Hypothesis that alpha-amylase evokes regulatory mechanisms originating in the pancreas, gut and circulation, which govern glucose/insulin homeostasis.

The anti-incretin theory involving the abolishment of diabetes type (DT) II by some of methods used in bariatric surgery, first appeared during the early years of the XXI century and considers the existence of anti-incretin substances. However, to date no exogenous or endogenous anti-incretins have been found. Our concept of the acini-islet-acinar axis assumes that insulin intra-pancreatically stimulates alpha-amylase synthesis ("halo phenomenon") and in turn, alpha-amylase reciprocally inhibits insulin production, thus making alpha-amylase a candidate for being an anti-incretin. Additionally, gut as well as plasma alpha-amylase, of pancreatic and other origins, inhibits the appearance of dietary glucose in the blood, lowering the glucose peak after iv or oral glucose loading. This effect of alpha-amylase can be interpreted as an insulin down regulatory mechanism, possibly limiting the depletion of pancreatic beta cells and preventing their failure. Clinical observations agree with the above statements, where patients with high blood alpha-amylase concentrations are seldom obese and seldom develop DT2. Obese-DT2, as well as DT1 patients, usually develop exo-crine pancreatic insufficiency (EPI) and vice versa. Ultimately, DT2 patients develop DT1, when the pancreatic beta cells are exhausted and insulin production ceases. Studies on biliopancreatic diversion (BPD) and on BPD with duodenal switch, a type of bariatric surgery, as well as studies on EPI pigs, allow us to observe and investigate the above-mentioned phenomena of intra-pancreatic interactions.

Difference in Performance of EPI Pigs Fed Either Lipase-Predigested or Creon®-Supplemented Semielemental Diet.

Pancreatic enzyme replacement therapy (PERT) and fat predigestion are key in ensuring the optimal growth of patients with cystic fibrosis. Our study attempted to highlight differences between fat predigestion and conventional PERT on body composition of young pigs with exocrine pancreatic insufficiency (EPI). EPI and healthy pigs were fed with high-fat diet for six weeks. During the last two weeks of the study, all pigs received additional nocturnal alimentation with Peptamen AF (PAF) and were divided into three groups: H-healthy pigs receiving PAF; P-EPI pigs receiving PAF+PERT; and L-EPI pigs receiving PAF predigested with an immobilized microbial lipase. Additional nocturnal alimentation increased the body weight gain of EPI pigs with better efficacy in P pigs. Humerus length and area in pigs in groups L and P were lower than that observed in pigs in group H (p value 0.005-0.088). However, bone mineral density and strength were significantly higher in P and L as compared to that of H pigs (p value 0.0026-0.0739). The gut structure was improved in P pigs. The levels of neurospecific proteins measured in the brain were mainly affected in P and less in L pigs as compared to H pigs. The beneficial effects of the nocturnal feeding with the semielemental diet in the prevention of EPI pigs' growth/development retardation are differently modified by PERT or fat predigestion in terms of growth, bone properties, neurospecific protein distribution, and gut structure.

Experiments suggesting extra-digestive effects of enteral pancreatic amylase and its peptides on glucose homeostasis in a pig model.

The studies presented were designed to highlight the impact of pancreatic enzymes on glycemic control and insulin response. Blood glucose and plasma insulin levels were monitored after intravenous, oral or direct gut glucose tolerance tests (GTT) in 6 pigs with an intact gastrointestinal tract and in 12 pigs following duodenal-jejunal bypass (DJB) surgery. In the intact pigs, pancreatic enzymes (Creon®) given orally 1 h prior to the GTT, lowered the blood glucose levels during the oral and meal GTT and reduced the plasma insulin response during the intravenous and meal GTT. In DJB pigs, blood glucose and plasma insulin levels were higher following glucose loading into the by-passed biliopancreatic limb as compared to that following glucose loading orally or into the common intestinal limb. Infusion of amylase or amylase peptides together with glucose into the biliopancreatic limb lowered blood glucose levels in DJB pigs. These preliminary data suggest new, extra-digestive, actions of enteral pancreatic enzymes - probably amylase or its peptides - on glucose homeostasis, with an reduction in net glucose absorption into the blood and in insulin response. This ability of digestive enzymes (amylase) to reduce post-prandial hyperglycaemia in an insulin-independent manner could aid in preventing the development of obesity and diabetes.

