Glucagon-like peptide-2 protects against TPN-induced intestinal hexose malabsorption in enterally refed piglets

Premature infants receiving chronic total parenteral nutrition (TPN) due to feeding intolerance develop intestinal atrophy and reduced nutrient absorption. Although providing the intestinal trophic hormone glucagon-like peptide-2 (GLP-2) during chronic TPN improves intestinal growth and morphology, it is uncertain whether GLP-2 enhances absorptive function. We placed catheters in the carotid artery, jugular and portal veins, duodenum, and a portal vein flow probe in piglets before providing either enteral formula (ENT), TPN or a coinfusion of TPN plus GLP-2 for 6 days. On postoperative day 7, all piglets were fed enterally and digestive functions were evaluated in vivo using dual infusion of enteral ((13)C) and intravenous ((2)H) glucose, in vitro by measuring mucosal lactase activity and rates of apical glucose transport, and by assessing the abundances of sodium glucose transporter-1 (SGLT-1) and glucose transporter-2 (GLUT2). Both ENT and GLP-2 pigs had larger intestine weights, longer villi, and higher lactose digestive capacity and in vivo net glucose and galactose absorption compared with TPN alone. These endpoints were similar in ENT and GLP-2 pigs except for a lower intestinal weight and net glucose absorption in GLP-2 compared with ENT pigs. The enhanced hexose absorption in GLP-2 compared with TPN pigs corresponded with higher lactose digestive and apical glucose transport capacities, increased abundance of SGLT-1, but not GLUT-2, and lower intestinal metabolism of [(13)C]glucose to [(13)C]lactate. Our findings indicate that GLP-2 treatment during chronic TPN maintains intestinal structure and lactose digestive and hexose absorptive capacities, reduces intestinal hexose metabolism, and may facilitate the transition to enteral feeding in TPN-fed infants.     

Total parenteral nutrition adversely affects gut barrier function in neonatal piglets

Sepsis is the most common morbidity in preterm infants, who often receive total parenteral nutrition (TPN). We hypothesized that gut barrier function is compromised in TPN-fed compared with enterally fed newborn piglets (ENT pigs). Colostrum-deprived newborn pigs were implanted with jugular venous and bladder catheters under general anesthesia. Pigs were either administered TPN (n = 15) or fed formula (ENT pigs, n = 15). After 6 days, pigs were gavaged a solution of mannitol, lactulose, and polyethylene glycol 4000 (PEG 4000) and urine was collected for 24 h. At 7 days, small bowel samples were assayed for myeloperoxidase activity, morphometry, and tight junction protein abundance. Intestinal contents and peripheral organ sites were cultured for bacteria. Urinary recovery (%dose) of mannitol (53 vs. 68) was lower, whereas that of lactulose (2.93 vs. 0.18) and PEG 4000 (12.78 vs. 0.96) were higher in TPN vs. ENT pigs, respectively (P < 0.05). Incidence of translocation was similar in TPN and ENT pigs. Myeloperoxidase activity was increased in TPN vs. ENT pigs in the jejunum (P < 0.001) and was weakly correlated with lactulose (R2 = 0.32) and PEG 4000 (R2 = 0.38) recovery. Goblet cell counts did not change, but intraepithelial lymphocyte numbers decreased with TPN. Only claudin-1 protein abundance was increased in the TPN group. We conclude that TPN is associated with impairment of neonatal gut barrier function as measured by permeability but not translocation.     

