Exocrine Pancreatic Insufficient Pigs Research Articles

Anti-Incretin Gut Features Induced by Feed Supplementation with Alpha-Amylase: Studies on EPI Pigs.

The acini-islet-acinar (AIA) axis concept justifies the anatomical placement of the Langerhans islets within the exocrine pancreatic parenchyma and explains the existence of the pancreas as a single organ. Amylase has been suggested to play a key role as an anti-incretin factor. Oral glucose tolerance tests (OGTT) were performed on 18 piglets in both a healthy (prior to pancreatic duct ligation (PDL) surgery, study Day 10) and an exocrine pancreatic insufficient (EPI) state (30 days after PDL, study Day 48)). Amylase (4000 units/feeding) or Creon® (100,000 units/feeding) was administered to pigs with the morning and evening meals, according to study design randomization, for 37 days following the first OGTT. Blood glucose levels, as well as plasma levels of insulin, GLP-1, and GIP, were measured, and the HOMA-IR index was calculated. EPI status did not affect the area under the curve (AUC) of insulin release, fasting insulin levels, or the HOMA-IR index, while amylase supplementation led to a significant (p < 0.05) decrease in the above-mentioned parameters. At the same time, EPI led to a significant (p < 0.05) increase in GLP-1 levels, and neither amylase nor Creon® supplementation had any effects on this EPI-related increase. Fasting plasma levels of GIP were not affected by EPI; however, the GIP response in EPI and Amylase-treated EPI animals was significantly lower (p < 0.05) when compared to that of the intact, healthy pigs. Orally administered amylase induces gut anti-incretin action, normalizing glucose homeostasis and reducing HOMA-IR as a long-term outcome, thus lowering the risk of diabetes type II development. Amylase has long-lasting anti-incretin effects, and one could consider the existence of a long-lasting gut memory for amylase, which decreases hyperinsulinemia and hyperglycemia for up to 16 h after the last exposure of the gut to amylase.

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.

Validation of an omega-3 substrate challenge absorption test as an indicator of global fat lipolysis.

The coefficient of fat absorption (CFA) quantifies fat that remains in stool after digestion and is not a direct measure of lipolysis. CFA has been used to assess treatment of pancreatic insufficiency but does not correlate with pancreatic enzyme replacement therapy dose. We explored use of an omega-3 substrate absorption challenge test as a sensitive test of lipolysis and absorption. We studied a novel microbially-derived lipase (SNSP003) employing an established surgical model commonly used to study the uptake of macronutrients, the exocrine pancreatic insufficient pig. Pigs were fed a high-fat diet and given a standardized omega-3 substrate challenge to test the effect of lipolysis on its absorption. Blood was drawn at 0, 1, 2, 4, 6, 8, 12, and 24 hours following the substrate challenge and was analyzed for omega-3 and total fat levels (c14:c24). SNSP003 was also compard to porcine pancrelipase. The absorption of omega-3 fats was significantly increased following administration of 40, 80 and 120 mg SNSP003 lipase by 51% (p = 0.02), 89%, (p = 0.001) and 64% (p = 0.01), respectively, compared to that observed when no lipase was administered to the pigs, with Tmax at 4 hours. The two highest SNSP003 doses were compared to porcine pancrelipase and no significant differences were observed. Both doses increased plasma total fatty acids (141% for the 80 mg dose (p = 0.001) and 133% for the 120 mg dose (p = 0.006), compared to no lipase) and no significant differences were observed between the SNSP003 lipase doses and porcine pancrelipase. The omega-3 substrate absorption challenge test differentiates among different doses of a novel microbially-derived lipase and correlates with global fat lipolysis and absorption in exocrine pancreatic insufficient pigs. No significant differences were observed between the two highest novel lipase doses and porcine pancrelipase. Studies in humans should be designed to support the evidence presented here that suggests the omega-3 substrate absorption challenge test has advantages over the coefficient of fat absorption test to study lipase activity.

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.

Pancreatic-like enzymes of microbial origin restore growth and normalize lipid absorption in a pig model with exocrine pancreatic insufficiency.