Dietary alpha-ketoglutarate increases cold tolerance in Drosophila melanogaster and enhances protein pool and antioxidant defense in sex-specific manner.

Alpha-ketoglutarate (AKG) is an important intermediate in Krebs cycle which bridges the metabolism of amino acids and carbohydrates. Its effects as a dietary supplement on cold tolerance were studied in Drosophila melanogaster Canton S. Two-day-old adult flies fed at larval and adult stages with AKG at moderate concentrations (5-10mM) recovered faster from chill coma (0°C for 15min or 3h) than control ones. The beneficial effect of AKG on chill coma recovery was not found at its higher concentrations, which suggests hormetic like action of this keto acid. Time of 50% observed mortality after 2h recovery from continuous cold exposure (-1°C for 3-31h) (LTi50) was higher for flies reared on 10mM AKG compared with control ones, showing that the diet with AKG enhanced insect cold tolerance. In parallel with enhancement of cold tolerance, dietary AKG improved fly locomotor activity. Metabolic effects of AKG differed partly in males and females. In males fed on AKG, there were no differences in total protein and free amino acid levels, but the total antioxidant capacity, catalase activity and low molecular mass thiol content were higher than in control animals. In females, dietary AKG promoted higher total antioxidant capacity and higher levels of proteins, total amino acids, proline and low molecular mass thiols. The levels of lipid peroxides were lower in both fly sexes reared on AKG as compared with control ones. We conclude that both enhancement of antioxidant system capacity and synthesis of amino acids can be important for AKG-promoted cold tolerance in D. melanogaster. The involvement of AKG in metabolic pathways of Drosophila males and females is discussed.

Dietary 2-oxoglutarate mitigates gastrectomy-evoked structural changes in cartilage of female rats.

Gastrectomy (Gx) leads to osteopenia/osteoporosis in humans and animals. However, little is known about the influence of Gx on the cartilage in this regard. Recent studies have demonstrated a protective effect of 2-oxoglutaric acid (2-Ox) on bone and cartilage. Hence, the purpose of this study was to investigate whether 2-Ox can mitigate eventual Gx-induced cartilage impairment. Twenty female Sprague-Dawley rats were subjected to Gx and randomly divided into two groups: Gx + 2-Ox and Gx. Another 20 rats were sham-operated (ShO) and randomly divided into two groups: ShO + 2-Ox and ShO. The daily dose of 2-Ox administered to the rats in the drinking water was 0.43 g per 100 g rat. After eight weeks, rats were euthanized and femora and tibiae were collected. Histology and histomorphometry analyses of the articular cartilage and the growth plate were done. Gx resulted in a 32% (±44.5 femur, ±35.8 tibia) decrease in overall thickness of articular cartilage in both bones (femur: ShO 279.1 ± 48.5 vs. Gx 190.2 ± 38.4 µm, tibia: ShO 222.9 ± 50.3 µm vs. Gx 151.3 ± 52.6 µm) (in some zones up to 58 ± 28.0%), and in the growth plate up to 20% (±22.4) (femur: ShO 243.0 ± 34.0 vs. Gx 207.0 ± 33.7 µm, tibia: ShO 220.0 ± 24.6 µm vs. Gx 171.1 ± 16.1 µm). Gx altered the spatial distribution of thick and thin collagen fibers, and chondrocyte shape and size. 2-Ox administration prevented the reduction in both cartilages thickness (Gx + 2-Ox: articular cartilage 265.2 ± 53.8 µm, 235.6 ± 42.7 µm, growth plate 236.7 ± 39.2 µm, 191.3 ± 16.5 µm in femur and tibia, respectively), and abolished the spatial changes in collagen distribution and structure induced by Gx. Gx affects cartilage structure and thickness, however, 2-Ox administration mitigates these effects and showed protective and stimulatory properties. Our observations suggest that dietary 2-Ox can be used to offset some of the changes in hyaline cartilage, in particular articular cartilage, following bariatric surgeries.