Glucagon-like peptide 2 has limited efficacy to increase nutrient absorption in fetal and preterm pigs

Exogenous glucagon-like peptide 2 (GLP-2) prevents intestinal atrophy and increases nutrient absorption in term newborn pigs receiving total parenteral nutrition (TPN). We tested the hypothesis that the immature intestine of fetuses and preterm neonates has a diminished nutrient absorption response to exogenous GLP-2. This was accomplished using catheterized fetal pigs infused for 6 days (87-91% of gestation) with GLP-2 (25 nmol.kg(-1).day(-1) iv; n = 7) or saline (n = 7), and cesarean-delivered preterm pigs (92% of gestation) that received TPN with GLP-2 (25 nmol.kg(-1).day(-1) iv; n = 8) or saline (n = 7) for 6 days after birth. Responses to GLP-2 were assessed by measuring intestinal dimensions, absorption of nutrients (glucose, leucine, lysine, proline) by intact tissues and brush border membrane vesicles, and abundance of sodium-glucose cotransporter mRNA. Infusion of GLP-2 increased circulating GLP-2 levels in fetuses, but did not increase intestinal mass or absorption of nutrients by intact tissues and brush border membrane vesicles, except for lysine. Administration of exogenous GLP-2 to preterm TPN-fed pigs similarly did not increase rates of nutrient absorption, yet nutrient absorption capacities of the entire small intestine tended to increase (+10-20%, P < 0.10) compared with TPN alone due to increased intestinal mass (+30%, P < 0.05). GLP-2 infusion did not increase sodium-glucose cotransporter-1 mRNA abundance in fetuses or postnatal preterm pigs. Hence, the efficacy of exogenous GLP-2 to improve nutrient absorption by the intestine of fetal and preterm pigs is limited compared with term pigs and more mature animals and humans.     

Glucagon-like peptide-2 induces intestinal adaptation in parenterally fed rats with short bowel syndrome

Glucagon-like peptide-2 (GLP-2) is an intestinal trophic enteroendocrine peptide that is associated with intestinal adaptation following resection. Herein, we investigate the effects of GLP-2 in a total parenteral nutrition (TPN)-supported model of experimental short bowel syndrome. Juvenile Sprague-Dawley rats underwent a 90% small intestinal resection and jugular catheter insertion. Rats were randomized to three groups: enteral diet and intravenous saline infusion, TPN only, or TPN + 10 microg.kg(-1).h(-1) GLP-2. Nutritional maintenance was isocaloric and isonitrogenous. After 7 days, intestinal permeability was assessed by quantifying the urinary recovery of gavaged carbohydrate probes. The following day, animals were euthanized, and intestinal tissue was processed for morphological and crypt cell proliferation (CCP) analysis, apoptosis (caspase-3), and expression of SGLT-1 and GLUT-5 transport proteins. TPN plus GLP-2 treatment resulted in increased bowel and body weight, villus height, intestinal mucosal surface area, CCP, and reduced intestinal permeability compared with the TPN alone animals (P < 0.05). GLP-2 treatment induced increases in serum GLP-2 levels and intestinal SGLT-1 expression (P < 0.01) compared with either TPN or enteral groups. No differences were seen in the villus apoptotic index between resection groups. Enterally fed resected animals had a significant decrease in crypt apoptotic indexes compared with nontreated animals. This study demonstrates that GLP-2 alone, without enteral feeding, stimulates indexes of intestinal adaptation. Secondly, villus hypertrophy associated with adaptation was predominantly due to an increase in CCP and not to changes in apoptotic rates. Further studies are warranted to establish the mechanisms of action and therapeutic potential of GLP-2.     

Qualitative and quantitative comparison of gut bacterial colonization in enterally and parenterally fed neonatal pigs

Total parenteral nutrition (TPN) has been associated with mucosal atrophy, impaired gut barrier function, and translocation of luminal bacteria with resultant sepsis in preterm human infants. Currently, we examined the effects of enteral (ENT) or TPN treatments on translocation events in neonatal pigs and on colonization and composition of microbiota in the neonatal gut. Newborn, colostrum-deprived pigs (<24 hours old) were fitted with intravenous catheters and were fed either ENT (n = 13) or TPN (n = 13) for 7 days. After 7 days of treatment, pigs were euthanized and samples were collected for bacterial culture from the blood, intestinal tract and organs. ENT pigs had increased numbers of bacterial genera isolated, higher concentrations of bacteria (CFU/g), and increased colonization of all segments of the intestinal tract compared to the TPN pigs. Translocation of bacteria from the intestinal tract to tissues or blood was similar (8 of 13) for both groups. The ENT group had 1/13 positive for Clostridium difficile toxin A whereas the TPN group had 5/13. We concluded that ENT favored increased bacterial concentrations comprised of more speciation in the gastrointestinal tract compared to TPN, and that TPN-treated piglets were at higher risk of colonization by toxin-expressing strains of C. difficile.     