IntroductionThe standard therapy for exocrine pancreatic insufficiency (EPI) is porcine-derived pancreatic enzyme replacement therapy (PERT). In the present study we tested a new approach with a mixture of pancreatic-like enzymes of microbial origin (PLEM) in a 1-week efficacy study in EPI pigs. In addition to the conventionally used coefficient of fat and nitrogen absorption (CFA and CNA), parameters that more accurately reflect the nutritional and health status, such as changes in the lipemic index (LI), plasma triglyceride (TG) and non-esterified fatty acid (NEFA) levels, and somatic growth, were determined.Material and methodsA PLEM dose containing 120 000 active lipase units, 80 000 active protease units and 12 000 active amylase units (all from Sigma, St. Louis, MO) was given as a powder, twice daily with a meal (40 g fat/meal) to 8 EPI pigs for 7 days. Ten healthy pigs were used as a comparator.ResultsThe PLEM enhanced fat and protein digestion, and reversed growth impairment in EPI pigs. With treatment, CFA and CNA increased by 59% and 43% (p < 0.05), respectively. Although fat and protein absorption were lower than in the comparator, the postprandial blood lipid profile was normal as in healthy pigs. The mucosal thickness significantly increased by 27%, 50% and 26%, in the proximal, middle, and distal jejunum (p < 0.05) with treatment and resembled that of healthy animals.ConclusionsPancreatic-like enzymes of microbial origin supported somatic growth and normalized the postprandial lipid profile. As a measure of efficacy, postprandial LI, TG and NEFA are viable endpoints to be explored in human trials.

Enhanced absorption of long-chain polyunsaturated fatty acids following consumption of functional milk formula, pre-digested with immobilized lipase ex vivo, in an exocrine pancreatic insufficient (EPI) pig model

An exocrine pancreatic insufficient (EPI) pig model was used in the study. The effects of milk formula pre-digestion with immobilized microbial lipase, on the absorption and tissue accretion of long chain polyunsaturated fatty acids (LCPUFA) were assessed. Thirteen male EPI pigs, 10weeks of age, were used in the study. Six healthy pigs, 6weeks of age, were used as controls. The pigs were fed either a regular or a pre-digested milk formula. The formula pre-digestion resulted in the decreased faecal total fat and LCPUFA excretion (by 43% and 38% respectively), increased plasma and tissue LCPUFA content (up to 38%). In conclusion, we postulate that feeding a pre-digested milk formula, is an efficient method to develop a functional milk formula, the consumption of which will ensure an increase in total fat absorption (in particular LCPUFA) in human infants as was indicated in the present study in the EPI pig model.

EPS05.1 Nutritional formula pre-hydrolyzed by novel point of care lipase device (EFIC™) leads to increased fat absorption in young pigs with exocrine pancreatic insufficiency (EPI)

Background Patients with CF and EPI have difficulty managing weight requiring liquid supplements and tube feeds. Administration of PERTs with enteral feeding is challenging, as enzyme availability is mismatched with formula, negatively affecting fat digestion, absorption and leading to fatty acid deficiencies. EFIC contains an immobilized lipase within a cartridge that is a part of enteral feeding circuit. Formula passes through the EFIC pre-hydrolyzing triglycerides including polyunsaturated fatty acids (PUFA) such as docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA) into absorbable fatty acids. This study measured fat absorption and changes in plasma profiles of DHA and EPA after feeding EPI pigs, with formula +/– EFIC. Methods Fasted EPI pigs were tube fed for 5 h (2 mL/min) with 500 mL Peptamen AF (Nestle, 32 g fat, 1.8 g DHA/EPA), either as EFIC pre-hydrolyzed (EPI+EFIC n = 6) or non-hydrolyzed (EPI n = 5) formula. 3 healthy age matched pigs served as controls. Ten plasma samples were collected over 24 h. Lipids were measured by lipaemic index (LI, turbidometry) and fatty acids by GC-MS and UPLC-MS. Results Fat absorption improved with EFIC compared to the non-hydrolyzed group with 5 h Cmax for LI and AUC 0–10 11.5±1.99 vs. 9.1±1.63 (p Conclusions Use of EFIC leads to an increase in total fat absorption and specifically, plasma levels of physiologically relevant PUFAs, such as DHA and EPA. EFIC provides an effective approach to optimizing fat hydrolysis, PUFA absorption and caloric intake during enteral feeding.