Melanoidins isolated from heated potato fiber (Potex) affect human colon cancer cells growth via modulation of cell cycle and proliferation regulatory proteins.

Melanoidins are brown, nitrogen containing, high molecular weight end products of Maillard reaction with poorly established activity towards tumor cells. The goal of present study was to verify whether both heated potato fiber Potex extract (180°C for 2h) and melanoidins isolated from the extract exerts growth-inhibiting activity in human colon cancer cells in vitro. The cells of LS180 colon cancer cell line were tested upon treatment with roasted potato fiber extract (AM4) as well as with high (HMW) and low (LMW) molecular weight fractions isolated from the extract, since both may be regarded as/or contain melanoidins. The tested compounds at concentration of 1000 µg/ml reduced cell growth down to 45%, 69% and 54%, respectively. Furthermore, deregulated ERK1/2 signaling was revealed upon treatment. Moreover, multiple alternations in cell cycle regulators activity were found (i.e. cyclinD1, cyclin-dependent kinase 4 and 6, p21, p27, p53, pRb) leading to cell cycle cessation in G0 phase. Importantly, LMW compounds revealed markedly stronger potential to alter specific molecular targets comparing to HMW compounds. Summarizing, the results emphasize that both high and low molecular weight melanoidins contribute to antiproliferative activity of heated potato fiber in LS180 colon cancer cells in vitro.

Alpha-ketoglutarate (AKG) inhibits proliferation of colon adenocarcinoma cells in normoxic conditions.

BACKGROUND AND OBJECTIVE: Alpha-ketoglutarate (AKG), a key intermediate in Krebs cycle, is an important biological compound involved in the formation of amino acids, nitrogen transport, and oxidation reactions. AKG is already commercially available as a dietary supplement and its supplementation with glutamine, arginine, or ornithine alpha-ketoglutarate has been recently considered to improve anticancer immune functions. It is well documented that AKG treatment of Hep3B hepatoma cells in hypoxia induced HIF-alpha (hypoxia-inducible factor) degradation and reduced vascular endothelial growth factor (VEGF) synthesis. Moreover, AKG showed potent antitumor effects in murine tumor xenograft model, inhibiting tumor growth, angiogenesis, and VEGF gene expression. However, the mechanisms of its anticancer activity in normoxia have not been examined so far. RESULTS: Here, we report that in normoxia, AKG inhibited proliferation of colon adenocarcinoma cell lines: Caco-2, HT-29, and LS-180, representing different stages of colon carcinogenesis. Furthermore, AKG influenced the cell cycle, enhancing the expression of the inhibitors of cyclin-dependent kinases p21 Waf1/Cip1 and p27 Kip1. Moreover, expression of cyclin D1, required in G1/S transmission, was decreased, which accompanied with the significant increase in cell number in G1 phase. AKG affected also one the key cell cycle regulator, Rb, and reduced its activation status. CONCLUSION: In this study for the first time, the antiproliferative activity of AKG on colon adenocarcinoma Caco-2, HT-29, and LS-180 cells in normoxic conditions was revealed. Taking into consideration an anticancer activity both in hypoxic and normoxic conditions, AKG may be considered as a new potent chemopreventive agent.

Potato fiber protects the small intestinal wall against the toxic influence of acrylamide.