Enteral bile acid treatment improves parenteral nutrition-related liver disease and intestinal mucosal atrophy in neonatal pigs

Total parenteral nutrition (TPN) is essential for patients with impaired gut function but leads to parenteral nutrition-associated liver disease (PNALD). TPN disrupts the normal enterohepatic circulation of bile acids, and we hypothesized that it would decrease intestinal expression of the newly described metabolic hormone fibroblast growth factor-19 (FGF19) and also glucagon-like peptides-1 and -2 (GLP-1 and GLP-2). We tested the effects of restoring bile acids by treating a neonatal piglet PNALD model with chenodeoxycholic acid (CDCA). Neonatal pigs received enteral feeding (EN), TPN, or TPN + CDCA for 14 days, and responses were assessed by serum markers, histology, and levels of key regulatory peptides. Cholestasis and steatosis were demonstrated in the TPN group relative to EN controls by elevated levels of serum total and direct bilirubin and also bile acids and liver triglyceride (TG) content. CDCA treatment improved direct bilirubin levels by almost fourfold compared with the TPN group and also normalized serum bile acids and liver TG. FGF19, GLP-1, and GLP-2 were decreased in plasma of the TPN group compared with the EN group but were all induced by CDCA treatment. Intestinal mucosal growth marked by weight and villus/crypt ratio was significantly reduced in the TPN group compared with the EN group, and CDCA treatment increased both parameters. These results suggest that decreased circulating FGF19 during TPN may contribute to PNALD. Moreover, we show that enteral CDCA not only resolves PNALD but acts as a potent intestinal trophic agent and secretagogue for GLP-2.     

Acute alcohol administration inhibits the refeeding response after starvation in rat skeletal muscle

This study determined whether an acute alcohol dose could inhibit the refeeding response in starved muscle. Rats starved for 24 h were pretreated with alcohol or saline before refeeding by intragastric or intravenous infusion of enteral diet (ENT), total parenteral nutrition (TPN), or saline. Refeeding by TPN or ENT stimulated increases in the fractional rate of protein synthesis (k(s)) in skeletal muscle. Alcohol prevented the increase in k(s) when refeeding occurred intragastrically (TPN or ENT) (P < 0.001) but not intravenously (TPN). Upon intragastric refeeding, alcohol inhibited the increase in both eukaryotic initiation factor 4E-binding protein-1 (4E-BP1) and p70 S6 kinase (p70(S6K)) phosphorylation in plantaris but caused only partial inhibition in soleus muscle (ENT only). When rats were refed intravenously, alcohol had no effect on the increased 4E-BP1 or p70(S6K) phosphorylation in either muscle. Plasma insulin levels were augmented by alcohol. Alcohol-related changes in plasma amino acid concentrations were similar irrespective of the route of feeding, whereas IGF-I levels showed differential changes. This is the first study to demonstrate that acute alcohol ingestion impedes the starved-to-fed response in skeletal muscle.     

