccharides that accumulated in the proximal jejunum after a test meal containing amylopectin. The levels of α-amylase activity in the intestinal juice of older patients with CSID were confirmed to be normal. Furthermore, the intestinal content of α-limit dextrins (α-LDx) of patients with CSID and a control were not different; the average degree of polymerization (DP) of total glucose oligosaccharides was low (2.15–2.53), and dextrins with higher molecular weights (DP >7) contributed only approximately 10% of total carbohydrate. Almost 90% of oligosaccharides had DP lower than 7; these were the branched dextrins (<27%; DP 5.11–5.85), as well as maltotriose, maltose, and glucose. No maltotetrose was found in this age group. In contrast, in patients with CSID and controls from 6 to 13 months of age, the level of α-amylase activity in the intestinal juice was low and there were no differences in the α-amylolysis of amylopectin detected. In these infant subjects, the α-amylolysis of amylopectin in the proximal part of the jejunum was less extensive than in the older children. Dextrins with DP >7 were present in larger amounts (>27%), and the content of small branched dextrins was lower (<16%). Lumenal maltotetrose was increased in young patients with CSID and acted as a “brake” on residual MGAM maltase activities. Furthermore, Auricchio reported, “The maltotriose oral tolerance test was normal in a 31-month-old patient, whereas it caused fermentative diarrhea and only a small increase of blood glucose in a 6-month-old patient” (17).
CLINICAL RECOGNITION OF PROLONGED CARBOHYDRATE INTOLERANCE
The broader clinical focus of this workshop was well described by Howland in 1921, as prolonged intolerance to carbohydrates (18). He wrote, “A more common condition and a more important one is the prolonged intolerance that develops after severe diarrhea, especially after numerous attacks of diarrhea.” A century ago, chronic diarrhea was the blight of all infant wards. Fifty years ago, one of the present authors (B.L.N.) found this problem as an intern when he was assigned the care of Stevie S. on the Grace-New Haven Hospital infant ward. Daily intravenous needlework was required to maintain his hydration while formulas were blindly switched; all of them exacerbated the infant's watery diarrhea. Nichols's attending, Nelson Ordway, had trained in Germany and was aware of Finkelstein's “protein milk” formula, which fed a constant protein and fat component but allowed modular change of carbohydrate. Through a series of systematic trials it was determined that Stevie tolerated a starch hydrolysate called maltodextrin. The substitution of this carbohydrate in the formula resulted in resolution of his chronic diarrhea, nutritional recovery, and discharge.
When Nichols arrived to supervise the pediatric house staff at Baylor College of Medicine in 1964, many infants with malnutrition and chronic diarrhea were discovered at the affiliated Texas Children's and Ben Taub Hospitals. Their number represented almost 3% of all infant hospitalizations (19). Nichols applied the Finkelstein method of management with some success, but there were long hospital stays. A modular formula similar to Finkelstein's protein milk was produced by Ross Laboratories (ProViMin; Seattle, WA) and used with progression of tolerated carbohydrate concentrations up to 7% (20). In 1967, a study of these infants was begun by mucosal biopsy with the modified Crosby capsule technique, and almost universal subtotal villous atrophy was discovered with increased intraepithelial lymphocytes (21,22). This pathology led the house staff to call the condition the slick-gut syndrome, but the faculty preferred to call it acquired monosaccharide intolerance (AMI). The diagnostic hallmark was clinical intolerance of >3% glucose in feeds. The use of constant nasogastric drips for nutritional support improved tolerance for higher concentrations of glucose (23), but patient recovery and hospital discharge remained slow. In 1968, Wilmore and Dudrick presented their study of total parenteral nutrition (TPN) in an infant with catastrophic loss of the small intestine (24). Implementing this intravenous mode of nutritional support greatly reduced mortality and morbidity in the patients with AMI, and ProViMin was used only for diagnostic purposes and weaning infants from TPN to oral feedings. The implementation of lifesaving TPN allowed investigations of the mucosal pathology in these fragile malnourished patients. Disaccharidase assays of mucosal biopsies revealed reduced lactase, sucrase, and maltase activities, but the clinical intolerance characterized by onset of watery diarrhea, with low fecal pH and presence of glucose, after feeding of these dietary substrates was not predictable (25). In contrast, reduced glucose absorption correlated with the degree of villous atrophy (26), and when the free glucose was replaced with equal energy from maltodextrins, water flux changed from secretion to absorption. Using an oral 13C-glucose substrate in a breath test showed that the reduction of glucose assimilation and oxidation in these patients was equivalent to the 13C-glucose malabsorption in congenital glucose-galactose malabsorption (27).The slick-gut syndrome disappeared when home-prepared infant formulas made with added corn syrup were abandoned. In retrospect, it was realized that light corn syrup is rich in maltotriose and maltotetriose and may act as “brakes” on MGAM maltase activities (28). The historical, clinical, and histologic aspects of AMI were summarized by Nichols et al in 1990 (19).