OBJECTIVE: Acrylamide is a neurotoxic, genotoxic substance present in many commonly consumed food products and has been shown to have carcinogenic effects in rodents. The protective effects (if any) of potato fiber preparations, composed of cell wall material from potatoes, against the toxic influence of dietary acrylamide on the small intestinal wall were investigated. METHODS: Male mice of the BALB/c strain were used in the study. Acrylamide was administered to the mice in their drinking water (0.5 mg/kg of body weight per day) and one of two types of potato fiber preparations (heated or raw potato fiber preparation) was added to their feed (2% addition to their feed). Histomorphometry of the small intestinal wall, hemoglobin adducts of acrylamide, animal weight, and feed and water consumption analyses were performed. RESULTS: Acrylamide altered the morphology and histology of the small intestinal wall, decreasing proliferation, myenteron and submucosal thicknesses, villus length, fractal dimension, crypt depth, crypt number, and the small intestinal absorptive surface. Conversely, apoptosis, hemoglobin adduct levels, intensity of epithelium staining, enterocyte number, villus epithelial thickness, and crypt width and parameters associated with nerve ganglia were increased. The two potato fiber preparations that were used abolished the negative influences of acrylamide on the small intestinal wall and had no influence on the hemoglobin adduct levels of acrylamide. CONCLUSION: The negative impact of acrylamide on the histologic structure, regeneration, and innervation of the small intestinal wall and the absorptive function of the small intestinal mucosa can be abolished by dietary potato fiber preparations.

The effects of enteral ghrelin administration on the remodeling of the small intestinal mucosa in neonatal piglets.

Ghrelin is a multifunctional peptide produced predominantly in the stomach, however substantial amounts have also been found in colostrum and milk. The aim of the study was to investigate the effect of exogenous ghrelin, administered intra-gastrically, on the processes of mitosis, apoptosis, autophagy, crypt fission and changes in histometry of the small intestine mucosa in neonatal pigs, fed with a milk formula. Three groups (n=6) of piglets were used in the study. The pigs were fed either milk formula (C7) or milk formula together with ghrelin, administered via a stomach tube (7.5 µg/kg body weight (BW), (LG)) and 15 µg/kg BW (HG), every 8h for 6 days. Compared to the control group (C7), feeding milk formula supplemented with ghrelin resulted in significant changes in the small intestinal morphometry and mucosa histometry. The observed changes were dependent on the dosage of hormone and the part of intestine investigated. Administration of ghrelin via the stomach tube (HG) significantly influenced epithelial cell renewal. Moreover, we demonstrated that autophagy is involved in the small intestine mucosa remodeling and ghrelin may be an important factor for its regulation. In conclusion, we found that enteral ghrelin influences the gut mucosa remodeling in a dose-related manner in the early postnatal period. Moreover in neonates, stomach activity does not interfere with the action of ghrelin in the small intestine.

Antiproliferative activity of melanoidins isolated from heated potato fiber (potex) in glioma cell culture model.

Potex constitutes a potato fiber preparation widely used as an ingredient to meat and bakery products which thermal treatment results in creation of new compounds. Melanoidins are high molecular weight brown end products of Maillard reaction, and few data presenting tumor cell growth inhibiting activity of melanoidins have been reported. Thus, in present study we utilized water extract of Potex roasted (180 °C for 2 h), whose chemical characterization revealed the presence of melanoidin complexes. Heated Potex extract inhibited C6 glioma cell proliferation in a dose-dependent manner measured by MTT method. High molecular weight components present in initial extract were responsible for stronger antiproliferative effect compared with low molecular weight fraction. Impaired MAPK (mitogen-activated protein kinase) and Akt signaling was found in cells treated with the extract. Moreover, flow cytometry analyses revealed the extract to induce G1/S arrest in glioma cells. Simultaneously, Western blot analysis showed elevated levels of p21 protein with concomitant decrease of cyclin D1. In conclusion, observed antiproliferative activity of melanoidins present in heated Potex was linked to disregulated MAPK and Akt signaling pathways, as well as to cell cycle cessation. These results suggest potential application of Potex preparation as a functional food ingredient and chemopreventive agent.

Immune suppression by cyclosporin A inhibits phytohemagglutinin-induced precocious gut maturation in suckling rats.