Role of luminal nutrients and endogenous GLP-2 in intestinal adaptation to mid-small bowel resection

To elucidate the role of luminal nutrients and glucagon-like peptide-2 (GLP-2) in intestinal adaptation, rats were subjected to 70% midjejunoileal resection or ileal transection and were maintained with total parenteral nutrition (TPN) or oral feeding. TPN rats showed small bowel mucosal hyperplasia at 8 h through 7 days after resection, demonstrating that exogenous luminal nutrients are not essential for resection-induced adaptation when residual ileum and colon are present. Increased enterocyte proliferation was a stronger determinant of resection-induced mucosal growth in orally fed animals, whereas decreased apoptosis showed a greater effect in TPN animals. Resection induced significant transient increases in plasma bioactive GLP-2 during TPN, whereas resection induced sustained increases in plasma GLP-2 during oral feeding. Resection-induced adaptive growth in TPN and orally fed rats was associated with a significant positive correlation between increases in plasma bioactive GLP-2 and proglucagon mRNA expression in the colon of TPN rats and ileum of orally fed rats. These data support a significant role for endogenous GLP-2 in the adaptive response to mid-small bowel resection in both TPN and orally fed rats.     

Glucagon-like peptide-2 acutely increases proximal small intestinal blood flow in TPN-fed neonatal piglets

Glucagon-like peptide-2 (GLP-2) is a gut hormone that is secreted in response to enteral feeding and stimulates small intestinal mucosal growth. We have previously shown that GLP-2 infusion acutely increases portal venous blood flow in TPN-fed piglets. The aim of this study was to localize the vasoactive effect of GLP-2 within the gastrointestinal tissues and other visceral organs in TPN-fed piglets. Tissue blood flow rates were quantified using fluorescent microsphere deposition in anesthetized TPN-fed piglets given intravenous infusion of GLP-2 at either 500 pmol x kg(-1) x h(-1) (low GLP-2, n = 7 pigs) or 2,000 pmol x kg(-1) x h(-1) (high GLP-2, n = 8 pigs) for 2 h. Compared with baseline, the low and the high GLP-2 treatment significantly increased the blood flow rate in the duodenum (+77%) and jejunum (+40% and 80%), respectively, but blood flow to the distal small intestine and colon (-15%) was unchanged or slightly decreased. Baseline mucosal blood flow was five-fold higher than serosal blood flow; however, high GLP-2 treatment increased serosal (+140%) to a larger degree than mucosal blood flow (+73%). The high GLP-2 dose increased pancreatic flow (+34%) but decreased blood flow in the kidneys (-14%) and stomach (-12%), whereas the spleen and brain were unaffected. These findings suggest that the acute GLP-2-mediated stimulation of portal blood flow in TPN-fed piglets occurs principally via increased blood flow through the superior mesenteric artery to the proximal small intestine, a tissue region where the GLP-2R mRNA abundance and trophic GLP-2 effects are greatest.     

Enteral feeding induces diet-dependent mucosal dysfunction, bacterial proliferation, and necrotizing enterocolitis in preterm pigs on parenteral nutrition

Preterm neonates have an immature gut and metabolism and may benefit from total parenteral nutrition (TPN) before enteral food is introduced. Conversely, delayed enteral feeding may inhibit gut maturation and sensitize to necrotizing enterocolitis (NEC). Intestinal mass and NEC lesions were first recorded in preterm pigs fed enterally (porcine colostrum, bovine colostrum, or formula for 20-40 h), with or without a preceding 2- to 3-day TPN period (n = 435). Mucosal mass increased during TPN and further after enteral feeding to reach an intestinal mass similar to that in enterally fed pigs without TPN (+60-80% relative to birth). NEC developed only after enteral feeding but more often after a preceding TPN period for both sow's colostrum (26 vs. 5%) and formula (62 vs. 39%, both P < 0.001, n = 43-170). Further studies in 3-day-old TPN pigs fed enterally showed that formula feeding decreased villus height and nutrient digestive capacity and increased luminal lactic acid and NEC lesions, compared with colostrum (bovine or porcine, P < 0.05). Mucosal microbial diversity increased with enteral feeding, and Clostridium perfringens density was related to NEC severity. Formula feeding decreased plasma arginine, citrulline, ornithine, and tissue antioxidants, whereas tissue nitric oxide synthetase and gut permeability increased, relative to colostrum (all P < 0.05). In conclusion, enteral feeding is associated with gut dysfunction, microbial imbalance, and NEC in preterm pigs, especially in pigs fed formula after TPN. Conversely, colostrum milk diets improve gut maturation and NEC resistance in preterm pigs subjected to a few days of TPN after birth.     