Diet supplemented with pancreatic-like enzymes of microbial origin restores the hippocampal neuronal plasticity and behaviour in young pigs with experimental exocrine pancreatic insufficiency

It is postulated that exocrine pancreatic insufficiency (EPI) can evoke neurological disorders. In the present study pancreatic-like enzymes of microbial origin (PLEM) were examined as a functional food component, with the goal of improving cognitive function and brain structure in a pig model of EPI. EPI in the present study induced alterations in the behaviour of the pigs as well as degenerative changes within the morphological structure of the hippocampus. EPI leads to a reduced number of pyramidal neurons and decreased levels of neural cell adhesion molecules (NCAM) in the CA1 area of the hippocampus. Here, we provide evidence that the use of PLEM as a functional food, in the form of dietary supplementation with PLEM for 10 days, restored pig behaviour and the histo-morphology of the hippocampus in EPI pigs. Thus, we suggest that the use of PLEM as a functional food ingredient should be considered in the prevention and/or postponement of the development of EPI-related encephalopathy.

A piglet with surgically induced exocrine pancreatic insufficiency as an animal model of newborns to study fat digestion.

The maldigestion and malabsorption of fat in infants fed milk formula results due to the minimal production of pancreatic lipase. Thus, to investigate lipid digestion and absorption and mimic the situation in newborns, a young porcine exocrine pancreatic insufficient (EPI) model was adapted and validated in the present study. A total of thirteen EPI pigs, aged 8 weeks old, were randomised into three groups and fed either a milk-based formula or a milk-based formula supplemented with either bacterial or fungal lipase. Digestion and absorption of fat was directly correlated with the addition of lipases as demonstrated by a 30% increase in the coefficient of fat absorption. In comparison to the control group, a 40 and 25% reduction in total fat content and 26 and 45% reduction in n-3 and n-6 fatty acid (FA) content in the stool was observed for lipases 1 and 2, respectively. Improved fat absorption was reflected in the blood levels of lipid parameters. During the experiment, only a very slight gain in body weight was observed in EPI piglets, which can be explained by the absence of pancreatic protease and amylase in the gastrointestinal tract. This is similar to newborn babies that have reduced physiological function of exocrine pancreas. In conclusion, we postulate that the EPI pig model fed with infant formula mimics the growth and lipid digestion and absorption in human neonates and can be used to elucidate further importance of fat and FA in the development and growth of newborns, as well as for testing novel formula compositions.

Effect of osteocalcin on human beta cell proliferation and function and its role on insulin secretion

Effect of osteocalcin on human beta cell proliferation and function and its role on insulin secretion

Exogenous pancreatic-like enzymes are recovered in the gut and improve growth of exocrine pancreatic insufficient pigs1

The exocrine pancreatic insufficient (EPI) pigs grow less due to different disturbances in feed digestion, absorption, and retention. Use of pancreatic-like enzymes of microbial origin in pigs may improve feed use and performance in slow-growing pigs. The aim was to study gut recovery and effectiveness of pancreatic-like enzymes of microbial origin supplementation on pig performance. Six male pigs 10 to 12 kg BW underwent pancreatic duct ligation surgery to induce total exocrine pancreatic insufficiency (EPI). Three cannulas to access the gastrointestinal tract content were installed in stomach, duodenum, and ileum in EPI pigs and in 3 control (healthy) pigs. One month after surgery, enzymes were given before feeding and digesta samples were collected for analyses. The BW of EPI pigs did not increase during 1 mo following surgery (11.7 vs. 11.6 kg BW); however, BW increased after 1 wk of enzyme supplementation (12.1 kg BW). Coefficient of fat and N absorption increased (P < 0.05) in EPI pigs after enzyme supplementation. Activity of amylase, lipase, and protease in chyme samples of EPI pigs was very low compared to controls. In EPI pigs after enzyme supplementation, amylase activity increased from 5.32 to 72.9 units/mL but remained lower than that of healthy pigs (162.7 units/mL). Lipase activity increased from 79.1 to 421.6 units/mL, which was similar to that of controls (507.3 units/mL). Proteolytic activity increased from 7.8 to 69.7 units/mL but still did not reach control pigs (164.3 units/mL). In conclusion, exogenous microbial enzymes mimic endogenous pancreatic enzymes being recovered along the lumen of the gastrointestinal tract. These enzymes might be a useful tool to stimulate growth of slower-growing pigs after the weaning period.