OBJECTIVES: Enteral exposure to the lectin phytohemagglutinin (PHA) provokes precocious gut maturation in suckling rats coinciding with an early expansion of intestinal mucosal T and B lymphocytes. Here, the role of the immune system in neonatal gut growth and maturation was further studied. MATERIALS AND METHODS: The effects of immunosuppression by cyclosporine A (CyA), 7.5 microg/g of body weight, injected 12 hours before and then daily after the intragastric gavage of PHA, 100 microg/g body weight, to 14-day-old suckling rats were studied after 4 and 12 hours and later after 72 hours. RESULTS: At 4 hours after PHA feeding, an early rapid increase in the intestinal levels of the proinflammatory cytokines interleukin-6, interleukin-1beta, and tumor necrosis factor was obtained, and the CyA treatment did not prevent the temporary PHA-induced intestinal disturbance seen at 12 hours. Later, at 72 hours after PHA gavage the CyA treatment significantly counteracted the PHA-induced gut changes with a decrease in small intestinal growth, a delay in the appearance of adult-phenotype enterocytes in the distal small intestinal, and total inhibition of the PHA-induced pancreas development. Additionally, the increase in plasma level of the acute phase protein, haptoglobin, after PHA feeding was dampened by CyA. CONCLUSIONS: The results indicate that proinflammatory cytokines are involved in the early recruitment of lymphocytes to the gut after PHA challenge, and that the ensuing precocious gut maturation is dependent on activation of the immune system, presumably T cells, in suckling rats.

Alpha-ketoglutarate decreases serum levels of C-terminal cross-linking telopeptide of type I collagen (CTX) in postmenopausal women with osteopenia: six-month study.

Several studies have shown that alpha-ketoglutaric acid (AKG) increases serum levels of proline and has beneficial effects on skeletal development. We studied the effect of alpha-ketoglutaric (AKG) acid calcium salt (6 g AKG and 1.68 Ca/day) or calcium alone (1.68 Ca/day) on serum C-terminal cross-linked telopeptide of type I collagen (CTX) and osteocalcin (OC), as well as on lumbar spine bone mineral density (BMD) in a randomized, parallel group, double-blind, 6-month study conducted on 76 postmenopausal women with osteopenia. The maximum decrease of the mean CTX level in the AKG-Ca group was observed after 24 weeks (37.0%, p = 0.006). The differences in CTX between study groups were statistically significant after 12 and 24 weeks. The OC serum level was not affected by treatments. The BMD of the AKG-Ca group increased by 1.6% from baseline; however, the difference between treatment groups was estimated as 0.9% (non-significant). This study suggests the potential usefulness of AKG-Ca in osteopenic postmenopausal women. AKG-Ca induced beneficial changes in serum CTX, which was consistent with preserving the bone mass in the lumbar spine; however, the long-term effect needs to be further investigated.

Binding and the effect of the red kidney bean lectin, phytohaemagglutinin, in the gastrointestinal tract of suckling rats.

Enteral exposure of suckling rats to phytohaemagglutinin (PHA) has been shown to induce growth and precocious functional maturation of the gastrointestinal tract. The aim of the present study was to explore the mechanism of this action. Suckling rats, 14 d old, were fed a single dose of PHA (0.05 mg/g body weight) or saline. The binding of PHA to the gut epithelium and its effect on the morphology and functional properties of the gut and pancreas were studied up to 3 d after treatment. Initially, at 1-24 h, the PHA bound along the gut mucosal lining, resulting in disturbed gut morphology with villi shortening and rapid decreases in disaccharidase activities and macromolecular absorption capacity. During a later phase, between 1 and 3 d, the PHA binding had declined, and an uptake by enterocytes was observed. An increase in crypt cell proliferation and gut growth became evident during this period, together with a functional maturation, as indicated by increases in disaccharidase (maltase and sucrase) activities and the low macromolecular absorption capacity. Pancreas growth also increased, as did its content of digestive enzymes. We conclude that enteral exposure to PHA in suckling rats temporarily causes mucosal disarrangement and functional impediment of the gut, which may be explained by binding to and disruption of the gut mucosa and a two-fold increase in the plasma corticosterone concentration. These findings may lead to a better understanding of the role of diet in gastrointestinal maturation and may constitute a basis for the treatment of mammals having an immature gut.