Interleukin-10-independent anti-inflammatory actions of glucagon-like peptide 2

Glucagon-like peptide 2 (GLP-2) is an important intestinal growth factor with anti-inflammatory activity. We hypothesized that GLP-2 decreases mucosal inflammation and the associated increased epithelial proliferation by downregulation of Th1 cytokines attributable to reprogramming of lamina propria immune regulatory cells via an interleukin-10 (IL-10)-independent pathway. The effects of GLP-2 treatment were studied using the IL-10-deficient (IL-10(-/-)) mouse model of colitis. Wild-type and IL-10(-/-) mice received saline or GLP-2 (50 microg/kg sc) treatment for 5 days. GLP-2 treatment resulted in significant amelioration of animal weight loss and reduced intestinal inflammation as assessed by histopathology and myeloperoxidase levels compared with saline-treated animals. In colitis animals, GLP-2 treatment also reduced crypt cell proliferation and crypt cell apoptosis. Proinflammatory (IL-1beta, TNF-alpha, IFN-gamma,) cytokine protein levels were significantly reduced after GLP-2 treatment, whereas IL-4 was significantly increased and IL-6 production was unchanged. Fluorescence-activated cell sorting analysis of lamina propria cells demonstrated a decrease in the CD4(+) T cell population following GLP-2 treatment in colitic mice and an increase in CD11b(+)/F4/80(+) macrophages but no change in CD25(+)FoxP3 T cells or CD11c(+) dendritic cells. In colitis animals, intracellular cytokine analysis demonstrated that GLP-2 decreased lamina propria macrophage TNF-alpha production but increased IGF-1 production, whereas transforming growth factor-beta was unchanged. GLP-2-mediated reduction of crypt cell proliferation was associated with an increase in intestinal epithelial cell suppressor of cytokine signaling (SOCS)-3 expression and reduced STAT-3 signaling. This study shows that the anti-inflammatory effects of GLP-2 are IL-10 independent and that GLP-2 alters the mucosal response of inflamed intestinal epithelial cells and macrophages. In addition, the suggested mechanism of the reduction in inflammation-induced proliferation is attributable to GLP-2 activation of the SOCS-3 pathway, which antagonizes the IL-6-mediated increase in STAT-3 signaling.     

Intestinotrophic effects of exogenous IGF-I are not diminished in IGF binding protein-5 knockout mice

IGF binding protein-5 (IGFBP-5) modulates the availability of IGF-I to its receptor and potentiates the intestinotrophic action of IGF-I. Our aim was to test the hypothesis that stimulation of intestinal growth due to coinfusion of IGF-I with total parenteral nutrition (TPN) solution is dependent on increased expression of IGFBP-5 through conducting our studies in IGFBP-5 knockout (KO) mice. IGFBP-5 KO, heterozygote (HT) and wild type (WT) male and female mice were maintained with TPN or TPN plus coinfusion of IGF-I [recombinant human (rh)IGF-I; 2.5 mg x kg(-1) x day(-1)] for 5 days. The concentration of IGF-I in serum was 73% greater (P < 0.0001) in mice given TPN + IGF-I infusion compared with TPN alone. IGF-I attenuated the 2-3 g loss of body weight associated with TPN in WT mice, whereas KO and HT mice did not show improvement in body weight with IGF-I treatment. KO and HT mice had significantly greater levels of circulating IGF-I binding proteins (IGFBPs) compared with WT mice. Intestinal growth due to IGF-I was observed in all groups treated with IGF-I based on greater concentrations of protein and DNA in small intestine and colon and significantly greater crypt depth and muscularis thickness in jejunum. Jejunal expression of IGFBP-5 mRNA was greater in WT mice, whereas IGFBP-3 mRNA was greater in KO mice treated with IGF-I. In summary, the absence of the IGFBP-5 gene did not block the ability of IGF-I to stimulate intestinal growth, possibly because greater jejunal expression of IGFBP-3 compensates for the absence of IGFBP-5.     