Behavioral changes in response to feeding pancreatic-like enzymes to exocrine pancreatic insufficiency pigs1

Behavioral changes during pancreatic enzyme therapy have never been studied. The present study investigated behavioral changes in exocrine pancreatic insufficiency (EPI) pigs when their feed was supplemented with pancreatic-like enzymes of microbial origin. A crossover design study was used to test the effect of enzyme supplementation in 2 × 4 EPI pigs that underwent pancreatic duct ligation (PDL). After 40 d of adaptation, the study commenced, comprising 2 control and 2 enzyme feeding periods of 10 d each in sequence. On days 7 and 10 of each experimental period, behavior was monitored for 24 h and feed consumption and BW were recorded. Behavioral observations focused on the pigs' activity-- lying down or passive, or sitting, or standing or active--and were expressed as percentage activity for 24 h. During the adaptation period, BW gain was completely inhibited after PDL whereas for the entire study period, the body weight increased from 10.5 ± 1.1 to 14.0 ± 1.4 kg (P < 0.01). Exocrine pancreatic insufficiency pigs were more active when fed the enzymes (21 vs. 18% per 24 h; P < 0.01). Microbial enzyme supplementation not only improved the growth of the EPI pigs but it also increased their activity. This behavior change contradicts the generally accepted norm that satiety evokes by digestion and subsequent nutrients absorption reduces human or animal motility.

Liprotamase, a Non-porcine Enzyme Therapy, Restores Growth, and Normalizes Lipid Absorption in Pigs with Exocrine Pancreatic Insufficiency (EPI)

Purpose: Liprotamase is a biotechnology-derived, non-porcine product that contains 3 highly purified and stable enzymes: crystalline cross-linked lipase, crystalline protease, amorphous amylase. The particular enzymes selected for inclusion in liprotamase were based on their stability in the pH of the GI tract without enteric coating and their broad substrate specificity without the need for co-factors. Because of this purity and stability, we are able to administer liprotamase utilizing a novel rapid disintegrating formulation (LipRDT) that is easily suspended in liquid. The LipRDT was tested in a surgical model of exocrine pancreatic insufficiency, young EPI pigs that have total lack of pancreatic enzymes, resulting in poor nutrient absorption and arrested growth. Efficacy of LipRDT was measured by changes in fat digestion (%CFA) and fat and protein absorption (TG, NEFA and growth). Methods: A cross over randomized dose study was used to test two daily doses of liprotamase, high (HRDT) and low (LRDT), or placebo in 8 EPI pigs fed a high fat diet (HFD, ˜80g fat). Study included 1 control and 2 treatment periods with a washout in between, each 1 week. During the fifth week all pigs were placed on LRDT. As a comparator, 6 healthy pigs, same age and breed, were maintained on the HFD. Fat absorption was assessed by postprandial changes in TG and NEFA in blood samples collected before the morning meal and then in 1h intervals on last day of the control and treatment weeks. Results: One week of treatment enhanced fat and protein digestion and reversed impairment of growth with body mass increase of 6% and 8% on HRDT and LRDT. Treatment CFA was 84% and 79% with HRDT and LRDT respectively, from a basal value of 24%, vs. 92% in healthy pigs. LRDT efficacy was confirmed during extension week when CFA was 79%. Enhanced fat absorption with both tested doses was associated with normalization of blood TG and NEFA levels. Calculated AUC values for TG were the same as in healthy pigs: 4.1±1.9 and 4.5±1.6 for each RDT dose, vs. 4.1±0.7 in healthy. Similarly NEFA AUCs were 1.61±0.4 and 1.6±0.6 for each vs. 1.4±0.1 in healthy animals. Conclusion: Both tested doses of LipRDT improved digestion of fat and protein. As an important measure of nutrient absorption, LipRDT normalized postprandial TG and NEFA serum levels, and facilitated significant growth in EPI pigs. Postprandial TG and NEFA, as a measure of efficacy, are a viable endpoint to the difficult CFA that should be explored in human trials. Thus, our study demonstrates successful in vivo activity of a liquid suspension of a pancreatic enzyme replacement therapy designed for the treatment of EPI in infants, children, and patients with a G-tube. Disclosure: Dr Grujic employee of Alnara Pharmaceuticals; Dr Pierzynowski-Consultant. This research was supported by an industry grant from Alnara Pharmaceuticals.