MOLECULAR BIOLOGY OF MUCOSAL CARBOHYDRATE DIGESTING ENZYMES
Lactase-phlorizin hydrolase (29) and SI (30) had already been sequenced and reported when Nichols took a 1992–1993 sabbatical in the Sterchi laboratory in Bern, Switzerland, to work toward cloning and sequencing of MGAM. This work was continued at the US Department of Agriculture/Agricultural Research Service Children's Nutrition Research Center at Texas Children's Hospital and Baylor College of Medicine. The deduced peptide sequence of MGAM was reported in 1998 (31) and the gene sequence was reported in 2003 (32). The peptide had close homology to SI, each having 2 catalytic domains, thus confirming the earlier studies of Semenza et al (8) and Dahlqvist et al (9). To understand the overlapping activities of the 4 maltase domains, a series of collaborative studies was carried out by immunoprecipitation of purified MGAM and SI complexes from normal human mucosal homogenates (33,34). The authors confirmed that SI accounts for approximately 85% of mucosal maltase activity, but MGAM is 10-fold more active (7–9). The role of recombinant α-amylase was tested in this in vitro system and shown to amplify the maltase activities of SI > MGAM (33). The authors found, however, that MGAM activities were inhibited by the α-LDx resulting from α-amylase hydrolysis of starch. This α-LDx inhibition of MGAM activity was termed a “brake,” which did not constrain SI activity (33,34). The relative in vivo roles of SI and MGAM complexes was tested in a MGAM knockout (KO) mouse model. Mgam (rodent terminology) ablation reduced total 13C-starch digestion to 13C-glucose and oxidation to breath 13CO2 by 40% (35), but did not alter prandial blood glucose response or total energy expenditure (35,36). Early α-glucogenesis from starch was equivalent in the wild-type and null mice on ad libitum intakes but higher in the wild type when starch intake was limited (Fig. 2). The measurement of the rate of gluconeogenesis in the MGAM KO mouse revealed that hepatic glucogenesis fully compensated for reduced prandial glucogenesis from starch digestion in the Mgam nulls (B.L.N., unpublished data, 2012). The hypothesis proposed that the high-efficiency MGAM complex is essential for digestion on low starch intakes, such as snacking, but is sensitive to the “brake” effect of α-LDx, and that “brake- resistant” SI has a dominant effect on the rate of glucogenesis from starch during feasting (Fig. 2).
NUTRITIONAL RECOGNITION OF MUCOSAL ENZYME ACTIVITIES
Because of the 2 catalytic sites on each of the SI and MGAM enzyme complexes, the individual maltase domains were expressed as recombinant peptides. The catalytic characteristics of the individual domains were investigated independently. The “brake” effect is limited to the MGAM glucosidase domain activity. The concept developed that the consortium of 4 mucosal maltases provides a range of substrate specificities needed for glucogenesis from the more than 400 different food starches consumed by humans. This hypothesis was termed “toggling” of the mucosal activities and was tested with a variety of glucosidase inhibitors. The hypothesis was supported by the differential modulation of the 4 different maltase activities (37,38). Many of the botanical sources of food starches also contain glucosidase inhibitors, which require further investigation using recombinant activities. In addition, the purified starches from different foods have structural features that modulate the rate and efficiency of glucogenesis (B.L.N., unpublished data, 2012). The individual recombinant α-glucosidase domains have been crystallized and studied at the structural level (37–40).
NONINVASIVE DIAGNOSIS OF CSID
The original diagnoses of CSID relied on clinical responses to sucrose elimination diets and were reinforced by flat blood glucose responses to sucrose challenges. An approximate 25% false-positive response resulted from the oral sucrose challenge (41). The reduced biopsy sucrase enzyme assays have become the standard for diagnosis of CSID; the biopsy technique, however, is invasive and the mucosal sample for enzyme assay requires rapid freezing and frozen transfer to the analytical laboratory. Confusion about the cutoff for normal sucrase activity persists. The H2 breath test for proximal glucose malabsorption was developed for lactose malabsorption and rapidly modified for detection of sucrose malabsorption (43–46). Not all patients with chronic diarrhea produce breath hydrogen (42,44), and the administered load of sucrose produces clinical symptoms. These considerations led to the development of 13C-substrate breath tests (46). Although a large dose of naturally enriched maize sugar had been used by others, Robayo-Torres et al (46) used a small dose of universally enriched 13C-substrates, believing that detection of deficiencies would be more precise if the normal baseline were highly enriched and the enzymes and transporters were not overloaded. The authors recognized that children and adults have different rates of glucose oxidation; they then normalized the test substrate oxidation to that of 13C-glucose. The digestion and oxidation of UL-13C-sucrose/glucose in CSID agreed with duodenal biopsy sucrase activity and was fully corrected by oral yeast sucrase supplements (46). Several clinical reviews have reported that some patients with CSID may also have starch maldigestion (2–4). A UL-13C-starch/glucose breath test demonstrates that starch digestion is poor in all patients with CSID. The correlation of starch breath test values with duodenal maltase activity is reduced because of the multiple enzyme activities that make up this mucosal activity (Fig. 3, Table 1). The universal poor starch digestion in CSID, however, confirms the biopsy sucrose- and maltase-enzyme assays (46) and Auricchio and colleagues’ pioneering CSID starch digestion studies (17).
ARE ALL STARCHES DIGESTED EQUALLY?
Given that starch digestion is poor in patients with CSID, are all food starches equally maldigested? Dietary management of type 2 diabetes mellitus recognizes that some starches are more slowly digested than others, resulting in variations of glucogenesis from starch, frequently recognized as the blood glycemic index. Parents of patients with poor starch digestion have suggested that slowly digested starches are better tolerated. Such reports deserve experimental confirmation.
Direct evidence from mouse experiments confirms Auricchio and colleagues’ early report (17) that SI enzyme activity plays a dominating role in normal mucosal starch digestion and that the α-amylase product of starch hydrolysis, α-LDx, containing abundant maltotriose and maltotetrose, can inhibit the residual maltase activity of MGAM in CSID. When feasting, MGAM KO mice, with only SI activity, have rates of starch digestion to glucose that are identical to those of wild-type mice with both activities; when snacking, however, the more efficient MGAM activities dominate (Fig. 2); thus, the dominant role of SI on α-LDx digestion to glucose determines the rate of early glucose absorption from starch during feasting. These insights support the general hypothesis that all patients with CSID with low sucrase activities will have poor digestion of starch. The hypothesis is supported by the poor 13C-starch digestion in patients with CSID with documented 13C-sucrose maldigestion (Fig. 3).
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