Proteomic analysis of plasma from rats following total parenteral nutrition‐induced liver injury

Total parenteral nutrition (TPN) is provided as the primary nitrogen source to manage patients with intestinal failure who were not able to sustain themselves on enteral feeds. The most common complication of long‐term TPN use is hepatitis. A proteomic approach was used to identify proteins that are differentially expressed in the plasma of rats following TPN‐related acute liver injury. Six male rats were randomly assigned to either the saline infusion control group or the TPN infusion group. Our results demonstrate that TPN infusion in rats resulted in hepatic dysfunction and hepatocyte apoptosis. Five proteins that were differentially expressed between TPN infusion and normal rats were determined and validated in vivo. Fascinatingly, the proteomic differential displays, downregulated proteins included peroxiredoxin 2 (PRDX2), alpha‐1‐antiproteinase (A1AT), and fibrinogen gamma chain (FIBG), which were involved in oxidative stress, inflammatory respondence and cells apoptosis. After TPN infusion, two protein spots showed increased expression, namely, the glucagon receptor (GLR) protein and apolipoprotein A‐1 (APOA1), which may mediate the effects of TPN administration on glycogen and lipid metabolism. In this study, proteomic analysis suggested TPN‐related acute liver injury could be involved in limiting cellular protection mechanisms against oxidative stress‐induced apoptosis. On the basis of the results, we also give molecular evidences replying TPN‐related hepatitis.     

Enteral nutrients potentiate glucagon-like peptide-2 action and reduce dependence on parenteral nutrition in a rat model of human intestinal failure

Glucagon-like peptide-2 (GLP-2) is a nutrient-dependent, proglucagon-derived gut hormone that shows promise for the treatment of short bowel syndrome (SBS). Our objective was to investigate how combination GLP-2 + enteral nutrients (EN) affects intestinal adaption in a rat model that mimics severe human SBS and requires parenteral nutrition (PN). Male Sprague-Dawley rats were assigned to one of five groups and maintained with PN for 18 days: total parenteral nutrition (TPN) alone, TPN + GLP-2 (100 μg·kg(-1)·day(-1)), PN + EN + GLP-2(7 days), PN + EN + GLP-2(18 days), and a nonsurgical oral reference group. Animals underwent massive distal bowel resection followed by jejunocolic anastomosis and placement of jugular catheters. Starting on postoperative day 4, rats in the EN groups were allowed ad libitum access to EN. Groups provided PN + EN + GLP-2 had their rate of PN reduced by 0.25 ml/day starting on postoperative day 6. Groups provided PN + EN + GLP-2 demonstrated significantly greater body weight gain with similar energy intake and a safe 80% reduction in PN compared with TPN ± GLP-2. Groups provided PN + EN + GLP-2 for 7 or 18 days showed similar body weight gain, residual jejunal length, and digestive capacity. Groups provided PN + EN + GLP-2 showed increased jejunal GLP-2 receptor (GLP-2R), insulin-like growth factor-I (IGF-I), and IGF-binding protein-5 (IGFBP-5) expression. Treatment with TPN + GLP-2 demonstrated increased jejunal expression of epidermal growth factor. Cessation of GLP-2 after 7 days with continued EN sustained the majority of intestinal adaption and significantly increased expression of colonic proglucagon compared with PN + EN + GLP-2 for 18 days, and increased plasma GLP-2 concentrations compared with TPN alone. In summary, EN potentiate the intestinotrophic actions of GLP-2 by improving body weight gain allowing for a safe 80% reduction in PN with increased jejunal expression of GLP-2R, IGF-I, and IGFBP-5 following distal bowel resection in the rat.     