An elemental diet fed, enteral or parenteral, does not support growth in young pigs with exocrine pancreatic insufficiency

Young individuals with exocrine pancreatic insufficiency (EPI) show growth reduction that can be reversed by dietary pancreatic enzyme supplementation. Here we investigated whether feeding an elemental diet could replace the growth-promoting effect of enzyme supplementation in EPI pigs. Weaned pigs with intact pancreas (control) or pancreatic duct-ligated (EPI pigs) were given a commercial pig feed, a fat-enriched diet, or an elemental diet, intragastrically and intravenously, with or without porcine pancreatin (Creon) supplementation for 1 week. Control pigs, irrespective of receiving pig feed or an elemental diet, increased their body weight by 13.4-20.1%, while EPI pigs showed negligible weight gain. Giving a fat-enriched diet did not improve growth of the EPI pigs. However, if the EPI pigs were supplemented with pancreatin in combination with fat-enriched feed or the elemental diet, i.v., their body weight increased by 16.6 %and 8.5%, respectively. Control pigs maintained normal growth, independently of the diet being given in polymeric or elemental form, while EPI pigs showed impaired growth when receiving the same diets without enzyme supplementation. Pancreatic juice and enzyme preparations, in addition to their digestive properties, also appear to affect nutrient assimilation and anabolism in young individuals.

The effectiveness of enzymatic replacement therapy measured by turbidimetry and the lipaemic index in exocrine pancreatic insufficient young, growing pigs, fed a high-fat diet

Conventionally, the management of exocrine pancreatic insufficiency (EPI) involves the consumption of a specific diet as well as the replacement of pancreatic enzymes, the effectiveness of which is usually measured by a classical method of blood analyses of non-esterified fatty acids (NEFA) and triglycerides (TG). Dietary supplementation with a pancreatic enzyme preparation (PEP), in conjunction with a high-fat diet, on growth performance, digestibility and absorption (analysed using turbidimetry) of dietary fat in pigs with EPI was investigated. EPI was developed by surgical ligation of the pancreatic duct of six male pigs, 6 weeks of age. The pigs were fed a high fat diet (twice daily). A PEP containing 1800 mg entero-coated pancreatin was included in the high fat meals. Blood, urine and faecal samples were collected. The urine and faeces were analysed for dry matter, crude protein and fat content. The lipaemic index and plasma lipid profiles were assessed. EPI completely stopped growth of the pigs. Treatment with PEP significantly increased (P<0.05) growth and body mass as well as the digestibility of dry matter and crude protein. PEP significantly improved the co-efficient of fat absorption, the lipaemic index (measured by turbidimetry methods) and caused significant changes in plasma nonesterified fatty acids and triglyceride concentrations. The short term enzymatic replacement therapy together with a high fat meal has immediate beneficial effects on diet digestibility and on the growth retardation observed in EPI pigs. The turbidimetry method used to measure lipaemic index is a reliable, quick and efficient technique in measuring plasma lipid profiles and thus a good tool for assessing fat absorption.

Pancreatic Enzyme Supplementation in Normal and Exocrine Pancreatic Insufficient Pigs

The effect of pancreatic enzyme preparation (Combizyme Forte granulate) on the growth rate of normal, sham-operated and pancreatic duct ligated growing pigs was studied in 2 trials. In the first trial 6 normal Yorkshire pigs were fed a pancreatic enzyme supplement and 6 other pigs from the same litter were fed without supplementation. The growth rate over a period of 4 weeks was not significantly greater in the supplemented group. In a second trial exocrine pancreatic insufficiency, without diabetic symptoms, was produced in 12 pigs (aged 1.5 months and weighing 14 kgs) by ligation of the main pancreatic duct. The growth rates over a period of 20 days after surgery were 240 g, 454 g and 483 g/day in ligated pigs, in ligated pigs supplemented with pancreatic enzyme and in sham-operated control pigs, respectively. The difference between the ligated groups was significant (p less than 0.01) indicating the efficiency of the pancreatic enzyme therapy.