Insulin-like growth factor I and glucagon-like peptide-2 responses to fasting followed by controlled or ad libitum refeeding in rats

Luminal nutrients stimulate structural and functional regeneration in the intestine through mechanisms thought to involve insulin-like growth factor I (IGF-I) and glucagon-like peptide-2 (GLP-2). We investigated the relationship between IGF-I and GLP-2 responses and mucosal growth in rats fasted for 48 h and then refed for 2 or 4 days by continuous intravenous or intragastric infusion or ad libitum feeding. Fasting induced significant decreases in body weight, plasma concentrations of IGF-I and bioactive GLP-2, jejunal mucosal cellularity (mass, protein, DNA, and villus height), IGF-I mRNA, and ileal proglucagon mRNA. Plasma IGF-I concentration was restored to fed levels with 2 days of ad libitum refeeding but not with 4 days of intravenous or intragastric refeeding. Administration of an inhibitor of endogenous GLP-2 (rat GLP-2 3-33) during ad libitum refeeding partially attenuated mucosal growth and prevented the increase in plasma IGF-I to fed levels; however, plasma GLP-2 and jejunal IGF-I mRNA were restored to fed levels. Intragastric refeeding restored intestinal cellularity and functional capacity (sucrase activity and sodium-glucose transporter-1 expression) to fed levels, whereas intravenous refeeding had no effect. Intestinal regeneration after 4 days of intragastric or 2 days of ad libitum refeeding was positively associated with increases in plasma concentrations of GLP-2 and jejunal IGF-I mRNA. These data suggest that luminal nutrients stimulate intestinal growth, in part, by increased expression of both GLP-2 and IGF-I.     

The role of polyamines in glucagon-like peptide-2 prevention of TPN-induced gut hypoplasia

Total parenteral nutrition (TPN) of rats has been demonstrated to produce hypoplasia of gut mucosa, and to be associated with reduced immune response and elevated translocation of bacteria from gut to mesenteric lymph nodes, spleen and liver. Treatment of rats being maintained on TPN with the proglucagon fragment, glucagon-like peptide-2 (GLP-2), has been shown to totally prevent small intestine mucosal hypoplasia. In the present study, we found that depletion of polyamines with alpha-difluromethylornithine (DFMO) significantly reduced the efficacy of GLP-2 in preserving gut mucosa in rats maintained on TPN for 8 days. Co-infusion of GLP-2 with TPN prevented loss of protein and mucosa in duodenum, jejunum and ileum, but not in colon. Addition of DFMO to the infusate prevented the protective effects of GLP-2 in the duodenum and jejunum. In the jejunum, putrescine and spermidine were reduced in DFMO-treated rats, while the ileum exhibited reductions of these polyamines in rats infused with TPN or TPN plus GLP-2. DFMO infusion further reduced these polyamines in the ileum, while levels of spermine were increased. Concentrations of ornithine decarboxylase were elevated in jejunum of rats infused with TPN or TPN plus GLP-2, but were reduced significantly in DFMO-treated rats. These results suggest that normal levels of polyamines are necessary for the expression of GLP-2-induced hyperplasia. Differential effects of GLP-2 and DFMO across gut segments may relate to regional differences in proliferative and anti-apoptotic effects of the treatments.     

Modulation of intestinal protein synthesis and protease mRNA by luminal and systemic nutrients

Route of nutrient supply is important in regulation of intestinal protein metabolism, because total parenteral nutrition, compared with enteral feeding, leads to profound atrophy. Participation of the fractional rate of protein synthesis (Ks), their degradation in regulation of gut protein balance, and their possible modulation by specific nutrients are the focus of our work. We developed an in situ experimental system that allows controlled exposure of intestinal mucosa to nutrients systemically, luminally, or both. We examined the effects of systemic glucose and amino acid (AA) infusion in overnight-fasted piglets. Jejunal segments within each piglet were simultaneously, luminally perfused with solutions containing various AAs or glucose. Intravenous infusion of glucose increased mucosal Ks by 16% (P < 0.05), whereas intravenous infusion of AA had no effect on Ks. Systemic glucose infusion had no effect on mRNA levels for components of the ubiquitin-proteasome proteolytic pathway. However, levels of these mRNA were reduced by intravenous or luminal AA supply. This effect was greatest (-50%) when highest tissue concentrations of AAs were achieved by the simultaneous infusion of AA by both routes (P < 0.05). Our findings suggest that not only is the modulation of protein balance in the intestine in response to nutrients in part attributable to anabolic stimulation of protein synthesis initiated by the systemic appearance of glucose, but a fall in protein degradation is also a likely contributor. AAs appear to be a key factor required to reduce expression of genes connected with proteolysis.     

Loss of exocrine pancreatic stimulation during parenteral feeding suppresses digestive enzyme expression and induces Hsp70 expression

Luminal nutrients are essential for the growth and maintenance of digestive tissue including the pancreas and small intestinal mucosa. Long-term loss of luminal nutrients such as during animal hibernation has been shown to result in mucosal atrophy and a corresponding stress response characterized by the induction of heat shock protein (Hsp)70 expression. This study was conducted to determine if the loss of luminal nutrients during total parenteral nutrition (TPN) would result in atrophy of the exocrine pancreas and small intestinal mucosa as well as an induction of Hsp70 expression in rats. In experiment 1, the treatment groups included an orally fed control, a saline-infused surgical control, or TPN treatment for 7 days. In experiment 2, the treatment groups included an orally fed control and TPN alone or coinfused with varying doses of glucagon-like peptide (GLP)-2, a mucosal proliferation agent, for 7 days. In experiment 1, TPN resulted in a 40% reduction in pancreatic mass that was associated with a dramatic reduction in digestive enzyme expression, enhanced apoptosis, and a 200% increase in Hsp70 expression. Conversely, heat shock cognate 70, Hsp27, and Hsp60 expression was not changed in the pancreas. In experiment 2, TPN resulted in a 30% reduction in jejunal mucosa mass and a similar induction of Hsp70 expression. The inclusion of GLP-2 during TPN attenuated jejunal mucosal atrophy and inhibited Hsp70 expression, suggesting that Hsp70 induction is sensitive to cell growth. These data indicate that pancreatic and intestinal mucosal atrophy caused by a loss of luminal nutrient stimulation is accompanied by a compensatory response involving Hsp70.     

Glucagon-like peptide-2 increases sucrase-isomaltase but not caudal-related homeobox protein-2 gene expression

To determine the effect of glucagon-like peptide-2 (GLP-2) on sucrase-isomaltase and caudal-related homeobox protein-2 (Cdx-2) gene expression, male Wistar rats were divided into total parenteral nutrition (TPN)-fed and GLP-2-treated, TPN-fed groups. TPN was given via a jugular line, inserted under anesthesia, for 7 days. The treatment group received 40 microg/day of GLP-2 intravenously with the TPN diet. The small intestine and colon were weighed and measured. Tissue was obtained from the jejunum, terminal ileum, and midcolon. RNA analysis, morphometry, and microdissection were performed. The weight of the small intestine of GLP-2-treated rats was greater than that of TPN-fed rats (P < 0.001). GLP-2 increased the mean metaphase arrests/crypt in both the jejunum and ileum (P < 0.001). Ileal expression of sucrase-isomaltase was increased by 1. 6-fold (P < 0.05). Jejunal expression was increased by a similar amount, although not significantly (P = 0.08). There was no change in Cdx-2 gene expression. Thus GLP-2 can maintain small intestinal morphology and function, but effects on gene expression are not mediated by gross changes in the level of the mRNA for the homeobox protein Cdx-2.